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NBS SPECIAL PUBLICATION 619

U.S. DEPARTMENT OF COMMERCE/National Bureau of Standards

ASBESTOS STANDARDS:

Materials and Analytical Methods
NATIONAL BUREAU OF STANDARDS

The National Bureau of Standards' was established by an act of Congress on March 3, 1901

.

The Bureau's overall goal is to strengthen and advance the Nation's science and technology

and facilitate their effective application for public benefit. To this end, the Bureau conducts

research and provides: (1) a basis for the Nation's physical measurement system, (2) scientific and technological services for industry and government, (3) a technical basis for equity in trade, and (4) technical services to promote public safety. The Bureau's technical work is per- formed by the National Measurement Laboratory, the National Engineering Laboratory, and

the Institute for Computer Sciences and Technology.

THE NATIONAL MEASUREMENT LABORATORY provides the national system of

physical and chemical and materials measurement; coordinates the system with measurement

systems of other nations and furnishes essential services leading to accurate and uniform

physical and chemical measurement throughout the Nation's scientific community, industry,

and commerce; conducts materials research leading to improved methods of measurement,

standards, and data on the properties of materials needed by industry, commerce, educational

institutions, and Government; provides advisory and research services to other Government

agencies; develops, produces, and distributes Standard Reference Materials; and provides

calibration services. The Laboratory consists of the following centers:

Absolute Physical Quantities' — Radiation Research — Thermodynamics and

Molecular Science — Analytical Chemistry — Materials Science. THE NATIONAL ENGINEERING LABORATORY provides technology and technical ser- vices to the public and private sectors to address national needs and to solve national problems; conducts research in engineering and applied science in support of these efforts; builds and maintains competence in the necessary disciplines required to carry out this research and technical service; develops engineering data and measurement capabilities;

provides engineering measurement traceability services; develops test methods and proposes

engineering standards and code changes; develops and proposes new engineering practices;

and develops and improves mechanisms to transfer results of its research to the ultimate user. The Laboratory consists of the following centers:

Applied Mathematics — Electronics and Electrical Engineering — Mechanical

Engineering and Process Technology : — Building Technology — Fire Research —

Consumer Product Technology — Field Methods.

THE INSTITUTE FOR COMPUTER SCIENCES AND TECHNOLOGY conducts

research and provides scientific and technical services to aid Federal agencies in the selection, acquisition, application, and use of computer technology to improve effectiveness and

economy in Government operations in accordance with Public Law 89-306 (40 U.S.C. 759), relevant Executive Orders, and other directives; carries out this mission by managing the Federal Information Processing Standards Program, developing Federal ADP standards

guidelines, and managing Federal participation in ADP voluntary standardization activities; provides scientific and technological advisory services and assistance to Federal agencies; and

provides the technical foundation for computer-related policies of the Federal Government.

The Institute consists of the following centers:

Programming Science and Technology — Computer Systems Engineering.

'Headquarters and Laboratories at Gaithersburg, MD, unless otherwise noted; mailing address Washington, DC 20234.

; Some divisions within the center are located at Boulder, CO 80303.
ASBESTOS STANDARDS: "»F"

Edited by:

John Small and Eric Steel

Center for Analytical Chemistry

National Measurement Laboratory

National Bureau of Standards

Washington, DC 20234

Materials and Analytical Methods MAY 14 1982

Proceedings of the NBS/EPA Asbestos Standards

Workshop held at the National Bureau of Standards,

Gaithersburg, MD, October 1-3, 1980

Jointly sponsored by: U.S. Environmental Protection Agency

Research Triangle Park, NC 2771 I and the National Bureau of Standards Washington. DC 20234

U.S. Department of Commerce, Malcolm Baldrige, Secretary

NATIONAL BUREAU OF STANDARDS, Ernest Ambler, Director

Issued March 1982
Library of Congress Catalog Card Number: 82-600506

National Bureau of Standards Special Publication 619

Nat. Bur. Stand. (U.S.), Spec. Publ. 619, 220 pages (Mar. 1982) CODEN: XNBSAV

U.S. GOVERNMENT PRINTING OFFICE WASHINGTON: 1982

For sale by the Superintendent of Documents, U.S. Government Printing Office, Washington, D.C. 20402

Price $7.00

(Add 25 percent for other than U.S. mailing)
FOREWORD

The Center for Analytical Chemistry of the National Measurement Laboratory, National Bureau of Standards, seeks to develop new techniques of chemical analysis and to constantly improve existing

analysis methods. Part of the mission of NBS is to disseminate

knowledge in the scientific and technical community. To aid in

achieving this objective, the Center for Analytical Chemistry has

sponsored a series of workshops on various topics in analytical

chemistry. The workshop topics are chosen to fulfill current needs

for detailed discussions on well-defined subjects in a wide variety

of specialized areas of interest. The objective of these workshops

is to bring together specialists from throughout the world to concentrate intensively on a particular subject in order to advance

the state-of-the-art. It is often very difficult to achieve this

goal at large international meetings where the size and diversity

of topics presented often limit detailed discussion of specialist

subjects. Past topics of these workshops and the published

proceedings include: Quantitative Electron Probe Microanalysis

(NBS Special Publication 298), Aerosol Measurements (NBS Special

Publication 412), Secondary Ion Mass Spectrometry (NBS Special

Publication 427), Use of Monte Carlo Calculations in Electron

Probe Microanalysis and Scanning Electron Microscopy (NBS Special

Publication 460), Characterization of Particles (NBS Special

Publication 533), and Energy Dispersive X-ray Spectrometry (NBS

Special Publication 604). These proceedings are available from

the Superintendent of Documents, Government Printing Office,

Washington, D. C. 20402. Further information on the workshops can

be obtained by writing to the Center Office, Center for Analytical

Chemistry, National Bureau of Standards, Washington, D. C. 20234.

This volume contains the proceedings of a Workshop on Asbestos

Standards. The three-day meeting involved participants from the

United States and Canada. The workshop format consisted of invited

talks and contributed papers on each topic followed by extensive

discussions.

There is an important need in the area of asbestos analysis

for physical standards as well as standard analytical methods.

This publication provides a detailed view of the state-of-the-art

in asbestos standardization and analysis. The papers should be of

interest to workers involved in programs associated with all

aspects of asbestos measurement.

i i i
PREFACE

Asbestos has been an important manufacturing and industrial

product since the beginning of the industrial revolution resulting

in its use in a variety of goods and materials. It was not until

the mid 1900s that asbestos was recognized as a health hazard and

by that time its widespread use had created a detectable global

background level. As a result of its potential adverse health

effects, the regulation and monitoring of asbestos has increased

dramatically during the past several years.

In 1976, the National Bureau of Standards (NBS) at the request

of the Occupational Safety and Health Administration (OSHA)

analyzed 80 industrial talc samples for asbestos. This analysis

marked the initial involvement of the Center for Analytical

Chemistry at NBS in the analysis for and characterization of

asbestos.

As a direct result of this program, NBS and OSHA jointly

sponsored a workshop on asbestos. The workshop was purposefully

broad in scope, enabl ing NBS and OSHA to evaluate information con- cerning the current state-of-the-art in asbestos definitions and

measurement methods. The proceedings of the workshop were published as NBS Special Publication 506.

Since this initial work, the emphasis of NBS involvement with

asbestos has appropriately shifted from routine analysis to a joint

program with the Environmental Protection Agency for the development of quality control specimens, reference standards, and refinement of analysis methodology. As part of this program, a joint

NBS-EPA workshop on Asbestos Standards was held in October 1980.

This workshop was designed to bring together representatives from

private and governmental organizations which are actively involved

in asbestos standards preparation and development of analysis

methods.

The editors gratefully acknowledge the excellent work of Joy

Shoemaker and the members of the Text-Editing Facility of the

Center for Analytical Chemistry in preparing this manuscript.

J. A. Small

E. B. Steel

Gas & Particulate Science Division

Center for Analytical Chemistry

iy
ABSTRACT

This publication contains the invited papers which were presented at a workshop on asbestos standards jointly sponsored by

the Environmental Protection Agency and the National Bureau of

Standards and held on October 1-3, 1980. The workshop was divided

into five sections: (1) Bulk Materials for Preparation of Asbestos

Standards. .. .This section includes descriptions of natural and

synthetic materials which have a potential use as standards for

asbestos analysis. In addition, it also includes a description of

the NBS Standard Reference Materials Program. (2) Standards Preparation. ... The electron-microscopy preparation procedures for

standards mimicking airborne and waterborne asbestos samples are described. (3) Asbestos Analysis for Standards Certification....

This section describes analytical procedures and problems asso- ciated with the intra- and i nterl aboratory analyses of asbestos

standards. (4) Error Analysis and Stati sties. ... Thi s section

describes the statistical considerations which are involved in

asbestos standards preparation and analysis. (5) EPA Provisional

Method. ... Current developments and selected problems with the EPA

Provisional Method for Electron Microscope Measurement of Airborne

Asbestos Concentrations are discussed.

The papers include general reviews on each of the subjects as well as specific papers detailing current research efforts.

Key Words: Asbestos standards; asbestos statistics; electron

microscopy; fibrous minerals.

V
TABLE OF CONTENTS

PAGE

FOREWORD iii

PREFACE iv

ABSTRACT v

INTRODUCTION

QUALITY ASSURANCE FOR AIRBORNE ASBESTOS MEASUREMENTS 1 Michael E. Beard

SESSION I. BULK MATERIALS FOR PREPARATION OF ASBESTOS STANDARDS

ASBESTOS REFERENCE MATERIALS: SOURCES AND CHARACTERIZATION 5 Jean L. Graf, Ronald G. Draftz, and Janet C. Haartz

GLASS AS A MATERIAL FOR ASBESTOS STANDARDS 21 David J. Cronin, Douglas H. Blackburn, and Wolfgang K. Haller

THE NBS PROGRAM FOR STANDARD REFERENCE MATERIALS 29

R. Keith Kirby

IDENTIFICATION AND QUANTIFICATION OF ASBESTOS IN CONSTRUCTION MATERIALS

USING POLARIZED LIGHT MICROSCOPY: THE NEED FOR STANDARDS 34

R. L. Virta, K. B. Shedd, and W. J. Campbell

PREPARATION OF ASBESTOS "STANDARDS" FOR METHODS VERIFICATION AND LABORATORY EVALUATION 44

D. E. Lentzen, E. P. Brantly, K. W. Gold, and L. E. Myers

SESSION II. STANDARDS PREPARATION

SAMPLE PREPARATION FOR QUANTITATIVE ELECTRON MICROSCOPE ANALYSIS OF ASBESTOS FIBER CONCENTRATIONS IN AIR 53

Philip M. Cook and David R. Marklund

DEVELOPMENT OF AN ASBESTOS REFERENCE SUSPENSION 68

Richard S. Feldman

PREPARATION OF AIRBORNE ASBESTOS STANDARDS 77 David R. Jones and George Yamate

CONCENTRATION AND SEPARATION OF CHRYSOTILE BY TWO-PHASE LIQUID SEPARATION. ... 85

Carl W. Melton, Sandra J. Anderson, Carolyn F. Dye,

W. Eugene Chase, and Charles H. Anderson

SESSION III. ASBESTOS ANALYSIS FOR STANDARD CERTIFICATION

ANALYTICAL PROCEDURES AND STANDARDIZATION FOR ASBESTOS FIBER COUNTING

IN AIR, WATER, AND SOLID SAMPLES 91 Eric J. Chatfield

DEVELOPMENT OF A RAPID SURVEY TECHNIQUE FOR THE DETECTION OF ASBESTOS

FIBERS 108

Peter Riis and Eric J. Chatfield

vi
Table of Contents Continued PAGE

INTERLABORATORY MEASUREMENTS OF THE CHRYSOTILE ASBESTOS FIBER AND MASS

CONCENTRATIONS IN WATER SAMPLES 121

K. S. Chopra, D. Beaman, and P. Cook

CONSIDERATIONS IN THE ANALYSIS AND DEFINITION OF ASBESTOS USING ELECTRON

MICROSCOPY 132

R. J. Lee, J. F. Kelly, and J. S. Walker

COMMENTS ON THE ACHIEVABILITY OF A VALID ASBESTOS STANDARD FOR TEM COUNTING. . . 138

I. M. Stewart

SESSION IV. ERROR ANALYSIS AND STATISTICS

MEMBRANE FILTER METHOD: STATISTICAL CONSIDERATIONS 145

Gerald R. Chase

SIMULATION OF THE EPA PROVISIONAL METHOD FOR AIRBORNE ASBESTOS

CONCENTRATIONS 154

Terence Fitz-Simons and Michael E. Beard

ANALYTICAL ERRORS IN ASBESTOS ANALYSIS BY ANALYTICAL ELECTRON MICROSCOPY .... 162

Eric B. Steel, John A. Small, and Patrick Sheridan

STATISTICAL CONSIDERATIONS IN THE PREPARATION OF CHRYSOTILE FILTER

REFERENCE MATERIALS: FILTER HOMOGENEITY 169

Stephan Leigh, Eric Steel, John Small, Patrick Sheridan, and

James Filliben

SESSION V. EPA PROVISIONAL METHOD

REFINEMENTS IN THE EPA PROVISIONAL METHODOLOGY 183

George Yamate and Michael E. Beard

FIBER IDENTIFICATION AND BLANK CONTAMINATION PROBLEMS IN THE EPA PROVISIONAL METHOD FOR ASBESTOS ANALYSIS 190

Steven Ring and Robert J. Suchanek

MSHA STANDARD METHOD FOR FIBER IDENTIFICATION BY ELECTRON MICROSCOPY 207

R. L. Clark

LIST OF ATTENDEES 211

Disclaimer: In order to adequately describe materials and experimental procedures, it was occasionally necessary to identify commercial products by manufacturer's name or label. In no instance does such identification imply endorsement by the National Bureau of Standards

nor does it imply that the particular products or equipment are necessarily the best available for that purpose.

vi i

National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gai thersburg , Maryland, October 1-3, 1980. (Issued March

1982)

QUALITY ASSURANCE FOR AIRBORNE ASBESTOS MEASUREMENTS

Michael E. Beard

Methods Standardization Branch, Quality Assurance Division

Environmental Monitoring Systems Laboratory

U.S. Environmental Protection Agency

Research Triangle Park, North Carolina 27711

Abstract

An EPA program designed to provide quality assurance foV

measurement of airborne asbestos fibers is outlined. The program

provides for the development of (1) a standardized measurement protocol

and (2) a characterized reference material suitable for performance

evaluation. A provisional methodology manual (EPA Report 600/2-77-178)

describing transmission electron microscopic identification of asbestos

fibers is being evaluated under contract. Critical subroutines

involving sampling, sample preparation, and analysis are being

investigated and the resulting information will be used to optimize the

current protocol. The resulting protocol will be subjected to a multi laboratory collaborative test designed to determine precision and

accuracy of the method. A concurrent EPA-NBS agreement is designed to produce reference materials for use in performance evaluation of

electron microscopic analysis of asbestos fibers. The material s wi 1 1 be

in the form of prepared grid specimens and coated sample filters. These

standards will allow identification of variabilities due to sample

preparation and counting. The program is scheduled for completion in

late 1981.

1. Introduction

The hazards associated with the widespread occurrence and use of asbestos fibers have

become a matter of increasing concern in recent years. The association between asbestos

workers and respiratory diseases such as asbestosis and lung cancer has resulted in air

quality standards for the work environment. Concern has also been raised for others who

breathe ambient levels of this hazardous material. It is in the public interest that any

potential health hazard due to breathing airborne asbestos fibers be assessed.

Measurement of airborne asbestos presents a unique analytical problem. These fibers

may range in size from a hundred or more micrometers to submi crometer lengths and submicrometer diameters. Reliable identification of the fibers requires the use of transmission

electron microscopy (TEM) which is costly, tedious, and time consuming. A recent study

focused on asbestos emissions from roadways surfaced with crushed stone obtained from a quarry containing small amounts of asbestos [l] 1

. Particulate samples analyzed for

asbestos by several laboratories using their own analytical protocol gave results on split

samples which differed by several orders of magnitude (Table 1). The need for a standardized measurement technique is obvious.

This paper reviews the Environmental Monitoring Systems Laboratory (EMSL), Research

Triangle Park, North Carolina (RTP) program to standardize and provide quality assurance for

airborne asbestos measurements. Detailed findings of each phase of the program will be presented by the respective investigators during this workshop. The cost of asbestos measurements and the cost and impact of controlling emissions of this hazardous material demand a reliable standardized measurement technique and a rigorous quality assurance program.

Figures in brackets indicate the literature references at the end of this paper.

1
Table 1. Ambient Air Asbestos Fiber Counts Measured by Eight Laboratories for Site 1 Adjacent to an Unpaved Graveled Road, Millions of Fibers/m3 of Ambient Air. ___________ Laboratory Designation

Relative Sample

Level Number A B c D E F G H

Low 1 BDL

3 BDL 0. 141 0. 145 BDL BDL 2. 16 0. 0334

2 BDL BDL 0, 217 0. 00739 0. 00874 BDL 1. 26 0. 199

3 BDL BDL 0, 160 0. 264 0. 00336 0. 00382 0. 180 1

.

03

Medi urn 11 0. 00805 0, 00064 0. 375 0. 135 0. 178 0, 0253 2. 74 6. 19 12 0. 0127 0, 0262 0. 297 1

,

48 0. 510 0. 0727 6. 61 17. 1 13 0. 00211 0. 0303 0. 253 1. 04 0. 273 0. 0422 3. 37 4. 29

High 21 0. 00521 0. 0417 0. 608 1

.

60 0. 201 28. 2 13. 7 22 0. 00159 0. 0333 1

.

15 0. 339 0. 100 0. 160 16. 8 54. 9 23 0, 00845 0, 0691 1. 20 0, 269 0. 282 9. 18 30. 7

Below detectable limits.

Data from EPA internal report: "Montgomery County Asbestos Study," October 1977.

1.1 Development of a Standardized Measurement Technique

There are several key elements which apply to the standardization of any environmental

measurement technique. First, a detailed protocol for accomplishing the measurement is

prepared. This protocol includes detailed descriptions of sampling, sample preparation,

analysis, calibrations, and data reporting procedures. The protocol may be based on information obtained from the literature, consultation with investigators, or limited laboratory

investigation. The protocol may be revised during subsequent evaluations as new information

is obtained which requires modification of the procedure in order to achieve more reliable

results.

Secondly, the protocol is subjected to a laboratory evaluation designed to determine

the performance of the procedure under controlled conditions. Critical subroutines may be

identified using multifactorial block design experiments (ruggedness tests). The optimum

values for these critical parameters are then determined and the protocol is revised

accordingly.

Thirdly, the protocol is subjected to various levels of field testing. This phase may

include single or multi laboratory measurements. Any critical parameters identified in this

phase of the study may be further evaluated in the laboratory and/or field and the information used to make revisions in the protocol.

Finally, the protocol may be subjected to a multi laboratory collaborative test designed

to determine the precision, accuracy, and overall utility of the method under monitoring

conditions. Performance of the procedure in monitoring networks can be measured by periodic

audits using various reference standards or comparisons.

A standardization program for measurement of airborne asbestos has been in progress at

EPA/RTP for the past several years. A provisional methodology manual describing the mea- surement of airborne asbestos was prepared under an EPA contract with I IT Research Institute

in 1977 [2]. The manual was developed through consultation with several investigators and

extensive laboratory evaluation [3]. In the procedure (figure 1), airborne particulate is

collected on polycarbonate membrane filters. A portion of the filter is carbon coated and

transferred to an electron microscope grid by means of a Jaffe wick. The grid is examined

by TEM. Fibers with aspect ratios of 3:1 or greater are identified by morphology and their

2
Sample

Air

Prepare

Sample

Ashing

Dilution

and

Redeposition

Prepare

Grid

Examine

Grid

Estimate

Concentration

Figure 1. Flow chart of EM procedure for estimating size distribution

and concentration of airborne asbestos.

crystal structure is determined by selected area electron diffraction (SAED). Chemical

composition may also be determined using energy dispersive x-ray fluorescence analysis

(EDXRF). Asbestos fibers are counted, their length and width determined, and a mass concentration is calculated using a density-volume relationship. Data reported includes

fiber count, mass, and particle size distribution.

The method was subjected to a mul ti 1 aboratory round-robin test which showed a precision

of 0.48 (spread between 95 percent confidence limits to mean) for fiber number concentration

and 0.40 for mass concentrations. While these results were indeed promising, some problem

areas were identified in the evaluation of the method which required further study. Subroutines involving the effects of sampling with polycarbonate vs. cellulose ester filters,

sampling face velocity, collection efficiency, ashing, dispersing and ultrasonic treatment

of the sample, and fiber bundle counting procedures are currently under investigation by

IITRI. Information obtained in the current investigation will be used to optimize the

method and, if appropriate, a collaborative study will be conducted. The investigation is scheduled to be completed by 1982.

3
1.2 Performance Evaluation of Asbestos Measurements

One means of evaluating the performance of a monitoring technique under actual use is to provide for a comparison between results obtained using the procedure and its associated

apparatus and those obtained using appropriate reference standards. For example, the flow

rate of a sampler used to collect airborne particulate for asbestos analysis may be calibrated by using a gas meter, calibrated orifice and manometer, or other flow measurement

device. An independent flow measurement device referenced to a primary flow measurement can then be used to obtain a comparative flow measurement. A criterion for acceptance of the user calibration can be established using established performance characteristics of the

apparatus. Performance audit devices for flow are in common use by EPA.

A reference material for determining the performance of the sample preparation and

analysis phases of the provisional method is currently being developed through an interagency agreement between EPA and the National Bureau of Standards. The material will

consist of a filter material having a deposit of characterized asbestos fibers in an urban

air particulate matrix. Electron microscope grid specimens prepared from the filter

material will be available on an interim basis until the development of the filter material

is completed. These materials will allow performance testing for the sample preparation

technique and the fiber identification and counting techniques. The reference materials

will contain several concentration levels of both chrysotile and amphibole asbestos fibers

in an urban air particulate matrix. The materials are scheduled to be available by 1982.

Other standards are available to the microscopist, such as latex spheres or carbon

grating replicas used to calibrate EM magnification, and gold layered grid specimens used

as internal standards for diffraction pattern measurements.

2. Summary

The effort to provide quality assurance for airborne asbestos measurements will be

greatly promoted with the development of (1) a standardized measurement protocol and (2) an asbestos reference material. Monitoring programs for airborne asbestos can be undertaken

and reliable data can be obtained. This information can be used to assess the need for

establishing any air quality standards required to protect the public health.

Progress reports from these projects are receiving review by several investigators representing federal and state government, private research institutes, and industry. Comments

and suggestions on the program are invited and welcome. Please address same to the Methods

Standardization Branch, Attn: Michael E. Beard, Quality Assurance Division (MD-77),

Environmental Monitoring Systems Laboratory, U.S. Environmental Protection Agency, Research

Triangle Park, North Carolina 27711

References

[1] Montgomery County Asbestos Study, EPA internal report, U.S. Environmental Protection

Agency, Environmental Monitoring Systems Laboratory, Environmental Monitoring Division,

Research Triangle Part, North Carolina 27711, October 1977.

[2] Electron Microscope Measurement of Airborne Asbestos Concentrations: A Provisional

Methodology Manual, EPA Report No. 600/2-77-178, Environmental Sciences Research Laboratory, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina

27711 , Revised June 1978.

[3] Evaluating and Optimizing Electron Microscope Methods for Characterizing Airborne

Asbestos, EPA Report No. 600/2-78-038, Environmental Sciences Research Laboratory,

U.S. Environmental Protection Agency, Research Triangle Park, North Carolina 27711,

June 1978.

4
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gaithersburg , Maryland, October 1-3, 1980. (Issued March

1982)

ASBESTOS REFERENCE MATERIALS: SOURCES AND CHARACTERIZATION

Jean L. Graf and Ronald G. Draftz

I IT Research Institute

10 West 35th Street

Chicago, Illinois 60616

and

Janet C. Haartz

Division of Biomedical and Behavioral Sciences

National Institute of Occupational Safety and Health

Cincinnati , Ohio 45226

Abstract

One of the keys to the successful development of any analytical

method is the availability of wel 1 -characteri zed reference materials.

The need for asbestos standards is especially acute since research

efforts to develop standardized methods of asbestos analysis are currently in progress even though there is an absence of high purity,

well-characterized asbestos reference materials. Current knowledge on sources for asbestos reference samples and the extent of their

characterization are summarized along with a list of recommendations for

the development of additional reference samples based on size and purity.

1. Introduction

The primary uses of standards or secondary reference materials are for instrument calibration, quality assurance, and methods development. In the case of known or potentially

hazardous materials such as asbestos, a primary need for standard reference materials is

also for bioassay studies.

Standard reference materials are used in instrument calibration to either check instrument response for the substance of interest or in preparation of calibration curves. Standard reference materials are essential for quality assurance programs in analytical

labs: "blind" samples prepared from known standards by the laboratory quality assurance officer must be analyzed routinely, or standard addition methods must be routinely incorporated in sample batches to verify a method's accuracy and reproducibility. No development

of analytical methods can logically proceed without standard reference materials.

Asbestos poses many problems to the conventional approaches of standard reference

materials production, the most significant of which is that asbestos is not simply one precise material. There are six naturally occurring silicate minerals that possess the

physical and chemical properties requisite for the commercial classification as asbestos.

Although only three of the asbestos minerals types - chrysotile, fibrous grunerite

(amosite), and crocidolite - are mined and processed in significant quantities, the other

three asbestos types - fibrous habits of the amphiboles anthophyl 1 ite, tremolite, and

actinolite - have either been used commercially in small quantities or can exist in non- asbestos commercial materials such as talc. Therefore, unless analysis methods are to be

restricted solely to one type of asbestos, standard reference materials of all six of the

asbestos types must be prepared.

5
The task of preparing suitable, broad application asbestos standard reference materials

is further complicated by the fact that one asbestos type can occur with widely varying

chemical and physical properties. In addition, the amphibole asbestos minerals also occur

in nonfibrous, but elongated crystal habits that are not readily distinguished from the

fibrous habit on a microscopic scale. Therefore, more than one standard reference material

of each asbestos type may have to be prepared, and the non-fibrous forms of the amphibole

asbestos minerals may also have to be prepared as standard reference materials to insure

that developed analytical methodologies are capable of distinguishing between the asbestos

and non-asbestos varieties of the mineral.

The desired properties of standard reference asbestos materials include high and

documented purity; availability, or more properly, accessibility to the research and analytical communities; and availability in forms amenable to the variety of uses to which they

will be put. These properties are not independent of each other - and therefore must be

considered very carefully as the plans for production of asbestos standard reference

materials are formulated.

2. Existing Needs for Asbestos Standard Reference Materials

Table 1 lists the most common analytical techniques currently employed in the detection

and quantitation of asbestos and the types of samples that are typically analyzed by the

techniques. There are, in fact, no fully developed, certified, so-called "standard" analysis methods for asbestos, 1 for the most part, because standard reference materials for

methods development and methods testing have not been available.

Table 1. Current Needs for Asbestos Standard Reference Materials.

Technique

Polarized Light Microscopy [l] 2

Type of Sample

bulk powders

Materials to be Analyzed

insulations, quarried rocks,

building materials

X-ray Diffraction [1] bulk powders insulations, quarried rocks,

building materials, talcs

X-ray Diffraction [2] thin film samples airborne fibers, liquidborne fibers, thin films

prepared from bulk powders

Electron Microscopy [3] thin film samples airborne fibers, liquidborne fibers, thin films

prepared from bulk powders

Some of the existing and proposed federal regulations pertaining to asbestos have

created urgent needs for asbestos standard reference materials in the analytical community

by either establishing limits for asbestos concentration (in the environment or in commercial products) or by specifying that a certain analytical technique be employed. For

example, the recently proposed rules of the U.S. Environmental Protection Agency [5] and the

U.S. Department of Education [6] for detection and control of asbestos containing building

materials in educational institutions require employment of a tentative polarized light

microscopy or x-ray diffraction techniques [1] for the analyses of the suspect building

materials. Since none of these tentative asbestos analysis methods has been sufficiently

tested to determine precision, accuracy, reliability and interferences, those proposed rules

may encounter stiff opposition, or if passed, may be nonenf orceabl e because adequate numbers

*The possible exception is the NIOSH P&CAM 239, Asbestos Fibers in Air, in which fiber

concentrations in the asbestos workplace environment are quantitated by a phase contrast

optical microscopy counting method. It should be noted that all particles meeting the

morphology and aspect ratio criterion of fibers, rather than only asbestos fibers, are counted in this technique.

2 Figures in brackets indicate the literature references at the end of this paper.

6
of qualified laboratories will not be available to provide the required analyses. A major

stumbling block to the testing and development of the methodologies has again been a lack

of asbestos reference materials. In fact, the tentative x-ray diffraction (XRD) methodologies will be useable only by a few government laboratories because both the bulk powder

and thin film XRD analysis methods require the preparation of a standard cal ibration curve . Logically, if standard reference asbestos materials are not used by commercial

analytical laboratories for instrument calibration, the XRD data produced cannot be con- sidered rel iable.

It should be obvious from Table 1 that a wide variety of asbestos standard reference

materials are required by the analytical community. In addition to standard reference

materials of each of the various minerals that fall under the commercial category, asbestos, each type of asbestos should be prepared in several different forms. For example,

analyses for determination of asbestos in bulk materials such as insulation by polarized

light microscopy will require preparation of standards with relatively large sized fibers,

while asbestos standards to be analyzed by x-ray diffraction or electron microscopy methods

must be reduced to very much smaller fiber sizes. Consideration should also be given to the preparation of standard reference materials of asbestos in the various matrix types

expected to be encountered in real world samples, since the sample preparation procedures

for PLM, EM, and XRD analyses are as important factors as the actual instrumental analysis

procedures are in ensuring accurate and reliable data. Therefore, pure asbestos bulk

powder standards, asbestos plus matrix bulk powder standards, pure asbestos thin film

(i.e., filter collected samples) standards, and asbestos plus matrix thin film standards

are the types of standard reference materials that are needed by analytical laboratories.

3. Existing Asbestos Samples

Table 2 lists some specific and generic sources of uncharacteri zed , processed, and

unprocessed asbestos samples. Asbestos samples of unknown purity, particle size, precise

chemical composition, etc. , are useful for observation of general analytical responses of

asbestos materials. For example, an uncharacteri zed asbestos sample can be used by an electron or optical microscopist for familiarization of the analyst with physical (e.g.,

refractive indices, birefringence for the optical microscopist), and morphological properties. However, such uncharacterized samples are of little use in the development of

quantitative analysis procedures, particularly those techniques that rely on some type of

asbestos concentration vs. instrument response calibration curve in the quantitation of

unknown samples.

Table 2. Sources of Uncharacterized Asbestos Materials.

U.S. Government Agencies

GSA Stockpiles

Bureau of Mines

State Geological Surveys

Public and Private Museums

Smithsonian

Mineral Dealers

Wards Natural Science Establishment

Minerals Unlimited

Mackinaw Geological Supply

Commercial

Johns-Manvi 1 1

Union Carbide

X-ray and Optical Microscope Analysis Supplies Dealers

Spex

Cargi 1 le Somar Labs.

Chemical Laboratories

Gooch Crucible Fibers

7
The GSA stockpiles currently maintain supplies of chrysotile, fibrous grunerite, and

crocidolite asbestos. A letter of authorization is usually required from another government

agency to obtain samples from the GSA stockpiles and other public institutions such as the

Smithsonian Museum. Therefore, non-government laboratories seeking samples of asbestos from

these government agencies must require the asbestos samples for completion of an existing

government sponsored contract.

Numerous private and commercial mineral dealers operate throughout the United States.

A brief search through the geology and mineralogy oriented professional journals will

provide many more dealers than those in Table 2 that can be contacted for asbestos mineral

specimens. Mineral dealers are especially useful as sources of the nonfibrous counterparts of the amphibole asbestos types. The three dealers listed have been especially

helpful to IITRI in obtaining asbestos mineral specimens in the past; the dealers are each

located in different areas of the United States that are rich in asbestos minerals or the

nonfibrous counterparts.

The major U.S. asbestos industrial companies (e.g., Johns-Manvi 1 1 e and Union Carbide)

may also provide small samples of uncharacteri zed or partially characterized asbestos.

Supply dealers that specialize in accessories, and chemicals for x-ray and optical

(polarized light) microscopy analysis techniques also sell powdered mineral sample sets that

include the asbestos minerals. Caution is advised in the interpretation of analysis data

from the powdered amphiboles since it cannot be determined with certainty if the minerals were of a true fibrous habit in the original, unground specimen and were therefore

asbestos.

One of the more interesting sources of amphibole asbestos (usually anthophyl 1 ite) has

been discovered to be the dusty back shelves of an old chemistry laboratory. The fibrous

filter bed material sold for Gooch crucibles was amphibole asbestos. Although laboratory

equipment supply catalogues as late as 1979 still listed amphibole asbestos for Gooch

crucibles as an available item, three major suppliers recently contacted (Fisher Scientific,

J. T. Baker Company, and Sargent-Welch) have indicated that they no longer sell asbestos.

Table 3 lists existing asbestos standard reference materials. It should be noted that

these are "existing" materials and are not necessarily available and accessible to the

general scientific community.

The UICC standard reference samples of asbestos are the most widely known, used and

probably also misused asbestos standard reference materials. The materials were prepared in

1966 by the collaborative efforts of the Llandough Hospital MRC Pneumoconiosis Unit

(United Kingdom) and the Pneumoconiosis Research Unit of the National Research Institute for

Occupational Diseases (South Africa) [7,8]. The materials were prepared specifically to reflect the purity and size distributions of the asbestos dusts encountered by industrial

workers in the mining, processing, milling, and fabrication of asbestos and asbestoscontaining products. The materials were intended primarily for use in biological response

studies, rather than as calibration standards for instrumental analysis and analytical

methods development. Therefore, the materials are relatively impure, in terms of classical

concepts of "standards".

Although the UICC standard reference samples were extensively characterized, including

elemental composition determinations, a major shortcoming of the UICC asbestos samples is

the lack of phase purity documentation. No data were obtained on the percentage of the

actual asbestos in each sample nor were data obtained for the concentrations and types of

all impurity mineral and non-mineral species present. The UICC anthophyl 1 i te is especially

impure and contains up to 10 percent talc.

The size distributions reported for the UICC standard reference asbestos samples based

on fiber length measurements were made by optical and electron microscopy fiber counts

[8]. These data are therefore not mass size distribution data and are not easily converted

into mass size data. Mass size distribution data officially supplied with the samples

simply list the percentage of "respirable" particles in each sample [8]; however, since

these data were obtained by sampling of dust clouds created from the samples, it is con- ceivable that these mass data are biased against the larger, non-respi rable particles

[9]. Our experience with the UICC samples suggests that they are in size ranges useful to

8
Table 3. Current Asbestos Standard Reference Materials.

UICC Standard Reference Samples - 500 kg each

(Prepared by National Research Institute for Occupational Diseases, South Africa)

Chrysoti 1

Chrysoti 1 Croci dol i te Anthophyl 1 i te

- Canada - Rhodesia

- South Africa

- Finland

NIEHS Bioassay Test Materials - 440-540 kg each

(Prepared by the U.S. Bureau of Mines)

Short Fiber Chrysotile

Long Fiber Chrysotile

Croci dol i te Fibrous Grunerite

Nonfibrous Tremolite

- Idria Range, California

- Quebec, Canada

- Kuruman Hills, South Africa

- Penge, South Africa

- Vanderbilt Mine, New York

NIOSH Analytical Reference Minerals - 1 kg each

(Prepared by I IT Research Institute)

Chrysoti le Crocidol ite

Fibrous Grunerite

Fibrous Anthophyl 1 ite

Fibrous Tremolite

Antigorite

Riebeckite

Gruneri te Anthophyl 1 ite

Tremol i te

Idria Range, California

South Africa

South Africa

Montana

Ra jasthani , Indi

West Chester, Pennsylvania

El Paso, Colorado

Luce Lake, Newfoundland

Bamble, Norway

Pennington, South Dakota

both optical and electron microscopists for the purposes of familiarizing analysts with the

physical and morphological properties of the various asbestos types. However, the size

distributions are too fine to be of use as analytical standards in quantitative analyses of

bulk samples by optical microscopy, and are too coarse for preparation of standard thin film

samples for electron microscopy. The coarse fiber contents have also caused difficulties

in sample mounting for x-ray diffraction analyses. Ideally, asbestos analytical standards

for bulk sample analysis by optical microscopy would consist predominantly of fibers in the

100 urn to 1 mm size range. For electron microscopy, the asbestos analytical standard

should contain fibers less than 100 urn in length, with no more than 10 percent of the

sample mass contained in fibers longer than 10 urn. The asbestos analytical standard

required for x-ray diffraction quantitative analysis of thin film type samples would be

similar in fiber length distribution to the electron microscopy standard.

There is now some indication that supplies of the UICC standards reference asbestos

samples are nearly exhausted [10]. Thus, the availability of these standard reference

materials in the quantities required for use by the U. S. scientific community is doubtful.

Large stocks of a few, fine-powder, wel 1 -characteri zed asbestos (and non-asbestos

tremolite) have been prepared for the U. S. National Institute of Environmental Health

Sciences (NIEHS) under the direction of the U. S. Bureau of Mines [11]. These materials were prepared principally to study the effects of orally injested asbestos in laboratory

animal s

Phase purities (i.e., percent of the desired asbestos in the total sample are estimated) [11] to be greater than 96 percent for the two types of chrysotile and 99 percent

for the crocidolite. However, the XRD data reported for the "long fiber" chrysotile does

suggest that the sample is less than 96 percent pure, since five other contaminant mineral

phases are definitely identified, two other possible contaminant phases are tentatively

9
identified, and one peak remains unidentified in the XRD pattern. If one simply assumes the

1 percent rul e-of-thumb detection level of a component in a complex matrix by XRD, at most,

the "long fiber" chrysotile can only be 95 percent pure.

The NIEHS nonfibrous tremolite and fibrous grunerite both reportedly contain other

asbestos types ^25 percent fibrous and nonfibrous serpentine in the tremolite and ~5

percent actinolite asbestos in the fibrous grunerites. Therefore, at least these two NIEHS

materials are not suitable for use as standards in development of quantitative analysis

methods specific for each asbestos type.

It should be noted that the NIEHS "long" 3 and "short" 3 fiber chrysotile samples were prepared from two entirely different starting materials - i.e., chrysotile mined from different geographical and geological origins. The reported chemical composition differences

between these two NIEHS chrysotile samples are not great (Table 4). However, the XRD data

reported suggests that non-asbestos mineral contaminants in the "long" fiber chrysotile

could be responsible for the nearly identical concentration of silicon (determined as Si0 2 ), magnesium (determined as MgO), and iron (determined as Fe 2 0 3 ) in the two chrysotiles. Since XRD detection limits for the mineral phases brucite (Mg(0H) 2 ), magnetite

(Fe 3 0 4 ), and apophyllite (KF Ca4 Si 8 0 2 o

* 8H 2 0) were not discussed in the analytical data

presented [11], the finding of these latter two minerals only in the "long" fiber chrysotile and the detection of five peaks for brucite in the "long" fiber chrysotile but only

one brucite peak in the "short" fiber sample is interpreted to indicate these contaminant

mineral phases were significantly higher in concentration in the "long" fiber chrysotile

sample. Thus, the near identical compositions of the two chrysotiles is questionable.

While both long fiber and short fiber chrysotile standard reference materials are urgently

needed for the different types of analysis methods, it is considered more desirable to obtain the required fiber size fractions from the same starting material.

Table 4. Chemical-Instrumental Analyses of NIEHS Chrysotiles [11].

Oxides

- - - Weight-percent - - -

Short Fiber Long Fiber Oxides

- - - Weight-percent - - -

Short Fiber Long Fiber

A1 2°3 0.66 1 .47 MnO 0.07 0.06

CaO 0. 32 0.05 SnO ND ND

FeO ND ND SrO ND ND

Fe 2°3 2.02 2.93 Bi

2 03 .... ND ND

MgO 40.62 40.26 Cr

2 0

3 .... 0. 17 0.06

K

2 0 ND 0.08 NiO 0.17 0.06

Si0

2 39.77 39.90 Co

2 03 .... 0.02 ND

Na

2 0 0.01 0.04 co

2

0. 78 0.51

Ti0

2 0.03 0.04 H

2 0" 1.54 1.17

Li 2 0 ND ND H

2 0

+ 12.69 12.81

Total 83.43 84.77 15.44 14.67

Grand Total

:

Short Fiber = 98.87 Long Fiber = 99.44

3 "Long" and "short" fiber chrysotile are designations given by NIEHS and the Bureau of

Mines to the two different types of chrysotile samples prepared. The short-fiber material

is mined as such; that is, the ore bodies from which the chrysotile is mined are practically devoid of fibers longer than 100 urn. The long-fiber chrysotile was prepared from a boarder size range commercial material that is sold to the plastics industry.

10
The fiber size distribution data reported for the short fiber chrysotile [11]

illustrates this material's suitability for use as a standard in electron microscopy

analyses and non-suitability as an optical microscopy analytical standard. Fiber size

distribution data on the long fiber chrysotile indicate its limited utility as an optical

microscopy analytical standard, and its unsuitabi 1 ity as an electron microscopy analytical

standard. The usefulness of either material as an analytical standard for XRD is not readily determinable from the data presented for the samples. The remaining long fibers

would likely cause difficulty in sample mounting for XRD analyses.

Fiber size distributions reported for the NIEHS crocidolite and fibrous grunerite

samples [11] indicate these materials will be of limited utility as analytical reference

standards for both optical and electron microscopy techniques. Mean fiber diameters and

volume mean fiber lengths are reported to be 0.53 and 88 urn, respectively, for the fibrous

grunerite and 0.27 and 10 urn, respectively, for the crocidolite.

Aside from the purity and particle size factors that render the NIEHS asbestos samples

of limited utility as analytical standard reference materials, the general availability of

these materials is somewhat doubtful. The bioassay study requirements of NIEHS will

likely limit the quantities of materials that NIEHS can make available for non-bioassay

studies. » Under contract to NIOSH, the I IT Research Institute (IITRI) prepared 1 kg samples of

the asbestos minerals chrysotile, crocidolite, fibrous grunerite, fibrous tremolite, and

fibrous anthophyl 1 ite. The nonfibrous mineral counterparts were also prepared under this

contract. All materials were prepared for asbestos analytical methods development being

conducted by NIOSH [12].

All the materials were prepared as comminuted powders, in particle size ranges

suitable for both electron microscopy and x-ray diffraction analysis techniques. The fine

particle sizes render the samples unsuitable as analytical standards for optical microscopy techniques.

The NIOSH asbestos samples all do contain small number (and mass) percentages of

fibers and fiber bundles greater than 40 urn in length. In addition, the crocidolite and

chrysotile samples contain significant numbers of highly tangled fiber agglomerates that

are not readily dispersed. These larger fibers and especially the tangled fiber agglomerates will present many problems in the applications of these two asbestos samples as quantitative analysis standards for electron microscopy techniques.

Purities of each of the NIOSH asbestos samples were inferred to be greater than

95 percent from the analyses conducted [12]. Thus, from the purity standpoint, these

materials would be useful to the general analytical community as asbestos standards.

However, the relatively small quantities of materials prepared and the anticipated requirements for these materials by NIOSH virtually render the samples unavailable.

4. Selection and Preparation of Samples as Asbestos Standard Reference Materials

Based on main analysis techniques and areas for which asbestos standard reference

samples are currently needed and the lack of availability of the current asbestos standard

reference materials to the general analytical community, it is obvious that a wellorganized effort must be put forth to prepare asbestos standard reference materials. That

is, none of the existing asbestos standard reference materials is either suitable for all

the analysis techniques for which they are currently required, or is available to all

those analytical laboratories that need them.

In selecting the asbestos materials that are to be processed into standard reference

materials, careful consideration must be given to the asbestos containing materials

against which the standards will be analyzed. That is, the asbestos types and origins of

the various asbestos types mined, processed, or used in the United States, should be

considered in deciding the numbers and types of asbestos standard reference materials to be prepared. As a natural material, the elemental composition of a particular asbestos

type is quite variable with geological origin. Since the variability in elemental

11
composition is dependent upon both the asbestos composition itself (i.e., degree of

substitution of one element by another within the asbestos mineral crystal structure), and

the types and amounts of nonremovable non-asbestos contaminants present, it is important

to determine if the i nterspecimen composition variability will have significant impact on pure asbestos quantitation analytical data obtained with various analysis protocols. If two different specimens of one asbestos type do not produce identical responses in one analytical protocol, where the response being measured is the key factor in obtaining

quantitative data, then either both specimens will have to be prepared as standard refer- ence materials, or the analytical procedure must be identified as nonuseable for asbestos

quantitation. For example, if an eastern Canada chrysotile gives a different calibration

curve than the Idria chrysotile in a standardized XRD procedure, then XRD can only be

applied as a quantitative analysis technique for samples in which the origin of the

asbestos is known and for which a calibration curve has been prepared.

It must be emphasized that the evaluation of specimens in the raw materials selection

process for asbestos standard preparation include all steps of the anticipated analytical

protocols, particularly sample preparation steps. In the case of an electron microscopy

asbestos fiber counting analysis of samples prepared from aqueous suspensions, differences

in such properties as asbestos cation solubility, hydrophi 1 i ty , surface water adsorptionability, etc., between the asbestos standard reference material and the asbestos in the

unknown sample will prevent establishment of meaningful confidence limits in quantitative

data.

Compositions of the individual amphibole asbestos types are more variable from geographical source to source than are chrysotile asbestos compositions. However, because

there are so many fewer geographical sources of each amphibole asbestos type that have

been and are being mined commercially than there are of chrysotile, determination of the

numbers and sources of each amphibole asbestos type that should be prepared as standard

reference materials is far less complicated. Consider that over the past three decades, more than 35 different countries, including the U.S. itself, are reported to have been or are currently producing chrysotile at a level of at least 1000 short tons annually [13].

In addition, within a country, chrysotile may be mined from several distinctly different

geological occurrences. Countries producing exportable quantities of all amphibole

asbestos types number less than a dozen. Therefore, it is probable that only one standard reference material (but in several forms) of each amphibole asbestos type need

be prepared for the various requirements of the analytical community, while multiple

source derived chrysotile asbestos standard reference materials will probably have to be

prepared.

Figures 1 through 5 illustrate the wide variability in appearance that chrysotile

asbestos assumes. Table 5 lists the chemical compositions of some of these as well as other chrysotile samples. The chemical composition data would seem to indicate very

little difference between chrysotile samples collected from various geographic locations.

However, as mentioned previously, chemical compositional data reflect the composition of

the actual chrysotile as well as the contaminant mineral phases that could not be

removed. Therefore, significant compositional differences between the asbestos portion of

various chrysotile samples may be obscurred by disseminated, separate mineral phases.

There undoubtedly are some significant differences in chemical-physical properties of the

chrysotiles derived from different sources that have given rise to the vastly different

gross appearances, but these appearances have not been or cannot be readily quantified.

While significant differences in the weight losses of chrysotile with time in aqueous

suspensions, pH of chrysoti 1 e-water suspensions, hydroscopi ci ty , etc. have been reported

[14], there is no reporting of how these differences relate to chemical composition of

the chrysotile phase vs. quantities and compositions of non-chrysoti 1 e impurities. Since

these factors, especially the weight loss in water and hydroscopic tendencies, are of

great importance in quantitative analyses, particularly where the quantitation is dependent upon preparation of a calibration curve, determinations of chemical compositions

as well as these types of physical properties must be made for the various chrysotile

asbestos types expected to be encountered in analyses of samples collected in the U.S. Determination of significant differences in these properties between chrysotile specimens

will clearly point to the necessity of preparing multiple chrysotile standard reference

sampl es

.

12
QUEBEC, CANADA

Figure 1. Chrysotile bulk sample (Quebec, Canada).
CASSSAR, BRITISH COLUMBIA

Figure 3. Chrysotile bulk sample (Cassiar, British Columbia).

Figure 4. Chrysotile bulk sample (Idria Range, California)

14
Figure 5. Chrysotile bulk sample (Globe, Arizona).

It is probable that a minimum of two chrysotile standard reference materials (each in

several different forms) will have to be prepared to adequately serve the needs of the

analytical community. Approximately 85-90 percent of the chrysotile imported into the U.S.

is derived from the eastern Canada deposits [13], and therefore chrysotile specimens from

the several eastern Canada geographic locales should first be evaluated for selection of one of the chrysotile standard reference materials. A second chrysotile standard reference

material should be prepared from the Idria Range (California) massive serpentine fiber

deposits because this asbestos deposit is considered unique amongst the commercially

developed asbestos deposits worldwide [13].

Consideration should also be given to the preparation of the nonfibrous counterparts of

the asbestos mineral types. Methods must be developed for distinguishing amphibole cleavage

fragments from true amphibole asbestos. Although the existing federal government definition

of "fiber" simply avoids the question of distinction between true amphibole asbestos fibers

and amphibole cleavage fragments, it is conceivable that on-going bioassay studies will

demonstrate that the cleavage fragments fitting "government fiber" definitions pose no health risks. Therefore, analytical methods for distinguishing between amphibole asbestos

fibers and amphibole cleavage fragments may have to be developed. The problem of distinguishing between cleavage fragments and asbestos fibers of amphiboles is especially

difficult for microscopic particles as figures 6 through 9 clearly demonstrate.

Processing steps in the preparation of asbestos standard reference samples must be

selected with respect to the forms in which the samples must be prepared and with respect

to the analysis techniques that require standards. For example, comminuted asbestos standards are required for electron microscopy and x-ray diffraction analyses. However, since

crystallinity is a physical property of asbestos which is utilized in both electron

microscopy and x-ray diffraction identification of asbestos, care must be taken in the

comminution process to not alter or reduce the bulk sample as well as individual fiber

crystallinity. Chrysotile is especially susceptible to crystallinity alteration by

grinding [15,16].

15
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16
Figure 6. Ground powder form of a grunerite that is classified as a nonfibrous grunerite in

the bulk sample form. (Secondary electron image, 3,0OOX).

Figure 7. Ground powder form of a grunerite that is classified as a fibrous grunerite (asbestos)

in the bulk sample form. Blocky, non-parallel

sided particles are observed in this sample as well as the nonfibrous sample. (Secondary

electron image, 10.000X).

17
Figure 8. Bulk sample of obviously fibrous tremolite

(photomicrograph, 1.8X).

Figure 9. Ground powder of the fib- rous tremolite depicted in

figure 8. Many fibers do

not have parallel sides.

(Secondary electron image,

3,000X).

Particle sizes required of asbestos standard reference materials for electron microscopy and x-ray diffraction bulk powder and thin film samples are quite small. In the

interest of preserving materials, preventing crystallinity alterations and contamination

during grinding procedures, a serial grinding-separation procedure is recommended, once benef iciation and other raw sample clean-up processings have been completed. That is,

rather than attempting to grind an entire portion of the sample to the desired size range,

grinding should proceed for a short period of time after which the desired size material is

removed from the mixture of partially ground sample (e.g., by flotation); the oversize

material is then returned to the grinder and ground for a further short period of time.

The processes of grinding and removal of the fines, though slow, minimizes heat build-up

in the sample that could result in crystallinity alteration and also minimizes tangling of

the fibers into non-dispersable balls [7].

18
The grinding-separation process also provides for simultaneous preparation of the

various size ranges of materials required by different analytical techniques. For example,

the coarser fibers required by polarized light microscopy analysis techniques could be the

remaining "oversized" material after the first one or two grinding-separation steps. By

preparing the various size ranges required from one starting material, ambiguities in the sameness of two asbestos samples from different sources is eliminated.

The chrysotile and tremolite bulk samples and ground asbestos samples portrayed in the

photographs were collected and processed under NIOSH Contract No. 210-75-0043. Mr. George

Yamate of I IT Research Institute provided the scanning electron micrographs of the tremolite and grunerite samples.

References

[1] U.S. EPA, OPTS, Interim Method for the Determination of Asbestiform Minerals in Bulk

Insulation Samples, prepared by Research Triangle Institute. Research Triangle Park,

N.C. ; June 1980.

[2] U.S. DHEW, NIOSH, Chrysotile; NIOSH Analytical Method No. P&CAM 309 in NIOSH Manual

of Analytical Methods, Vol. 5. D. G. Taylor, coordinator. U.S. Government Printing

Office, Washington, D.C.; 1979.

[3] U.S. EPA, ESRL, Electron Microscope Measurement of Airborne Asbestos Concentrations:

A Provisional Methodology Manual. EPA Publication No. EPA-600/2-77- 1 78. Revised

June 1978.

[4] U.S. DHEW, NIOSH, Asbestos Fibers in Air; NIOSH Analytical Method No. P&CAM 239 in

NIOSH Manual of Analytical Methods, Vol. 1. D. G. Taylor, coordinator. U.S. Government Printing Office, Washington, D.C.; 1977.

[5] U.S. EPA, OPTS, Friable Asbestos-Containing Materials in Schools: Proposed Identification and Notification. Federal Register, Part VI. Vol. 45. No. 182; Sept. 1980.

pp. 61965-61997.

[6] U.S. Department of Education, Asbestos Detection and Control: Local Educational

Agencies; Asbestos Detection and State Plan: State Educational Agencies. Federal

Register, Part V, Vol. 45. No. 182; Sept. 1980. pp. 61950-61964.

[7] Timbrel 1, V.; Rendall, R. E. G. Preparation of the UICC Standard Reference Samples of

Asbestos. Powder Technology 5; 1972.

i [8] Rendall, R. E. G. The Data Sheets on the Chemical and Physical Properties of the

UICC Standard Reference Samples. Shapiro, H. A., ed. Pneumoconiosis Proceedings of

the International Conference; 1969 Johannesburg. Oxford University Press; 1970.

[9] Timbrell, V. Characteristics of the International Union Against Cancer Standard

Reference Samples of Asbestos. Shapiro, H. A. , ed. Pneumoconiosis Proceedings of

the International Conference; 1969 Johannesburg. Oxford University Press; 1970.

[10] Rendall, R. E. G. personal communication.

[11] Campbell, W. J.; Huggins, C. W. ; Wylie, A. G. Chemical and Physical Characterization

of Amosite, Chrysotile, Crocidolite, and Nonfibrous Tremolite for Oral Ingestion

Studies by the National Institute of Environmental Health Sciences. U.S. Department

of the Interior. Bureau of Mines Publication RI-8452; 1980.

[12] Graf, J. L. ; Ase, P. K. ; Draftz, R. G. Preparation and Characterization of Analytical

Reference Minerals. NIOSH Technical Report. Publication No. 79-139; June 1979.

19
[13] LeFond, S. J. ed. Industrial Minerals and Rocks, 4th Edition. American Institute

of Mining, Metallurgical and Printing Engineers, Inc. Baltimore, Md. ; 1975.

[14] Michaels, L. ; Chissick, S. S. , eds. Asbestos, Vol. 1. Properties, Applications and

Hazards. John & Wiley & Sons; New York, N.Y. ; 1979.

[15] Martinez, E. The Effect of Particle Size on the Thermal Properties of Serpentine

Minerals. Ameri can Mi neral ogi st . 46; 1961.

[16] Caillere, S. ; Henin, S. The Chlorite and Serpentine Minerals in Mackenzie R. E. , ed. The Differential Thermal Investigations of Clays. Mi neral ogi cal Society, London;

1957.

[17] Stewart, L. A. Mining Methods and Costs, Regal Asbestos Mine. Jaguays Corp., Gila

County, Arizona. U.S. Dept. of the Interior. Bureau of Mines Information Circular

No. IC-7986; 1961.

20
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gai thersburg , Maryland, October 1-3, 1980. (Issued March

1982)

GLASS AS A MATERIAL FOR ASBESTOS STANDARDS

David J. Cronin, Douglas H. Blackburn, and Wolfgang K. -Haller

Ceramics, Glass, and Solid State Science Division

National Bureau of Standards

Washington, D. C. 20234

Abstract

Glasses are finding increased use as analytical standards, particularly for micro-analytical techniques, because they can be produced with

a broad composition range and excellent homogeneity and durability. The

nature of glass also allows the production in several forms, e.g., bulk,

spheres , or fibers.

Glass may prove useful in two areas of asbestos standards, for

chemical composition and for physical dimensions.

Some compositions of interest in asbestos studies have been

produced as glasses. Others cannot be made as glasses without additions

to aid particular properties, such as to reduce melting temperatures or decrease the crystallization tendency. It does appear several

compositions of interest could be produced as homogeneous glasses for use

as composition standards if desired.

For size standards it would be desirable to have fibers of carefully

controlled diameter and length in the range of 0.1 urn diameter and 5 pm

long. Suitable fibers for producing such standards are not currently

available. However, the theory and experimental work on fiber drawing

suggest that it may be possible to alter present techniques to produce

such fibers.

1. Introduction

Glass has found increasing use in recent years as a material for a variety of standards, particularly as analytical standards for mi croanalyti cal techniques. The use of

glass for such standards is related to some of the unique properties of the glassy state.

Glass has been defined in a number of different ways; in structural terms it can be

defined as a non-crystalline solid. That is, glass lacks the long range repetitive order

of a crystal. Figure 1 shows a schematic view of a glass and a crystal produced from the

same structural units. The glass shows a more or less random orientation of structural

units while the crystal is constructed of the same structural units in a specific, repeated

orientation.

The glassy state is always metastable in terms of thermodynamics. A material can lower

its free energy by going from the disordered glassy state to an ordered crystalline arrangement. The kinetics of the transition from liquid to crystalline structure control

whether a given material can be obtained in the glassy state. For a wide variety of

materials the nucleation and crystallization behavior is such that it is possible to avoid

crystallization on cooling from the melt and thus produce glasses that are stable, with

respect to crystallization at room temperature, for an indefinite period of time.

21
Figure 1. (a) Regular crystalline lattice, and (b) corresponding irregular glass network.

Figure 2 shows a plot of specific volume vs. temperature for a glass and a corresponding crystalline material. When crystallization occurs on cooling a melt to its liquidus

or crystallization temperature, there is an abrupt volume change at the transition from the

liquid to crystalline state due to the related structural differences. In the case of

glass formation there is no sudden structural change, the volume decreases smoothly and

continuously even below the liquidus temperature. At a point near where the melt becomes

essentially a solid (viscosity of 10 13 poise) there is a change in slope in the V vs. T curve. This point is called the glass transition point, Tg. Below Tg the material is considered a solid, above Tg the material is considered a liquid. The material would be

considered a glass only below the glass transition temperature.

Materials which can be formed as glasses generally show a relatively high viscosity at their liquidus temperature. This is a major factor that results in nucleation and crystallization behavior such that the melt can be brought through the crystallization region without crystals being formed. Materials which cannot be formed as glasses or only with great

difficulty generally show a low viscosity at the liquidus temperature. The viscosity of a material during glass formation increases smoothly as temperature is decreased, this allows

the use of a number of different forming techniques for producing glasses in a variety of

shapes directly from the melt.

This brief overview of the glassy state points out some of the advantages of glasses

as materials for standards and also some of the problems that must be considered. Synthesizing a glass, one starts with a melt, that is with a liquid which can be homogenized by

stirring. This high homogeneity is retained when the melt is cooled into the glassy

state. The random structure of a glass allows a wide variety of ions to be incorporated

into a glassy matrix and still maintain the homogeneity required. In a crystalline

material the type of sites available and ions that can be accommodated are usually quite

limited. In a glass, because of the disordered structure no specific sites exist but rather

the structure can change and accommodate a variety of ions. This is particularly useful

when producing standards for analytical use where a number of elements are desired in a single matrix. The problems of partitioning between different crystalline phases or grain

boundry segregation that occur in multicrystal 1 ine materials are avoided by using a glass.

As mentioned above, glasses are not thermodynami cal ly stable and not all materials or compositions will form glasses. This presents a problem when attempting to produce a glass

to simulate the composition of certain crystalline materials.

The smooth increase in viscosity as temperature decreases allows the liquid forming of

glass by casting, pressing, drawing, or blowing. Therefore, it is possible to produce

22
LIQUID

TEMPERATURE

Figure 2. Plot of specific volume vs. temperature for glass and crystalline material.

standards in a variety of forms, e.g., bulk form, spheres, fibers, or thin films. This can

be particularly useful for producing analytical standards for certain microanalytical

techniques.

2. Glass for Use as Asbestos Standards

There are two different areas where glass may prove useful as a material for standards

related to asbestos characterization. The first as standards for chemical composition, the

second as standards for fiber dimensions.

2.1 Chemical Composition Standards

There are numerous minerals that are of interest in asbestos studies covering a wide

compositional range. Since it is desired to simulate a typical mineral the major as well

as minor constituents are specified. This brings up the problem, as mentioned above, that

not all compositions will form a glass. However, the asbestos compositions contain

significant amounts of Si0 2 , which is an excellent glass former and it is frequently

possible to produce glass near the composition of interest by making minor composition

23
alterations. Such alterations frequently include additions to reduce the melting temperature to a reasonable level and alterations to decrease the tendency to crystallize. There

are few general rules and each composition must be considered individually.

Frequently the minerals of interest will contain significant amounts of H 2 0 in the

structure. Glasses, as normally produced, however contain only trace amounts of water.

While it is possible to incorporate significant amounts of H 2 0 in a glass it involves

considerable technical difficulties. Therefore one alteration which would normally be made

would be to exclude H 2 0 from the composition.

Table 1 gives some examples of typical asbestiform materials and glasses which were produced to simulate them.

To produce a glass to simulate chrysotile the composition is approximately 50-50 MgO

and Si0 2 , on a weight basis since H 2 0 is eliminated. Li 2 0 was added to decrease the

melting temperature. This was effective but the melt crystallized rapidly on casting.

B 2 0 3 was added to the second trial melt, but the melt still crystallized rapidly. However,

using a rapid quench technique it was possible to produce thin sheets of homogeneous glass

having the composition shown in column 3.

Another composition of interest is that of crocidolite as shown in column 4. The

glass produced to simulate the composition is shown in column 5. It was necessary to decrease the total iron content and increase MgO to avoid crystallization problems.

Addition of Ti0 2 and MnO were requested and do not materially affect glass formation in this

case. The composition given is a nominal composition and indicates only that iron was added as Fe 2 0 3 and is not an indication of the redox state of the glass.

A third composition of interest is that of tremolite. Two glasses were produced to simulate this material, one with A1 2 0 3 and one without. These glasses have been certified

as Standard Reference Materials and are available from NBS as SRM 470, mineral glasses for

microanalysis.

In terms of standards for chemical composition it is possible to produce homogeneous

glasses closely simulating many abestiform compositions for use as analytical standards.

2.2 Glass for Use as Standards for Fiber Dimensions

This is an area where glass may be useful because of the unique forming properties of

glass. This is an area of importance because of the work of Stanton [l] 1 and others which

suggests that the ability of a fiber to cause mesothelioma may be related more to fiber

geometry than fiber chemistry, at least for inorganic oxides. From this standpoint it

would be desirable to have fibers of carefully controlled diameter and length for use as standards in materials characterization work. If such fibers can be produced they would

also be valuable for use in medical testing as a further test of Stanton's suggestion since

a variety of fibers having a carefully controlled diameter and length, but identical

chemistry, have not previously been available. It would be desirable to have fibers

approximately 0.1 pm in diameter and 5 urn in length.

To produce the desired fibers it is necessary to have a method not only to produce

fibers with close control of diameter but also a method to accurately cut and size the

fibers to length. A method has been developed, by the Thermal Insulation Manufacturers

Association [2], to cut continuous glass fibers into the length range desired. The method

has been used successfully to prepare fibers for animal studies, using fibers of 1-2 pm

diameter. Therefore, if fibers of 0.1 urn diameter were available there is reason to believe

they could accurately be sized with respect to length.

Unfortunately continuous glass fibers are not currently available below 1 pm diameter.

Discontinuous glass fibers can be produced having diameters down to 0.1 pm diameter. These

fibers are not suitable for use as size standards since the diameter varies along the

figures in brackets refer to the references at the end of this paper.

24
00

0) -p

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-a CO s_ o Q. 03 CO 1— CO o o

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in

Z.

<u

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Q) XI

CO

QinO

Q.EO

c_>

X5

to o o o o o c CM CM CM CM CM o o O o o O in +->

in 03 +1 + 1 +1 +1 + 1 (0 3 LT> CO LO

«J E CO CM cn cn CM CO m en cn cn

<V

O o O o +-> c CM CM CM CM o O O O o O in -t-> E 10 nj +1 +1 +1 +i

a> to

S- 33

O CM l>» r-- h- CJ E CO IX) CO O") i

o

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CJ E m

co

in

<0 in _U >, cn Q. rc >) c in

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CM

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ro

13 o E <* COCo

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CM CM

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CM CM o O o O + 1 o o o CM o CM CM O o o CM o O o CO CM CM

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CO oCM

25
length and the fibers are randomly oriented as a result of the manufacturing process.

These drawbacks make it extremely difficult to produce the sharp size distribution required

for standards or medical studies.

However, an examination of the theory and practice of producing continuous glass

fibers suggests that alterations to current methods may allow the production of continuous

'fibers in the desired diameter range. A theory of the drawing process developed by Bruckner

and Stehle [3] predicts that using a simple bushing system, as is normally used, the minimum

fiber diameter attainable is approximately 1.5 urn. This is in excellent agreement with

experimental results of fiber drawing. A simplified drawing configuration is shown in

figure 3. It consists of the bushing or orifice plate, through which the glass flows, and

a winding mechanism. A glass fiber is pulled from the orifice and attached to the drum

which is then rotated thus drawing the fiber. The flow rate of glass through the orifice is

independent of drawing speed of the drum. Therefore independent control of the glass flow

rate and the winding speed determine the final fiber diameter, if breakage does not

intervene. Such a drawing process provides very close control over the fiber diameter.

Studies have been carried out to understand the hydrodynamics of the drawing process

and to determine critical engineering parameters. These studies point out several areas of

present practices that might be altered in an effort to produce submicron fibers. To model

the drawing process the glass stream is divided into three separate areas as shown in

figure 4. The upper jet region is important not for attenuation, only a factor of 5-10,

but as a source of instability in the process. The instability is caused by surface

tension which tries to pinch off the glass stream. The surface tension forces have to

BUSHING

FIBER

Figure 3. Schematic representation

of apparatus for drawing

continuous fibers.

WINDING APPARATUS

26
final CONSTANT RADIUS

REGION

I TO WINDER

Figure 4. Glass Stream

overcome the viscosity of the glass and it is therefore desirable to have the viscosity of

the glass as high as possible at this point. However, the viscosity must be sufficiently

low to sustain flow through a small orifice. Therefore it is desirable to have the

viscosity increase rapidly with the distance from the orifice.

The central jet region is where most of the attenuation occurs, normally a factor of

50 to 100. To produce very fine fibers it is desirable to have the incoming diameters as small as possible to decrease the necessary attenuation. It is further desirable to have

the viscosity low at the start of the region and change slowly with temperature to produce

a long zone of gradual attenuation.

In the constant radius region there is no further attenuation but breakage can occur

before the fiber reaches the drum. Air turbulence caused by fiber/air and drum/air

friction as well as fiber vibration contribute to breakage of the solidified fiber.

It can be seen from this brief description that different conditions are required in

each section. It is not possible to optimize the conditions required to produce submicron fibers solely through the use of a simple bushing, natural cooling, and glass

selection. A more promising approach is to attempt to produce the desired conditions in

each section by controlling the temperature distribution of the glass stream. To increase

the stability in the upper jet region the viscosity should be as high as possible consistent

with maintaining glass flow. This can be achieved by low melt temperatures within the

crucible and by cooling the glass stream immediately as it leaves the orifice. To provide

sufficient glass flow with low melt temperatures it may be necessary to increase hydrostatic melt pressure at the orifice. This can be done by either a high hydrostatic column

or by applying gas pressure at the melt surface.

The attenuation zone could be lengthened through the use of auxiliary heaters.

Observations of the drawing process also indicate that the finer the initial diameter

the smaller the final fiber diameter. Very fine orifices may prove useful, though again

pressurization may be required to control glass flow. These alterations are shown

schematically in figure 5.

27
Experimental evidence exists which suggests that each of these modifications can be

effective. However, to date these approaches have not been combined in a system to produce

the submicrometer diameter fibers that would be required for asbestos size standards.

In conclusion, fibers suitable for use as size standards are not currently available.

The theory and practice of fiber drawing suggests modifications to current practices that

could result in the production of the submicron fibers required.

INCREASE PRESSURE

KEEP MELT TEMPERATURE LOW

REDUCE ORIFICE SIZE

COOLING MECHANISM

AUXILIARY HEATER

Figure 5. Schematic representation of suggested alterations to the drawing apparatus

for the production of submicrometer fibers.

References

[1] [a] Stanton, M. F. ; Layard, M. ; Tegeris, A.; Miller, E. ; May, M. ; Kent, E. Carcinogenicity of fibrous glass: Pleural response in the rate in relation to fiber

dimension, J. Natl. Cancer Inst., Vol 58 (1977), 587-603.

[b] Stanton, M. F. ; Wrench, C. Mechanisms of mesothelioma induction with asbestos and

fibrous glass, J. Natl. Cancer Inst., Vol. 48 (1972), 797-821.

[c] Stanton, M. F. ; Layard, M. The Carcinogenicity of Fibrous Minerals, NBS Special

Publication 506, Proceedings of Workshop on Asbestos: Definition and Measurement

Methods, July 18-20, 1977.

[2] Thermal Insulation Manufacturers Assn. , Mount Kisco, New York.

[3] Stehle, M. ; Bruckner, R. Simultaneous Rheological and Thermal Analysis of Glass

Fiber Drawing, Glasstechn. Ber. , Vol. 52 (1979), No. 4, 82-91, No. 5, 105-115.

28
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gai thersburg , Maryland, October 1-3, 1980. (Issued March

1982)

THE NBS PROGRAM FOR STANDARD REFERENCE MATERIALS

R. Keith Kirby

Office of Standard Reference Materials

National Bureau of Standards

Washington, D. C. 20234

The Standard Reference Materials (SRM's) program will be described and specific

examples given of SRM's and how they aid in measurement compatibility and traceability in

the environmental area. An SRM is a material or device for which a chemical or physical

property has been certified by the National Bureau of Standards (NBS). SRM's can be used to calibrate measurement systems, to evaluate measurement methods, and to provide traceability

of the measurement to NBS. In general, measurements are made at NBS or in cooperating

laboratories in such a way that accurate values for the property are obtained and the

present best estimate of the true value and its uncertainty are certified. Current SRM's

in the environmental and industrial hygiene areas include analyzed gases for atmospheric

pollutants such as S0 2 and N0 2 ; lead, sulfur and mercury content in fuels and water;

quartz, beryllium, and other metals on filters to be used in determining their level in an industrial atmosphere; and trace element concentrations in coal, fly ash, fuel oil, urban

aerosols, and water.

The National Bureau of Standards (NBS) has issued Standard Reference Materials (SRM's)

since 1906. During most of these years the emphasis has been on metallic compositional

reference materials. During the last 10 years, however, there has been a large increase in

requests for SRM's in clinical chemistry, nuclear materials, physical science, and electronics as well as in environmental and occupational health areas. Some examples of SRM's

in the environmental and industrial hygiene areas are shown in Table 1. Although the con- centrations of more than one element are generally certified in these SRM's the concentration of lead, which is of particular interest to the health of individuals, has been

indicated in this table.

Table 1. Some Examples of Standard Reference Materials for Use in The Environmental and

Industrial Hygiene Areas.

Lead

SRM Material Certi f ied (uq/cj)

1643a Water 17 trace elements 0. 0027

1577 Bovine Liver 16 major and trace elements 0. 34

1566 Oyster Tissue 19 minor and trace elements 0 48

1635 Subbituminous Coal 14 trace elements 1 9 1632a Bituminous Coal 18 trace elements 12 4 1636a Lead in Reference Fuel lead only, 4 levels 11 2 to 76.4

1633a Coal Fly Ash 20 major and trace elements 72 4 1645 River Sediment 13 major and trace elements 714 0 1648 Urban Particulates 9 elements 6550 0 2676a Metals on Filter Media 4 elements, 3 levels 6 9 to 29.64 ug/f ilter

2661b Benzene on Charcoal 16 to 54 ug/tube, 4 levels

1661-1664 SO 2 in N

2 for Stack Gas 480 to 2521 ppm

29
Standard Reference Materials are well-characterized, homogeneous, stable materials

or simple artifacts with specific properties that have been measured and certified

by NBS. They are used to help develop test methods and to calibrate measurement systems.

By using SRM's the long-term reliability and integrity of measurement processes and the

development and enforcement of equitable regulations can be assured.

Two general procedures are followed in the certification of an SRM:

1. Measurement by a so called "definitive" method of known accuracy having very small

systematic errors.

2. Measurement by two or more independent methods having systematic errors that are estimated to be sir ^11 relative to the desired certification accuracy. Measurements

are generally made at NBS but may be made by a group of cooperating laboratories,

preferably using previously issued SRM's as controls.

When a user purchases an SRM, a certificate is provided (see figure 1) with the

material that clearly states the certified value(s), the uncertainty of the value(s),

directions for using the material, limitations on its use, and information on its possible

instability. The certified value is the present best estimate of the "true" value and is

not expected to deviate from the "true" value by more than the stated uncertainty. Ideally

the uncertainty includes the systematic error of the measurement process, the measurement

imprecision, and the material i nhomogeneity.

The difference between primary reference materials (SRM's) and secondary or working

reference materials should be made clear. While both should be homogeneous and stable, the

working standards should relate very closely to the real world and should as much as possible match the material being measured. Working standards are not only more economical

but if they are carefully calibrated they can very often lead to a more accurate calibration of the measurement system. The SRM on the other hand should be free of unnecessary

complications and unambiguity in the certified property. A good example of this difference

is found in the determination of the level of an anticonvul sent drug in a patient's blood.

The method generally used in a clinical laboratory can be very inaccurate so that calibration is needed. Working standards can easily be made in the clinical laboratory by

spiking whole serum with the drug at several weighed-in levels. The drug and the whole

serum, however, can have interferences that still make the calibration questionable. To

help this measurement problem, SRM 900 was prepared with four high-purity drugs and a specially processed serum so that all interferences were removed. After preparation the

concentrations of the drugs were determined by using two independent methods which gave

excellent agreement. This SRM can now be used sparingly by a clinical laboratory to calibrate its working standards and to verify the measurement procedure.

The role of the Office of Standard Reference Materials is to provide funds and directions for the development, procurement, and certification (measurement) of SRM's. Funds

for these activities come from regular appropriations and the NBS Working Capital Fund.

Monies used for the direct production of an SRM are returned to the Working Capital Fund as the units are sold. A surcharge on each unit of an SRM provides funds for the operation of

the office which includes packaging, storage, inventory, sales, and shipping as well as the

management of the program.

Criteria for the production of an SRM include a positive evaluation of its technical

impact on a measurement system, the user's acceptance and support, material availability in

suitable form and as a homogeneous lot, and the lack of any serious technical risk in its

preparation and certification. The production cycle for an SRM typically takes about two

years for planning and development and two years for preparation and certification. This

cycle includes the following steps:

1. Identification of the needs of a measurement system. (In the case of asbestos detection and analysis there seems to be a need for a bulk powder for assurance checking of

overall sample handling and measurement techniques for both chrysotile and amphibole

forms, for powder in solution to evaluate sample preparation, and for fibers on a filter to qualify a TEM and the operators.)

30

i
U.S. Department of Commene

Juanita M. Kreps

Secretary

National Bufeifti "f Standard

Urnesl Anibler, Acting Director Rational ^Bureau of Standards

(Certificate

Standard Reference Material 1642a

Mercury in Water- ng/mL

This Standard Reference Material is intended for use in the primary standardization of instruments and tech- niques used for the determination of mercury in water. It is intended for use as received, without dilution or other alteration. The concentration of mercury in this Standard Reference Material is at, or near, the detection

limit of most commercial instruments used for the determination of mercury in water. It is to be used for the primary standardization of these instruments near these detection limits where many analytical problems occur. Mercury Concentration 1.10 ± 0.06 ng/mL

The uncertainty value shown, ±0.06, expresses an estimate of the overall uncertainty of the certified value. The

uncertainty value, ±0.06, includes twice the standard error of the average by two analytical techniques (a total of 36 determinations) plus an estimated upper bound for possible systematic errors.

Stability: Trace mercury solutions have been a constant problem when long-term storage is required. Below

the fxgj mL level, mineral acid stabilization is not sufficient. A new stabilizing technique has been applied to this Standard Reference Material that allows for prolonged storage. Gold, as the tetrachloride, has been added in a concentration 25 times that of the mercury. The gold ion, in conjuction with the normal mineral acid, has proven to be an effective stabilizer. It is recommended that this Standard Reference Material not be used after ONE YEAR FROM DATE OF PURCHASE.

This Standard Reference Material was prepared by J. R. Moody. Atomic absorption analyses were performed

by T. C. Rains and J. D. Messman, and neutron-activation analyses were performed by H. L. Rook.

The overall direction and coordination of the technical measurements leading to the certification were per- formed under the chairmanship of H. L. Rook. The statistical evaluation was done by J. Mandel.

The technical and support aspects involved in the preparation, certification, and issuance of this Standard

Reference Material were coordinated through the Office of Standard Reference Materials by R. Alvarez.

Washington, D.C. 20234 J. Paul Cali, Chief

August 24, 1977 Office of Standard Reference Materials

(over)

Figure 1. Example of an SRM certificate.

31
4.

Development of the candidate SRM and resolution of all problems associated with its

preparation, homogeneity, measurement, and use.

Procurement and preparation of the material in sufficient quantity to provide a 5 to

10 year supply.

Evaluation of the bulk material or the artifacts to assure the homogeneity of the

entire supply.

5. Measurement of the property to be certified to provide the basis for certification.

6. Preparation of the certificate and making the SRM available for sale.

This cycle can be illustrated by following the development and final certification of

an artifact to calibrate the magnification of a scanning electron microscope (SEM). It had

been documented (by about 1970) that the uncertainty in the magnification scale of an SEM

was on the order of 10 to 30 percent. To calibrate this scale, laboratories had to use artifacts such as ruled gratings and wire mesh screens. The difficulties in using these

artifacts were in preparing them for viewing in an SEM, in accepting an average value for

the spacing', and in the variable and ill-defined image of the line or wire. Research

started at NBS in 1973 resulted in the development of an electroplating technique for

building up various thicknesses of nickel between very thin layers of gold (about 0.06 pm

thick). When this sandwich structure is cut and viewed edge-on the gold layers appear as very thin straight lines. Problems that had to be overcome included uniformity across a sheet, delamination of the layers, gas bubbles, and strain. Production was started in 1976

which resulted in the successful preparation of a composite sheet from which about 200

pieces were sheared. Each piece was mounted in a copper filled epoxy inside a short length

of stainless steel tubing and metal lographical ly polished. A Knoop identification placed

on the specimen is used to identify the measurement area (See figure 2). Patents for this

device and process were issued to Ballard, Ogburn, and Young of NBS in 1978 and 1979.

Evaluation of homogeneity and measurement for certification were made at the same time.

Each unit was measured by comparing to a master unit which had been directly measured with

an interferometry technique. Nominal 1, 2, 3, 5, and 50 pm spacings are certified. Units

which had unacceptable line spacings were discarded. For this initial lot the total

uncertainty for each spacing of 1 through 5 pm was 0.04 pm and for the 50 pm spacing was 0.48 pm. These uncertainties were the linear sums of the uncertainties associated with

the calibration of the master unit and the comparison of each unit with the master unit.

Line 012 3

Figure 2. Standard Reference Material 484, SEM magnification standard

(A Stage Micrometer Scale).

32
This Standard Reference Material, designated as SRM 484, can be used to calibrate the

magnification scale of an SEM to an accuracy of five percent or better in the range of

1000 to 2,000X. One hundred and fifty units of this SRM went on sale in 1977 and were sold

out within six months. Since then, SRM 484a (the second lot) has been certified and nearly

sold out while the third lot is in the process of being certified.

Not all SRM's that are useful in the verification of measurement systems have a chemical or physical property that is certified. Examples are SRM 1010, the Microcopy Resolution

Test Chart and SRM 469, the SEM Resolution Test Sample. SRM 1010 contains 26 5-line

patterns (see figure 3) and is only certified to have high contrast and to be defect free.

Standard Reference Material 1010 (Microscopy Resolution

Test Charts). This Standard Reference Material is

intended to be used to determine the resolving power of

microscopy systems. It meets all of the requirements

for ISO Test Chart No. 2, as described in International

Standard ISO 3334-1976(E).

In summary it can be noted that a compatible measurement system contains at best five

components:

1. A uniform system of units;

2. Primary and secondary reference materials;

3. Definitive and reference methods of measurement;

4. Field methods of measurement;

5. Long term stability and quality assurance.

Figure 3.

1

1.25 1

1.4 I

L6

Ideally, a definitive measurement method is used to certify an SRM which in turn is

used to develop and calibrate reference methods and secondary standards. These in turn are used to calibrate field methods of measurement. The long term availability of the SRM from

NBS assures the continued quality control of the system.

3 3
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gaithersburg, Maryland, October 1-3, 1980. (Issued March

1982)

IDENTIFICATION AND QUANTIFICATION OF ASBESTOS IN CONSTRUCTION MATERIALS

USING POLARIZED LIGHT MICROSCOPY: THE NEED FOR STANDARDS

R. L. Virta, K. B. Shedd, and W. J. Campbell

United States Department of the Interior

Bureau of Mi nes Avondale Research Center

4900 LaSalle Road

Avondale, Maryland 20782

Abstract

The Bureau of Mines Particulate Mineralogy Unit, in cooperation

with the Environmental Protection Agency (EPA), conducted a round robin

program to evaluate the reliability of analyses of asbestos-containing

building materials by polarized light microscopy. This microscopic

technique was selected by EPA as the principal analytical method for

asbestos identification and quantification in their program to evaluate

the potential health risk from exposure to airborne asbestos in public

bui 1 dings.

Results of the round robin show a need for monomi neral ic reference

samples of asbestos and nonasbestos components of these materials to aid

in identification training. Also necessary are bulk standards containing known amounts of asbestos to be used in verification and quality

control of quantification techniques.

1. Introduction

The identification and quantification of asbestos in insulation materials were investigated by the Bureau of Mines Particulate Mineralogy Unit (PMU). One component of

the PMU's mission is to provide assistance to other Federal agencies in mineral particulaterelated problems. The Environmental Protection Agency (EPA) requested PMU assistance in

the evaluation of polarizing light microscopy (PLM) for determination of asbestos in

insulation and construction materials in public buildings, particularly schools.

In the guidelines developed by EPA for evaluating the potential health risk from

exposure to airborne asbestos in public buildings [l] 1

, PLM was selected as the principal

analytical method for measurement of asbestos in various materials used in construction

prior to the EPA ban on the use of sprayed asbestos-containing materials. The choice of

PLM was made on the basis of the need to determine asbestos in moderately complex samples,

the speed and relatively low cost of the visual method, the wide availability of PLM capability, the sensitivity of the method at very low concentrations, and the ability to distinguish between fibrous and nonfibrous forms of a mineral, for example, chrysotile and

antigorite. No other instrumental method such as x-ray diffraction or infrared spectrophotometry offers all of these features.

Since visual observation is the basis of the PLM method, reliability is strongly

operator dependent. To obtain a measure of the reliability of the identification and

quantification versus operator training and experience, the Bureau of Mines, with assistance

from EPA, conducted a round robin PLM evaluation using samples of insulation and related

materials collected from various public buildings. The need for suitable standards for PLM analyses is based on the results of this round robin program together with the experience

figures in brackets refer to the literature references at the end of this paper.

34
gained in conducting PLM training in asbestos determination for State and Federal personnel. This report consists of a brief summary of the round robin program, followed by

discussion of two approaches to the development of PLM standards.

2. Round Robin Study

2.1 Samples

Fourteen samples of insulation materials submitted to the PMU for evaluation were used

in the round robin program. The samples were selected to cover a wide range both in

asbestos concentration and type of asbestos and in variations in the fillers and binders

used in the construction materials. These samples were characterized in detail by Avondale

personnel using microscopic and x-ray diffraction methods of analysis. Of the 14 samples,

there were four with chrysotile, three with amosite, two with amosite and chrysotile, and one with chrysotile and crocidolite; there was no detectable asbestos in four samples. The

asbestos concentration ranged from not detectable to approximately 80 weight-percent.

Fibrous nonasbestos constituents included rockwool , fiberglass, and organic fibers.

No attempt was made to blend each sample prior to submittal to the cooperating laboratories. Although blending is an essential step in most round robin analytical studies, it was not included in this program because of the nature of the samples. The only effective

way to blend samples of this type would be to extensively mill the materials prior to blending; however, this milling step may not be incorporated in a general analytical procedure to be adopted by EPA. The objective of the round robin program was to evaluate the

various laboratories in terms of their ability to correctly identify and semiquantitatively

determine the asbestos minerals in samples as they would be received from public buildings

for analysis rather than to generate concentration values accurate to two or three significant figures. Preliminary studies at Avondale indicated that grab sampling of the materials

as received was adequate for the purpose of the round robin. The adequacy of the grab

sampling was confirmed by subsequent x-ray diffraction analyses discussed later in this

report.

2.2 Participating Laboratories

The participants in the roundrobin consisted of both Federal and commercial laboratories interested or involved in the analysis of asbestos by polarized light microscopy.

The 26 laboratories were instructed to identify the asbestos minerals and estimate their

concentration using the PLM methods with which they were familiar. Avondale personnel

provided guidelines for identification of the asbestos minerals based on their optical

properties (see figure 1). Quantification techniques used by the various laboratories

included visual estimates, point counting, and the use of grid graticule (multiple point

count technique).

2.3 Results

Several of the participating laboratories used more than one analyst in the roundrobin

program. The participants ranged in education and experience from graduate degree mineralogists with extensive experience in PLM to physical science technicans with limited PLM training (see Table 1). As expected from this diverse group of participants, there were

some obvious difficulties in the identification of the specific variety of amphibole

asbestos. For example, low concentrations of crocidolite were incorrectly identified as either amosite, anthophyl 1 ite-asbestos , or tremol ite-asbestos. In general, from the

results of the 33 analysts there were one to two incorrect identifications per sample, and

up to four incorrect identifications on one sample. In terms of the EPA proposed algorithm

[2] for assessing the potential health hazard, this mi si denti f i cati on is not a serious

problem since all of the asbestos minerals are assigned equal rank in the hazard evaluation. This problem with misidentif ication should be resolved by additional training and

experience using reference slides prepared from asbestos and other minerals commonly found

in construction materials.

35
Immerse in

1.556 oil

V

Verify

f i bergl ass or rockwool

Veri fy

chrysoti 1

Yes

Yes

Verify

organics

<-

Yes

Isthe

fiber .

i sotropic?

v

No

Do fibers

match parallel

to length?

V

Immerse in

1.670 oil

V

Do fibers

match

perpendicular

to length?

No V

Are

fibers

blue?

V

No

Are

fibers

length

fast?

No V

Do fibers

exhibit wavy

exti nction

Yes Veri fy

amosite

Yes

Yes

Verify

croc idol ite

No ->

Verify organics. Rare

tremol ite-actinol ite

asbestos or anthophyl1 ite asbestos

Verifications should be based on all optical properties.

Fiberglass may be strained. This results in weak birefringence. Fiberglass identification must be checked.

'Organics may have the same refractive index as chrysotile.

j Chrysotile may appear to possess wavy extinction but should have been . identified earlier.

ure 1. Guideline for petrographic analysis of fibrous constituents

commonly found in construction materials 3

.

36
Table 1. Listing of Training, Experience, and Technique of Participants.

Analyst Training and Experience Technique

1A Mineralogist, moderate experience Polarizi ng microscope

IB Mineralogist, moderate experience Polarizing microscope

1C Mineralogist, moderate experience Polarizing microscope

2A Mineralogist, extensive experience Polarizing microscope

2B Mineralogist, extensive experience Pol ari zi ng mi croscope

3 Mi neral oqi st , extensive experience Pol ari zi ng mi croscope

4 Metallurgist, limited mineralogy

trai ni ng

Pol ari zi ng microscope

5A Mineraloqist, extensive experience Pol ari zi ng mi croscope

5B Geologist, limited microscopic

experience

Pol ari zi ng mi croscope

6 Mineralogist, extensive experience Polarizing microscope

7 Physical science technician,

extensive mineralogical experience

Pol ari zi ng microscope

8 Geologist-biologist Pol ari zi ng microscope

9 Not stated Dispersion staining

10 Materials scientist Pol ari zi ng mi croscope

11 Mineraloqist, extensive experience Pol ari zi ng mi croscope

12 Microscopist (novice mineraloqist),

experienced

Pol ari zi ng mi croscope

13 Chemist, traininq in microscopy Pol ari zi ng mi croscope

14 Not stated Pol ari zi ng

di spers i on microscopestai ni ng

15 Technician, moderate experience Dispersion stai ni ng

16 Mineralogist, experienced Polarizing microscope

17 Chemical engineer, training in

mi croscopy

Polarizing microscope

18 Mineralogist, experienced; geologist,

moderate experience

Pol ari zi ng mi croscope

19 Chemist, training and experience in

microscopy

Di spersi on stai ni ng

20A Mineralogist, extensive experience Pol ari zi ng mi croscope

20B Mineralogist, very limited experience Pol ari zi ng mi croscope

20C Mineralogist, very limited experience Polarizing microscope

21A Microscopist, extensive experience Pol ari zi ng dispersion

microscope

stai ni ng 21B Microscopist, extensive experience Pol ari zi ng

di spers ion

microscope

stai ni ng

22 Chemist, limited experience in

mi croscopy

Pol ari zi ng dispersionmicroscope

stai ni ng 23 Experience in polarized light

microscopy

Polarizing microscope

24 Not stated Not stated

25 Not stated Not stated

26 Geologist, moderate experience

37

Polarizing microscope
The most serious problem noted was the quantification of the asbestos minerals.

Table 2 lists the number of participants reporting asbestos concentrations of <1 percent,

1 percent to 10 percent, and >10 percent for each sample. Where a participant listed

"trace" concentrations, the estimate was placed in the < 1.0 percent category. When

estimates were given as ranges which overlapped two categories, they were placed in the

higher concentration category; for example, 5 to 15 percent would be placed in the

>10 percent category. If two asbestos types were identified, the percents were summed,

and if the ranges were given, the two upper ends of the ranges were summed to get the

maximum asbestos estimate.

Table 2. Quantification of Asbestos Content by Polarized Light Microscopy. 3

Number of Participants Reporting

b Asbestos, weight-percent . . <1 .0 1 to 10 >10

Sample No.

7154A . .... 0 3 29

7154B .... 8 17 7 7335 .... 6 15 11 7336 .... 4 22 6 7385 .... 17 0 0 7389 .... 5 0 0 7500 .... 1 0 31 7504 .... 0 3 29

7505 .... 29 2 1 7506 .... 25 7 0 7507 .... 2 16 14 7508 .... 0 2 30

7510 .... 0 0 32

7512 .... 30 2 0

laboratory No. 19 results not included.

Value is the sum of all asbestos minerals reported.

On the samples with zero to trace asbestos content such as 7385, 7389, 7505, 7506, and

7512 there was general agreement of <1 percent with one exception. Similarly, for some samples with high asbestos content such as 7154A, 7500, 7504, 7508, and 7510 there was good

agreement. In contrast, there were several samples such as 7154B, 7335, 7336, and 7507

where the reported values ranged from zero to high concentrations with a significant

number of laboratories in each range. For the latter samples the range of values could

reflect either problems in quantification or unacceptable variability in the grab samples

submitted to the participating laboratories. Two of these samples, 7154B and 7507, were

layered. Sample 7335 had areas of high fiberglass concentration. These types of inhomogeneous samples can pose difficulties to the analyst. Sample 7336 has an asbestos content

near 10 percent, so high and low estimates could put it into different categories.

To check the variability of the grab samples, several of the laboratories reporting

diverse results were requested to return the unused portion of specific samples to Avondale

for homogeneity evaluation. Each of these returned samples was reduced to minus 20 mesh in

a Wylie mill, then blended for 4 minutes with a Spex mixer. One-half gram amounts of each

sample were prepared as 1-1/2-inch-diameter pellets using a starch-type binder for x-ray

diffraction intensity measurements on the chrysotile and amosite peaks.

38
Sample 7510 had total asbestos concentrations that were fairly consistent among

laboratories. However, the reported concentrations for amphibole asbestos and chrysotile

ranged from 20 to 95 and 0 to 40 weight-percent, respectively. X-ray diffraction data for

five grab samples returned to Avondale by the participating laboratories are listed in

Table 3. There is no significant difference in the measured x-ray intensities among the

five samples considering the reproducibility of the sample preparation and x-ray measurement. The reader should be cautioned that no attempt was made to achieve high-precision

x-ray measurements since this level of precision was not warranted for this study.

Similar agreement in x-ray diffraction intensities was obtained on other samples that had

large variation in reported concentrations. Therefore, it was concluded that the grab

sampling technique employed was adequate for this round robin study.

Table 3. Comparison of PLM Determined Asbestos Concentrations and X-ray Diffraction

Intensities - Sample 7510.

Concentration, Weight-Percent X-ray Diffraction Intensity

Laboratory

Estimates by

Amphibole Asbestos

PLM

Chrysoti le

Counts

Amosite

per Second

Chrysoti le

3 75 0 570 320

4 50 5.5 575 365

9 20 5 440 290

12 40 40 480 280

20A 75 25

B 90 0 470 300

C 95 5

Two factors that can account for the large variations in reported concentrations are systematic bias by the analysts, and sample heterogeneity at the microscopic level.

Systematic bias is defined as the bias imposed by a particular operator's sampling and/or

quantification techniques that results in a tendency to have high or low estimates for all

samples examined. This bias can be reduced by training and experience using primary or secondary standards of known asbestos content.

Microscopic heterogeneity imposes a serious limitation on the precision, and hence the

accuracy, of the concentration measurement. Sample heterogeneity at the microscopic scale

is illustrated by the data in Table 4. These data represent visual estimates on six different slides prepared for each sample. At the high concentration levels represented by

these samples, there is no serious problem in placing the asbestos concentrations in a trace, minor, or major classification. However, as the asbestos concentration decreases to approximately 1 percent level, the PLM data become increasingly less precise on a relative

scale. The data in Table 5 were obtained on blended samples prepared as possible standards

for EPA by the Research Triangle Institute. These data are actual point counts by PLM. Samples A, C, H, and K indicate the large relative uncertainty per slide at approximately

the 1- to 2-percent level. The 1 -weight-percent level is given significance by regulatory

agencies in that materials containing 1 percent or more asbestos are to be regulated as asbestos-containing materials [21. The objective here is to point out the analytical

uncertainty at this critical concentration level. This example also points out that at low concentration levels it is possible to have no asbestos present in some slides

examined, thus increasing the chances of false negative results. False positive results

are the result of misidentif ication or contamination; both can be eliminated through

proper training in sampling and in identification and analytical procedures.

39
Table 4. PLM Visual Estimate of Asbestos Concentration Using

Unmilled Samples.

Concentration, weight-percent

Sample Asbestos Type

Slide

No. 1 SlideNo. 2 Slide

No. 3 SlideNo. 4

Slide

No. 5 SlideNo. 6

2029 Chrysoti le 20-30 20-30 30-40 10-20 30-40 40-60

Amosite 40-50 50-60 60-70 40-50 40-50 30-40

2030 Chrysoti le 70-80 50-60 20-30 40-50 40-50 30-40

Amosite 20-30 30-40 40-60 40-60 30-40 50-60

2031 Amosite 5-20 30-50 5-10 15-25 5-10 15-30

2032 Amosite 8-15 5-15 5-10 5-20 5-10 25-40

Table 5. Variation in Counting Asbestos Fibers Using Polarized

Light Microscopy.

Asbestos Fibers per 50 Particles Counted

Sample A B C D E F G H I J K

lide:

1 .... 0 17 0 9 4 0 2 2 10 26 0

2 .... 0 13 1 7 5 0 3 2 26 20 0

3 .... 1 12 0 8 4 0 2 1 19 24 1 4 .... 1 16 0 6 4 ND 3 2 24 30 0

5 .... 1 8 0 9 7 ND 2 0 22 23 0 6 .... 0 1

1

0 10 2 ND 1 1 25 28 1

7 .... 0 10 0 7 1 ND 1 1 20 27 0 8 .... 1 8 0 12 2 ND 4 0 22 22 0

ND = Not determined.

The Department of Education Asbestos Task Force recently proposed that the critical

concentration level for asbestos-containing materials in place be changed from 1 percent

to 5 percent [3]. This change would reduce the analytical uncertainty by expanding the

<l-percent concentration category, where the analytical estimates are most difficult, to <5 percent. Table 6 shows the roundrobin data categorized using this proposed 5-percent

concentration level. With samples 7154B, 7335, 7336, and 7507 excluded for reasons mentioned earlier, there was closer agreement among concentration estimates using <5

percent than with the <l-percent critical concentration level.

40
Table 6. Quantification of Asbestos Content by PLM Using the

5-Percent Critical Concentration level.

3

Number of Participants Reporting

Asbestos, weight-percent . . <5 >5

Sampl e No.

7154A . 2 30

7154B 15 17 7335 13 19 7336 17 1 5 7385 17 0 7389 5 0 7500 1 31 7504 .'<::; 1 31 7505 31 1 7506 31 1 7507 7 25

7508 2 30

7510 0 32

7512 32 0

laboratory No. 19 results not included.

Value is the sum of all asbestos minerals reported.

3. Standards for Polarizing Light Microscopy

Based on the round robin and experiences gained in training personnel, two types of

standards are reguired: one for identification training, and another for establishing and

monitoring the reliability of the guantitative analyses. The first need would be met by a set of essentially monomi neral i c standards comprising each of the fibrous minerals and

synthetics used in insulation and construction materials. These "pure" standards, both as bulk materials and as prepared slides, would be used to train personnel on the correct

identification of asbestos and how to distinguish asbestos from other fibrous materials.

The UICC asbestos standards suitable for this purpose are available from Duke Scientific

Corporation, 445 Sherman Avenue, Palo Alta, California 94306. This identification training

can be greatly assisted by the use of textbooks such as the McCrone Asbestos Particle Atlas

[4], as well as basic mi neral ogi cal ly oriented microscopy texts. Short courses of 3- to 5-day duration are also available.

A second group of standards is reguired for quantification of the asbestos content

using PLM. Two suggested approaches are (1) the use of analyzed real world samples for

secondary standards and (2) the preparation of primary standards by blending known quantities of materials. The values for selected amphibole and serpentine asbestos in real world

samples can be established on a consensus (relative) basis using optical microscopy, x-ray

diffraction, and infrared spectrophotometry. However, these techniques are not necessarily

absolute because of possible systematic errors; therefore, these secondary standards would

be limited in use as relative controls only.

In special cases, absolute values can be obtained from insulation samples or standards.

One example of absolute values is the case where the insulation sample is composed of

chrysotile plus other constituents that do not lose water of hydration over the same tem- perature range as chrysotile. With this type of sample, a reliable value of the chrysotile

41
concentration can be obtained by careful measurement of the quantity of released water over

a selected temperature range, as measured by weight loss, water absorption, gas chromatography, or mass spectrometry. Because of the moderately high water content of chrysotile,

approximately 13.5 weight percent, as compared with the amphiboles' approximately 2 weight

percent, quantitative determinations based on measurement of water content are limited to chrysotile except for higher concentrations of amphiboles. Theoretically, the chrysotile

content can be determined with an absolute accuracy of ±1 percent using large samples and

careful measurement of the analytical signal or the water absorption tubes before and after

the controlled heating cycle.

The other approach is to prepare synthetic standards using careful ly weighed quantities

of the various constituents. Table 7 lists the common components of asbestos insulation

materials. The fibrous constituents are asbestos in the form of chrysotile, amosite, or crocidolite; manmade inorganic fibers such as fiberglass and rock wool; and organic fibers

such as cellulose and synthetic organics. Wollastonite and fibrous talc are rare components

sometimes observed in insulation samples. The nonfibrous components are usually calcite,

quartz, talc, vermiculite, mica, clays, gypsum, and perlite. Lime or gypsum mortar is

often used as a binding agent.

Table 7. Possible Constituents in Asbestos-Containing Insulation.

Fibrous Constituents

Amosite Mineral wool/fiberglass

Chrysotile Paper fibers

Crocidolite Synthetic fibers

Nonfibrous Constituents

Bassanite Lime

Calcite Mica

Chlorite Perlite

Clays Pumice

Diatoms Quartz

Ground glass Talc

Gypsum mortar Vermiculite

Two major problems with these synthetic standards are the degree of purity of the

asbestos minerals and adequate blending of the materials. Although one can accurately

weigh the asbestos minerals used in each standard, there is some uncertainty as to their

mi neral ogi cal purity. As examples, the two chrysotile samples used in the National

Institute of Environmental Health Sciences feeding study contained minor amounts of calcite,

brucite, talc, quartz, and opaques [5]. The chrysotile content was judged to be >96 percent

so that the maximum systematic error due to impurities is less than 5 percent absolute.

The mechanical mixing of the fibrous and platy constituents offers a challenge. A practical method found by the authors was the use of a Wylie mill that consists of a set

of rotating knife edges with a small adjustable distance from the knife edges to the inner

surface of the mill. The milled particles pass through a screen of selectable mesh size

and are collected in a suitable container. Standards prepared by this approach should be

suitable for milled unknowns; however, the use of milled standards might be of questionable

value for quantification of unmilled samples.

42
4. Summary

The asbestos minerals in construction materials can PLM by analysts with appropriate mi neral ogi cal education and

cipants who incorrectly identified some of the samples would

using monomi neral ic standards.

Quantification of the asbestos concentration by PLM is a more difficult problem because

of several factors including range of operator experience, lack of suitable standards, and

sample heterogeneity at the microscopic level. The reliability of the quantification is particularly serious at the 1- and 5-weight percent concentration levels, decision point

concentrations for the regulatory agencies. Improvement in PLM quantification can be

anticipated with additional training using primary and secondary standards, and by the use of milled unknowns and standards.

References

[1] Sawyer, R. N. ; Spooner, C. M. Sprayed Asbestos-Containing Materials in Buildings:

A Guidance Document. EPA-450/2-78-014; March 1978.

[2] Dorsey, L. C. Numerical System for Assessing Exposure to Asbestos in Schools.

Draft EPA memorandum dated March 23, 1979.

[3] Federal Register, Part XIV, Department of Education, Friday, January 16, 1981.

Asbestos Detection and Control: Local Education Agencies: Asbestos Detection and

State Plan: State Education Agencies; Final Regulations.

[4] McCrone, W. C. The Asbestos Particle Atlas. Ann Arbor, Michigan; 1980. 122 pp.

[5] Campbell, W. J.; Huggins, C. W. ; Wylie, A. G. Chemical and Physical Characterization

of Amosite, Chrysotile, Crocidolite, and Nonfibrous Tremolite for Oral Ingestion

Studies by the National Institute for Environmental Health Sciences. BuMines RI 8452; 1980. 63 pp.

be correctly identified using

experience. Round robin partibenefit by supervised training

43
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS , Gaithersburg , Maryland, October 1-3, 1980. (Issued March

PREPARATION OF ASBESTOS "STANDARDS" FOR METHODS VERIFICATION AND LABORATORY EVALUATION

D. E. Lentzen, E. P. Brantly, K. W. Gold, and L. E. Myers

Research Triangle Institute

P. 0. Box 12194

Research Triangle Park, North Carolina 27709

Abstract

The Research Triangle Institute (RTI) has supported the

Environmental Protection Agency's "Asbestos in Schools" program by

producing a method for the determination of bulk material asbestos

content. The utility of the method was investigated in an

i nterl aboratory test program which included formulated samples and

samples of in-place sprayed insulation. Preliminary analysis of the

results suggests that a reasonable estimate of asbestos weight percents

can be made by application of appropriate correction factors to real

percent determinations of optical methods. Laboratory evaluation may be

best performed through the use of nonparametric ranking of reported

results when standard samples are not available.

The potential health hazards of asbestos exposure are now well documented through

epidemiological studies of occupational exposures to dusts and fibers. Increased incidence

of lung cancers, mesothelioma, and asbestosis are correlated with duration and level of

exposure. Presently the Occupational Safety and Health Administration is enforcing standards in work exposure for the 2-3 million workers involved in mining, milling, and

production operations involving asbestos. The Bureau of Mines, the Mine Safety and Health

Administration, NIEHS, NIOSH, and others are also concerned with various aspects of this

same problem.

In addition to the clearly recognized hazards of occupational exposures to asbestos,

there is increasing concern over the potential hazards to the general population of

exposures resulting from release of asbestos into the environment from widely distributed

asbestos-containing materials. At present, over 850,000 tons of asbestos are used in the

U.S. each year, a significant percentage of which is eventually released into the environment. Preliminary attempts have been made to assess the levels of such exposures and the

associated health hazards; however, further investigation is needed.

The Environmental Protection Agency has a clear responsibility both in the area of

health risk assessment and in control strategy development. As a part of the Agency's

effort to meet this responsibility, initial attention was focused on the problem of asbestos

exposures in schools resulting primarily from release of asbestos from friable insulating

materials (figure 1). For the past two years, RTI has been involved in providing technical

assistance to the EPA in this program, including both the evaluation of laboratories

performing asbestos analysis and the development of reliable methods for quantitative

analysis. As a part of this work, it was necessary to develop wel 1 -characteri zed asbestoscontaining samples for validation of the proposed method. This presentation will focus on the use of "standard" samples in evaluating analytical methodologies and laboratory performance. The word "standard" is qualified to the extent that final values for the weight

percent of asbestos in each sample series were calculated using mean values for matrix

weight loss due to drying and dissolution during sample preparation.

44
SPRAYED ASBESTOS - CONTAINING CEILING MATERIAL

FALLOUT

4>

RATE

R.

MODE

FALLOUT

IMPACT

SECONDARY

DISPERSAL

IMPACT

RATE R

CAUSES

AIR MOVEMENT

VIBRATION

MAINTENANCE

ACCIDENTAL IMPACT

USUAL ACTIVITY

CUSTODIAL SERVICE

SECONDARY

DISPERSAL

FREQUENCY

CONSTANT

OCCASIONAL

FREQUENT

RATE

LOW

HIGH

(R,

(R

c

)

LOW TO (R

HIGH

1

Figure 1. Modes and rates of fiber dispersal. (From "Sprayed Asbestos-Containing

Materials in Buildings", EPA-450/2-78-014.

)

A method was developed for the analysis of asbestos in bulk insulation material by

polarized light microscopy (PLM) as shown in figure 2. An x-ray powder diffraction (XRD)

protocol was included as an auxiliary method to be used when independent confirmatory

analysis is required. Procedures for qualitative identification and quantitation were

included for both techniques; quantitation in the PLM protocol is performed by point

counting. This method was carefully drafted from available source information and was based in large part on information obtained in an RTI coordinated symposium at the Bureau

of Mines Avondale Research Center, October 23-25, 1979 for the EPA Office of Pesticides and

Toxic Substances. Symposium participants and several commercial laboratories reviewed the

method prior to validation testing.

In an effort to validate the proposed method, an i nterl aboratory testing program was designed and executed with the following objectives:

1. Evaluate the inter- laboratory precision and accuracy and intra-laboratory

variation in applying this interim method;

2. Compare the resulting measure of precision with that obtained in other

studies using different PLM protocols; and

3. Evaluate the error rate of the method relative to the federal 1-percent

weight criterion for asbestos content of sprayed-on insulation materials

(38 FR 8830; April 6, 1973).

45
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46
Adequate evaluation of the proposed method required the use of well -characteri zed

samples. Factors to be considered were:

]. asbestos type,

2. Vpnppcpnt at ii/pnpcc ac rnmnflrpri to TPfll world c> a.rrinlp c;

3. homogeneity,

4. matrix interferences,

5. degree of characterization obtainable, and

6. end use of data.

When the use of actual ceiling insulations was considered for the evaluative studies,

it was clear that there were major difficulties in the areas of homogeneity, matrix interferences, and degree of characterization obtainable. As a result, a decision was made to formulate a series of samples containing known amounts of asbestos (either chrysotile or amosite) in a relatively simple matrix material.

Eight series of samples, targeted at specific weight-percents of asbestos fiber, were

formulated. Table 1 presents target weights and allowable limits for matrix material and

asbestos fiber in each series. The formulated samples contained a single matrix to assure comparability within and hopefully between asbestos types. The actual weight percent of

asbestos in each sample series was determined after applying a correction factor for

weight loss due to dissolution of part of the matrix in the mixing step.

Table 1. Target Weights and Allowable Limits for Matrix Material

and Asbestos Fibers.

Target Weight of Asbestos Weight of Matrix

Series wt % Fiber Type (g) (g)

C 1 Chrysoti 10. 05 ± 0. 005 4. 95 ± 0. 05

A 4 Chrysoti 10. 2 ± 0. 01 4. 80 ± 0. 05

E 16 Chrysoti le 0. 8 ± 0. 01 4. 20 ± 0. 05

I 64 Chrysoti le 3. 2 ± 0. 01 1. 80 ± 0. 05

H 2 Amosite 0. 1 ± 0. 01 4. 90 ± 0. 05

G 8 Amosite 0. 4 ± 0. 01 4. 60 ± 0. 05

D 16 Amosite 0. 8 ± 0. 01 4. 20 ± 0. 05

B 32 Amosite 1. 6 ± 0. 01 3. 40 ± 0. 05

Blanks were provided as controls and for determining the method's potential for producing false positives. A "real world" sample containing approximately 50 percent chrysotile

was included for comparison to determine whether precision estimates from formulated

samples are misleading. Duplicates were included without the laboratories' prior knowledge

to estimate within-laboratory variance.

The sample components were carefully weighed for each sample. The asbestos was suspended in a 0.05 percent sodium dodecyl sulfate solution and sonicated to ensure good

separation of the mill grade asbestos. This suspension was mixed with the predominantly

gypsum matrix in a blendor, vacuum filtered and oven dried. Quality control procedures

employed included microscopic examination and duplicate weighings.

Twenty laboratories returned PLM results on the sample sets. There were several false

negatives at the target one percent loading and one false negative at the target four percent loading. No false negatives were reported for samples containing amosite or more

than five percent chrysotile. There was one false positive.

47
Six laboratories returned XRD results; three of these performed the requested analyses

using some variation of the proposed thin-layer method of quantitation; three used alternative bulk or thick-layer methods of quantitation. However, none of the methods were strictly equivalent.

The following statements are preliminary observations which are subject to revision

pending further evaluation.

(1) The point count method appears to be a reasonable but somewhat high estimator

of asbestos content.

(2) Laboratories using the point count procedure reported more accurate results

than those laboratories using their own quantitation procedures.

(3) XRD results were generally good, and of comparable accuracy and precision to the point-count method.

Because the result produced by XRD quantitation is directly related to the amount of

material present, calculation of precision and accuracy for the XRD evaluation is straightforward. In the PLM evaluation, however, area is measured and then related to weight . This theoretically necessitates an adjustment of the PLM data before reporting the

analysis. The following geometric and physical considerations apply.

1. Weight W is proportional to volume V;

2. V is proportional to the cube of linear measure L;

3. Area is proportional to the square of linear measure L.

Therefore, an anticipated model for the area-weight relation is the power law, A = bw or log(A) = c log(W) + log(b), with c = 2/3.

Reported PLM results are area proportion estimates (A), which require numerical adjustment in order to be regarded as weights (W). If data from all point count laboratories are pooled and the model is fit with c = 2/3 and b allowed to vary with asbestos type, then the

Pearson correlation coefficient r equals .91.

If laboratories are individually calibrated, so that the constant b is allowed to vary with asbestos type and laboratory, then r equals .98.

After fitting the model to the PLM data, empirical values for the asbestos-typedependent parameter b, with c = 2/3 are used to transform the reported areas to weight

percent values. Precision and accuracy are then determined by comparison of the derived

weight estimates with the actual weight loadings.

Although the areas reported by PLM cannot be reasonably regarded as weights, and the area vs. weight calibration changes with asbestos type and matrix, area values as reported

seem to carry considerable information, in that they tend to preserve the correct order of

the asbestos concentration. That is, if one sample has a greater weight proportion of

asbestos than another, then the PLM reported area will generally be greater. In this

study, it appeared to be the case without adjustments for asbestos type.

A statistical approach which utilizes this information involves the use of a correlation coefficient. The ordinary, or "Pearson" correlation coefficient is a measure of the

association between two variables, x and y (figure 3). In the method evaluation discussed

above, x was the loaded weight and y was the transformed area data. The value of r may

vary between negative one and positive one, a value of zero indicating no relationship

between the variables.

48
r(x,y) is a measure of the association between

two variables, x and y

• r(x,y) = [1/n E(xj - x)(yj - y)]/[(Sx)(Sy

)]

• -1 < r < 1 • If x and y are unrelated, then r

= 0

• If x and y are linearly related, then |r| =1

Figure 3. Pearson correlation coefficient r(x,y).

The Spearman rank correlation coefficient (r

s

) is a non-parametric measure of the

relationship between two variables. It may be computed by replacing area and weight values

by their relative ranks, and then computing the ordinary (Pearson) correlation between the

pairs of ranks (figure 4).

• rs is a nonparametric measure of the association

between two variables, x and y

• Replace xj and yj by their within-group ranks

R(xj) and R(yj); • - 1 < rs < 1 • If x and y are unrelated, then rs = 0

• If x's and y's are in same relative order, then

IrJ

= 1

Figure 4. Spearman rank correlation coefficient r (x,y).

49
This approach is applied to results reported by a laboratory in the method evaluation

study for illustration (Table 2). For samples containing amosite, the loaded weight (X),

and the reported area (Y), are each ranked in ascending order. The ranks match exactly,

and r

s equals 1 (see Table 2). If two ranks are reversed, r

g equals 0.9. If three ranks

are misplaced, r equals 0.7.

Table 2. Spearman Rank Correlation within Laboratory,

(i) Amosite results of Lab #7.

R(x)

Loaded Weight (%)

X Reported Area (%)

y R(y)

5 35.6 63 5 4 17.8 48 4

3 9.0 36 3

2 2. 3 17 2

1 0.0 0 1

(ii) With positions reversed,

r = 0.9 = 0.7

g r

s R(x) R(y) R(x) R(y)

5 4 5 3

4 5 4 5

3 3 3 4

2 2 2 2

1 1 1 1

In our preliminary results on the method validation study we find that, with no calibration or correction for asbestos type, the average Spearman correlation coefficient (r ) between weights and reported areas (not transformed) is 0.91. This compares favorably with

the Pearson coefficient (r) of 0.92 obtained by transforming data and calibrating chrysotile separately from amosite. That is, if area ranks (unadjusted for asbestos type) are analyzed, then the corresponding correlation is as high as is obtained by analyzing the

original data separately for each asbestos type.

A rank based approach may also be used to check whether a laboratory tends to report

excessively high or low results. This involves the same type of analysis applied across

laboratories. For example, in figure 5, laboratory H (1 of 20 laboratories in the point

count group) was ranked either first or second on all samples. The chances of a result

this extreme occurring by chance alone is less than one in a million. Clearly laboratory

H produces outlying values. (They are, however, internally consistent [Spearman correlation = 0.95]. )

Another variation on this approach may be used to test for significant withinlaboratory imprecision.

For the past year a quality assurance program has been conducted for laboratories which

analyze bulk materials for asbestos using mill-grade materials. The purpose of the program

is two-fold: first, to provide laboratories with simple characterized samples for quality

control; second, to preliminarily evaluate laboratory performance on qualitative and quantitative analysis of bulk samples for asbestos. Over 130 commercial and non-commercial laboratories have participated. Quarterly mailings of 4-6 samples per laboratory are planned

50
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51
for the coming year. This will require >2500 samples to be distributed over the next

year. Preparation of each of these samples in the RTI laboratory would not be cost effective. Neither would it allow adequate representation of the range of sample conditions

which occur in in-place sprayed materials.

Future i nterl aboratory studies will use samples taken from in-place sprayed-on insulation since individual sample preparation and statistical analysis is not always possible or warranted. Bulk materials for these samples will be screened by RTI for within-batch

variability, and distributed to participating laboratories for analysis. Each laboratory

results will then be compared with the means of all laboratories results to evaluate

within- laboratory consistency. Laboratories may be tested for extreme bias by looking at within-sample ranks to see if the laboratory is consistently high or low. Finally, laboratories may be evaluated with respect to precision by a related non-parametric rank-based

procedure as illustrated in this paper. This approach reduces the burden of assuring that

each sample is individually characterized and less information on each sample is required.

Laboratory reports nonetheless yield information with which to evaluate their respective

performance. Larger sample distribution programs for the purpose of evaluating laboratory

performance are therefore feasible.

52
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gai thersburg , Maryland, October 1-3, 1980. (Issued March

1982)

SAMPLE PREPARATION FOR QUANTITATIVE ELECTRON MICROSCOPE ANALYSIS

OF ASBESTOS FIBER CONCENTRATIONS IN AIR

Philip M. Cook

U. S. Environmental Protection Agency

Environmental Research Laboratory-Dul uth

6201 Congdon Boulevard

Duluth, Minnesota 55804

and

David R. Marklund

Lake Superior Basin Study Center

University of Mi nnesota-Dul uth

Duluth, Minnesota 55804

Abstract

The ability to determine asbestos fiber concentrations and size

distributions depends on the electron microscope observation of the

particles in very small fields of view that constitute microcosms

representative of the total sample. Sample preparation procedures are a critical consideration in the quest for acceptable accuracy and

precision because they have a direct influence on the number and size of

particles observed. Loss of particles, physical or chemical degradation, particle contamination, alteration of particle size

distribution, interference by debris, and non-uniform particle

distribution are problems associated with sample preparation procedures.

Basic methods for collecting particles suspended in air or water on membrane filters and then transferring them to electron microscope grids

will be discussed with respect to these problems. Preparations of

biological samples (tissues, food, fluids) and standard samples from dry

powders are complicated by the need to manipulate the sample to obtain

particle suspensions in water suitable for filtration on a membrane

filter. The desirable effects of the use of ashing, chemical digestion,

surfactants, sonif ication, centrifugation, and agitation techniques must

be weighed against the possibilities for sample alteration associated

with each manipulation of the sample. Results from i nterlaboratory

comparison studies have indicated that different sample preparation

procedures frequently result in poor agreement between the laboratories

using the procedures.

1. Introduction

In the past decade electron microscopic techniques for identifying and counting asbestos particles in water and air have been extensively developed and applied. The most

useful exposure data for evaluating health risks associated with non-occupational exposures

to asbestos and analogous mineral and synthetic fibers is fiber concentration and size

distribution determined by electron microscopy. The identification techniques of morphology, selected area electron diffraction and energy dispersive x-ray spectroscopy available

for transmission electron microscopy (TEM) are essential for the discrimination of asbestos

fibers from organic or non-asbestos mineral particles with aspect ratios exceeding 3:1.

53
The ability to determine asbestos fiber concentrations and size distributions depends

on the electron microscopic observation of the particles in very small fields of view that

constitute microcosms representative of the total sample. Sample preparation procedures

are a critical consideration in the quest for optimum accuracy and precision because they

have a direct influence on the number and size of particles observed. Loss of particles,

physical or chemical degradation, particle contamination, alteration of size distribution,

interference by debris, and non-uniform particle distribution are problems associated with

sample preparation procedures.

Although preparation procedures for air, water, tissue and other types of samples

incorporate common steps and principles, the sample type (e.g., water, air, tissue, sediment) may influence the behavior of particles during sample preparation. For example,

water filtration of particles onto a membrane filter produces a more stable deposit than

that received from filtration of suspended particles in air. This difference seems to cause more extensive particle loss or rearrangement for air samples than for water samples

during direct transfer of uncoated filter pieces to TEM grids. With these sample related

differences in mind, this discussion of the influence of sample preparation procedures on asbestos fiber analysis will concentrate on methods for asbestos fibers in air. An excellent comprehensive review of fiber counting techniques for air and water samples, including

sample preparation procedures, has been prepared by Chatfield [l] 1

.

2. Basic Procedures for Transfer of Particles from Membrane Filters to TEM Grids

Most air analysis techniques in use involve filtration of particles from the air with

either a Millipore or Nuclepore membrane filter2

. The estimation of how much air to filter

has an important impact on sample preparation. The loading of particles on the filter must

be controlled to ensure adequate sensitivity without accumulating so much sample that

particles are obscured by other particles on the TEM grids. Sample preparation procedures

can be chosen to either increase or decrease the number of particles per unit area on the

TEM grid in comparison to the original filter loading. This requires resuspension and

refiltration of the particles removed from the original membrane filter.

Particles on the surface of membrane filters can be transferred directly to a carboncoated TEM grid by solvent dissolution of the filter (figure la). Modified Jaffe washer

[2] techniques or condensation washers [3] are frequently used to dissolve the filter. The

extremely flat surface of the Nuclepore membrane filter allows a thin carbon film (approximately 50 nm) to be applied with a vacuum evaporator so that particles on the filter surface

are embedded in the carbon film. Filter pieces placed on TEM grids can be dissolved in

chloroform with a Jaffe wick technique (figure lb) to leave the carbon film with particles

embedded in it on the TEM grid [4]. This retention of particles in a carbon film prevents

movement and loss of particles during solvent dissolution of the filter. Other membrane

filters such as the Millipore type do not produce satisfactory carbon replicas because of

their very rough surface texture. Unfortunately, direct transfer preparation of TEM grids

from air particulates on uncoated membrane filters with rough surfaces can result in

greater than 90 percent loss of the particles, especially when the filter is heavily

loaded. The EPA provisional method for electron microscopic measurement of airborne

asbestos concentrations [5] utilizes the carbon coated Nuclepore filter direct transfer

technique.

Brief exposure of Millipore filters to acetone vapor results in the reduction of the

rough surface and sponge-like inner filter structure to a continuous film with most particles on the surface [6]. This allows carbon coating of the filter and TEM grid preparation

in the same way as previously described for carbon coated Nuclepore filters (figure lb)

except that acetone is used as the solvent. This "collapsed membrane filter" method is used

by NIOSH for air samples [7]. Some fibers may be lost if they are captured in the interstices of the filter rather than on the surface where they can be embedded in the carbon

film. The acetone vapor treatment of filters prior to transport from the field would

prevent loss of particles from the filter surface prior to the TEM grid preparation in the

1 aboratory

figures in brackets indicate the literature references at the end of this paper.

2 Mention of trade names or commercial products does not constitute endorsement or recom- mendation for use by the U.S. Environmental Protection Agency.

54
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55
A technique used at this laboratory for preparing TEM grids for particulates collected

from large volumes of air on 0.45, 0.8 or 1.2 micrometer pore size Millipore filters is

illustrated in figure 2. The ratio of the Nuclepore filter area used for filtration of

resuspended low temperature ashed (LTA) sample to the area of the original Millipore filter

ashed typically ranges from five to twelve. Less crowding of particles on the TEM grid is

thus achieved. This method allows longer sampling times and the analysis of average samples

by combining pieces from several filters. Thus, far greater information is obtained about

average human exposure concentrations independent of short term fluctuations caused by

varying meteorological and emission conditions.

Air Somp,e

O.ljum Filtered

Low Temperature

Ashing

Distilled H20.

0.8pm

Millipore Filter

filter piece

Filter

O.I jum or 0.2jum

Nuclepore Filter

ELECTRON MICROSCOPE

Figure 2. Technique for preparation of TEM grids from air samples collected on membrane

filters suitable for low temperature ashing.

Low temperature ashing and resuspension of ashed particles is useful for insuring a uniform distribution of particles, optimum particle areal density, and removal of interfering organic debris and coatings. However there is concern for the alteration of the

original mineral fiber size distribution and concentration with this technique. This is

most likely to happen in the ash resuspension step and is dependent on the particular procedures used. The tendency for mineral fiber particles to be comminuted during ash resuspension, particularly by separating into thinner fibers or fibrils, is also greatly influenced

by particle mineralogy. Samples containing amphibole fibers on Nuclepore filters have been

prepared by both the direct carbon-coated filter technique and the LTA/resuspension

technique described in figure 2. The TEM grids produced differed only in the sample preparation technique used (the resuspended samples were neither concentrated nor diluted).

56
Amphibole fiber concentrations determined for the two preparation techniques indicate a tendency for underestimation (up to 30%) of amphibole fibers following LTA and resuspension

with low energy ultrasonic energy. This may result from failure to separate some ash

particle aggregates so that some amphibole fibers are not identified or from the loss of some particles during low temperature ashing.

Chrysotile fibers are potentially far more susceptible to comminution during low tem- perature ashing and resuspension because of the mineral's fibrilar structure and relatively

weak interfibril bonding. Different chrysotile mineralizations produce variations in

chrysotile fiber properties which create variable resistance to ultrasonic energy and other

resuspension techniques. The use of surfactants such as aerosol 0T for resuspension of

chrysotile particles dramatically reduces the mineral to individual fibrils. Chrysotile

often occurs in air samples as bundles, clumps, and loose mats of fibrils in addition to single fibrils. It is unlikely that any LTA and resuspension procedure can prevent the

dislocation of some fibrils making up these complex aggregates. The increase in number of

fibers and fibrils counted following LTA and resuspension can be minimized, however, by

using the least ultrasonic energy necessary to achieve a uniform distribution of particles

on the TEM grids.

An outline of options used for air sample preparation is provided in Table 1. Besides

the previously discussed direct techniques and LTA variations, other techniques involving

resuspension of particles prior to grid preparation are listed. High temperature ashing is

not recommended because severe clumping of particles can occur as well as alteration of

asbestos fibers. Chrysotile, for example, begins to undergo thermal decomposition at

approximately 500 °C [8]. The micropipette method of preparing TEM grids by drying a drop

containing resuspended particles on a carbon coated grid [9] is also not recommended due to the difficulty in consistently achieving a uniform distribution of particles.

Table 1. Techniques for Air Particulate Transfer from Membrane Filters to TEM Grids: An Outline.

I. Direct Preparation from Filter

A. Nuclepore Filter

1. Without carbon coat/Jaffe wick

2. With carbon coat/Jaffe wick

B. Mi Hi pore Filter

1. Condensation washer

2. Without carbon coat/Jaffe wick

3. With carbon coat/Jaffe wick

4. Collapsed filter with carbon coat/Jaffe wick

II. Ashing

A. High Temperature/Resuspension/Filtration/Direct Transfer

B. Low Temperature/Resuspension/Filtration/Direct Transfer

C. Low Temperature/Resuspension/Micropipette (drop on carbon

coated grid)

D. Low Temperature/Rubout

III. Solvent Dissolution of Filter/Rususpension/Filtration with

Nuclepore/Direct Transfer

57
The rubout technique [10] involves grinding the low temperature ashed sample on a slide

with the edge of a watch glass and embedding the dispersed particles in a nitrocellulose

film. Portions of the film are mounted on TEM grids. The purpose of the rubout technique

is to reduce fiber bundles to individual fibers (fibrils in the case of chrysotile) so that

a fiber mass concentration can be more precisely estimated. Since the original sample size

distribution is radically changed for chrysotile, the rubout technique cannot be used to determine the original fiber number concentration. The same problem exists when extensive,

high energy ultrasound treatment of ashed air samples is used, often with a surfactant, to eliminate fiber bundles and clumps.

The dissolution of the membrane filter or a piece of the membrane filter in a solvent

which can then be refiltered through a Nuclepore filter has been suggested as a method

which would avoid ultrasonic resuspension of particles. Aside from the problem of finding

a reliable solvent for completely dissolving the filter and allowing a clean ref i ltration,

it is difficult to ensure that the resulting membrane filter will have a uniform deposit of

particles without the use of ultrasound prior to ref i ltration. Little or no data exists

for the adequacy of other agitation techniques (shaking, swirling) as a substitute for

ultrasonic treatment.

A clean room facility and/or a laminar flow hood to provide a filtered air environment

for sample preparation is necessary to minimize sample contamination particularly with

chrysotile fibers. The worst contamination threat may be from improperly cleaned glassware

or unfiltered solvents. Membrane filters themselves contain measurable levels of chrysotile fibers [11] which must be considered when ashing the filter. If a large piece of

membrane filter is ashed and refiltered on a much smaller area filter, the number of

chrysotile fibers attributable to the filter can become unacceptable. Blank samples which

reflect all sources of contamination are essential for each preparation of grids.

3. Interlaboratory Comparisons as Influenced by Sample Preparation Technique

Several sets of air samples have been collected in recent years and analyzed by different laboratories with wide variation in fiber concentrations reported. Each laboratory

used basically the same fiber identification and counting criteria. Personnel or electron

microscope capability differences do not explain the large systematic differences found

between laboratories. The different sample preparation procedures used seem to explain all or most of the interlaboratory disagreement.

3.1 Amphibole Fibers

Table 2 provides amphibole fiber concentrations reported by six laboratories for a series of 12 air samples collected by the Minnesota Department of Health in 1975. Each

laboratory received a piece of 1 . 2 urn pore size Millipore membrane filter to analyze for

each air sample. Particulate collection on the filters was accomplished with Hi-Vol air

samplers operating for approximately 55 hours so that visibly heavy sample loadings were achieved (approximately 5 cubic meters of air per square centimeter of filter). Because

small amphibole fibers such as those in these air samples are not easily comminuted, they serve as good indicators of fiber concentration underestimates introduced by fiber loss

during sample preparation or fiber obscuration by debris or overlapping particles.

In addition to the amphibole fiber concentration reported for different sample preparation techniques used by each laboratory, Table 2 contains x-ray diffraction measurements of

total amphibole mineral mass concentration on the filters. The heavy loading of particles

on the membrane filters made them suitable for direct x-ray diffraction analysis and

provided an independent measurement to which the amphibole fiber concentrations can be

compared [12]. Similar amphibole fiber size distributions and ratios of amphibole fibers to total amphibole particles are indicated by electron microscopic measurements for the 12 samples, so amphibole fiber concentrations should correlate with the x-ray diffraction

measurements of amphibole mass.

The three laboratories using LTA followed by different TEM grid preparation steps

reported fiber concentrations averaging thirty or one hundred times greater than fiber con- centrations obtained from samples prepared by the two laboratories using direct grid

58
Table 2. Amphibole Fiber Concentrations (10 3 fibers/m3

) for Air Samples Collected on 1.2 um Millipore Filters: Six Laboratories Using Different Grid Preparation Techniques.

Amphibole Cone.

Sampl

Number

(ug/m3

) X-ray

Di f fraction LTA/Rubout

LTA/C-Coat

Nuc-Jaf fe LTA/Drop Direct-CW

Di rectJaffeC-Coat

Direct/Jaffe

7144A 4 .08 335 262 390 5 9 5 5 99

7144B 2 64 164 235 177 2 7 5 4 110

7144C 2 34 323 178 174 3 0 6 6 91

9040 8 74 384 513 450 3 9 12 8 100

9041 8 89 502 448 351 2 5 6 1 160

9042 9 82 583 516 569 0 8 6 2 291

9061 1 66 53 33 67 1 0 1 6 74

9062 3 05 358 71 112 5 8 12 4 215

9063 3 19 240 76 120 0 6 3 8 20

4221 3 73 252 158 138 4 4 10 4 50

4222 2 28 100 99 96 1 4 8. 0 70

4223 4 3 394 230 221 3 2 20. 6 84

Average 307 230 239 2 9 8. 3 114

preparation techniques. One laboratory used a condensation washer and the other laboratory

the Jaffe wick method to dissolve away pieces of Millipore filter placed on carbon coated

TEM grids. These great differences in results have been attributed by Peters and Doerfler

[13] to an increase in the number of fibers in the LTA/resuspens i on techniques due to fracturing of large amphibole fibers into many smaller fibers. They conducted comparisons

of uncoated Mi 1 1 ipore/di rect and LTA/ul trasonic resuspension techniques for air samples

from the same set (Table 2) and concluded that the fiber size distributions for resuspended

samples were significantly shifted to smaller sizes, proving that fiber comminution had

occurred. We disagree with these conclusions for the following reasons:

1. Ultrasound even of long duration and high energy is not of sufficient energy

to cause amphibole fiber cleavage.

2. Although the percentage of large fibers measured in resuspended samples was

less than that measured in direct preparation samples, the number of fibers in

every size category was far greater for resuspended samples.

3. Simple carbon coating of the Millipore filters followed by direct transfer to TEM grids produced fiber concentration estimates averaging fifteen to forty

times the concentration estimates from direct preparation of uncoated filters

(Table 2) despite the severe crowding of particles and rough carbon replica of

the 1.2 um Millipore filter surface. In our opinion the carbon film embedding

the particles prevents the loss of particles during grid preparation.

4. Our own attempts to prepare grids from these and other air samples by the

direct Jaffe wick transfer of filter pieces without carbon coating produces

grids with non-uniform particle distributions radically lower in particle

numbers than expected from optical microscope examination of the original

filter.

59
5. Particle loss during grid preparation should occur preferentially for

smaller particles on uncoated filter pieces.

6. Samples prepared by LTA followed by resuspension

particle density on the filter was much lower

filter. This allows easier identification of

smal 1 fibers.

were filtered so that

than on the original

particles, particularly

Additional comparisons are available in Table 3 which contains linear regression analysis correlation coefficients for comparison of each set of results from the six different

sample preparation methods. Significant correlations at the 99.5 percent confidence level were found between each of the LTA/resuspension methods and also the x-ray diffraction

results. The two direct uncoated Millipore technigues produced data with no correlation to each other, any of the other data sets, or the x-ray diffraction results. The carboncoated Millipore direct technique produced data which correlated with the LTA/resuspension

and x-ray diffraction data at the 95 percent confidence level. These correlations provide

powerful evidence for our observation that direct transfer of air samples without carbon

coating results in large and variable loss of particles and that gentle resuspension

techniques do not result in alteration of the original amphibole fiber size distribution or number concentration.

Table 3. Interl aboratory Comparison of 12 Air Samples: Amphibole Fiber

Analysis Correlation Coefficients.

X-ray LTA/C-Coat Di rect- C-Coat/

Direct- DirectDi f fraction LTA/Rubout Nuc-Jaffe LTA/Drop CW Jaffe Jaffe

X-ray Diffraction 1. 00 0. 33

a

0. 94

a

0. 89

a

0. 06 0. 15 0. 59

LTA/Rubout 0. 83

a

1 . 00 0. 77

a

0. 82

a

0. 18 0. 33 0. 69

LTA/C-Coat Nuc-Jaffe 0. 94

a

0. 77

a

1

00 0. 95

a

0. 01 0. 15 0. 52

LTA/Drop 0. 89

a

0. 82

a

0. 93

a

100 0. 07 0. 05 0. 59

Direct-CW 0. 06 0. 18 0. 01 0. 07 1. 00 0. 43 0. 06

Di rect-Jaf fe 0. 15 0. 33 0. 15 0. 05 0. 43 1. 00 0. 04

C-Coat/Di rect-Jaf fe 0. 59 0. 69 0. 52 0. 59 0. 06 0. 04 1. 00

Significant correlation at 99.5% confidence level.

Portions of an air sample collected on a 1.0 urn pore size Nuclepore filter for amphibole fiber analysis have been prepared by the carbon coated Nuclepore Jaffe wick technique

and the LTA/ul trasonic resuspension/carbon coated Nuclepore Jaffe wick technique [14]. No significant difference existed for amphibole fiber size distribution or fiber concentration

for the two preparation techniques. This direct comparison of the two sample preparation

techniques provides conclusive evidence for the efficacy of the LTA and resuspension

techni que

3.2 Chrysoti le Fibers

Table 4 provides chrysoti le fiber concentrations reported by seven different laboratories for a series of nine air samples provided by the EPA Environmental Monitoring and

Support Laboratory, Research Triangle Park, NC in 1977 [15]. Samples were collected on 102 mm diameter 0.45 urn pore size Millipore membrane filters with membrane samplers

located near a gravel road covered with crushed serpentine rock containing chrysoti le. The

dust generated from this rock contains chrysotile [16] in a great variety of forms including small fibrils, fibers, large bundles and clumps, and chrysotile locked in a matrix of

antigorite. Most of the chrysotile mass was present in particles other than a single

chrysotile fibers or bundles. Filter loading achieved was heavy (1.3 cubic meters of air

60
per square centimeter • of filter). Some sample loss appeared to occur during shipment of

the filters in glycine envelopes and uneven distribution of particulate was observed on the filters probably due to uneven air flow through the filter during sample collection.

Chrysotile fiber concentrations reported for each sample range over four orders of

magnitude. When sample preparation techniques used are considered, these results are not

surprising. Very large fiber concentrations were obtained only for those sample preparations which utilized harsh resuspension steps such as grinding (rubout), high energy

ultrasound, or surfactant treatment plus ultrasound. These techniques not only separated

chrysotile fibers from fiber bundles and clumps, but probably caused the release of chrysotile fibers from chrysotile originally locked in a matrix of non- fibrous minerals.

Evidence for the latter effect is provided by each laboratory's estimate of chrysotile fiber

mass concentration: the three preparation techniques using harsh resuspension methods

produced mass estimates ten to one hundred times greater than those from the other preparation techniques. The comparison is complicated by the possibility of fiber loss especially

for the condensation washer and direct Jaffe wick preparation of grids from Millipore

filter pieces without carbon coating.

Table 4. Road Dust Air Samples: Chrysotile Fiber Concentration.

(10 3 fibers/m3

)

Sample Lab 1 Lab 2 Lab 3 Lab 4 Lab 5 Lab 6 Lab 7

BDL

a BDL BDL 14 220 3 1350

BDL BDL BDL BDL 120 20 1530

BDL BDL 0.4 26 17 100 1260

0.8 0.06 2.5 14 270 620 3330

1.3 2.6 7.3 150 660 1710 2430

0.2 3.0 4.2 100 340 430 2340

0.5 4.2 170 2820 1370 5310

0.2 3.3 16 34 1670 5480 9450

0.8 6.9 27 920 3070 10200

BDL = Below detection limit.

Grid preparation technique used by each laboratory:

Lab 1 - Uncoated filter direct transfer by condensation washer.

Lab 2 - Uncoated filter direct transfer by condensation washer.

Lab 3 - Uncoated filter direct transfer by Jaffe.

Lab 4 - LTA/low energy ultrasound/carbon coated Nuclepore/Jaffe. Lab 5 - LTA/surfactant, ultrasound/carbon coated Nucl epore/Jaf f e. Lab 6 - LTA/high energy ultrasound/carbon coated Nucl epore/Jaffe. Lab 7 - LTA/rubout.

4. Intralaboratory Comparison of Chrysotile Fiber Concentrations for Different

Sample Preparation Techniques

In order to test the EPA provisional method for asbestos in air, the Environmental

Monitoring Systems Laboratory at Research Triangle Park, NC recently collected air particulates on 0.4 urn pore size Nuclepore filters in the vicinity of asbestos mining operations

[17]. Eight by ten inch Hi-Vol , 37 mm personal, and 102 mm membrane samplers were used.

The filters were carbon coated prior to shipment. Direct preparation from these filters

with the Jaffe wick technique produced grids which when examined by TEM revealed very light

and non-uniform particle distribution. A set of samples containing the most chrysotile

61
was chosen for detailed study. These 37 mm filters had been collected simultaneously on side-by-side personal samplers with filtration times regulated to produce samples with 0.0,

0.017, 0.034, and 0.60 cubic meters of air per square centimeter of filter. Thus, the

greatest volume of air per unit area of filter was twenty times less than for the air

samples reported in Table 4.

TEM grids prepared by Jaffe wick direct transfer of carbon coated Nuclepore filter

pieces were difficult to examine for chrysotile fibers because of non-uniform distribution

of particles and clusters of fibers and fibrils which were imperfectly sized and counted

with procedures provided by the provisional method. Some of the complex chrysotile fiber

groupings on the filter were suggestive of fiber separation upon impact of loose clumps of

short fibrils on the filter surface (figure 3a). Other chrysotile fibrils appeared to be

either attached to the surface of other mineral particles or locked within a non-chrysoti le

mineral matrix (figure 3b).

In order to determine if LTA and resuspension of the samples could improve the fiber

distribution for counting and evaluate the influence of gentle resuspension techniques on fiber concentration estimates, two pieces of each Nuclepore filter were submitted to LTA. One subsample ash was resuspended in filtered water by five minutes of ultrasonic bath

treatment and the other subsample ash was resuspended in filtered water by five minutes of

swirling with a vortex device. Ashed air sample suspensions were filtered on to 25 mm

0.1 urn pore size Nuclepore filters, rather than 47 mm filters, in order to increase the

particle loading on the TEM grids by a factor of almost four. This nevertheless produced a particle loading approximately one sixth that on the original filters. A comparison of

chrysotile fiber concentrations obtained for the three different sample preparation techniques is provided in Table 5.

The previously mentioned problems with sizing and counting fibers and clumps, non- uniform distribution of particles on the original filters, and the small number of fibers

counted for some samples contributed to poor precision. Operator difficulty with existing

guidelines for counting chrysotile fiber clumps and loosely associated fibril aggregates

contributed to some of the largest differences. For example, there were more single fibrils

counted for sample 2312/direct preparation as evidenced by the very small mean and median

fiber size. Thus that sample has a large fiber number concentration but a very small mass concentration as a result of data based on an estimate of the number of fibrils in each of

several aggregates observed. In other samples fibrils tended to occur more in bundles or clumps that were counted as single large particles. Chrysotile identification was based

primarily on morphology rather than positive selected area electron diffraction of each

fibril. Median fiber widths were slightly reduced for both resuspension techniques as compared to the direct preparation.

The LTA/resuspension techniques may have separated loosely associated fibrils to an extent that some were counted as separate particles rather than parts of clumps. An overall

two- to three-fold increase in chrysotile fiber number concentrations for the resuspension

techniques was observed in contrast to differences of several orders of magnitude reported

for some other preparation procedures in Table 4. Chrysotile mass concentrations from

individual sample analysis are quite divergent from mean values for the three analyses of

each sample. This is not surprising in view of the statistical problem of estimating mass concentrations from samples containing large, complex chrysotile fiber aggregates which are few in number but represent a very large fraction of the chrysotile mass. The mass concen- trations for each sample preparation technique as calculated for the sum of the four air

samples were in better agreement perhaps because of the larger number of particles contributing to each estimate.

Sample 2310 was a blank filter sample. The LTA/sonify subsample contained 20 chrysotile fibrils in 10 grid squares examined. The blank chrysotile fiber concentration calculated from the average air volume for the four LTA/sonify preparations equals one fourth

the mean of the four sample estimates. Thus contamination, probably from the cross sample

contamination during LTA or from ashed filter residues [11], may significantly raise the

fiber concentration particularly when few fibers are counted per grid square.

62
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5. Conclusions

Inter laboratory comparisons of mineral fiber concentrations determined by electron

microscopy for air particulates collected on membrane filters have been shown to be greatly

influenced by sample preparation procedures. Large systematic and predictable differences

exist between results obtained by direct transfer of particles from the filter to TEM grids

and results from grids prepared following harsh resuspension procedures which cause fiber

size reduction and thus greatly increased numbers of particles. Fiber loss during direct

transfer preparation of TEM grids is prevented by use of the carbon coated Nuclepore

filter/Jaffe wick technique. Resuspension procedures following low temperature ashing of

membrane filters can be chosen to minimize but not prevent chrysotile fiber comminution

while removing interfering material, optimizing particle loading and uniformity of distribution, and allowing air particulate collection on ashable membrane filters other than the

Nuclepore type.

Improvement in methodology for chrysotile fiber analysis of air samples requires work

in a number of areas. The development of optimum sampling strategies and techniques is

needed. The definition of what sizes of particles and clumps are hazardous and the need

for fiber mass measurements must be made. This information will assist in the development

of better guidelines for counting and sizing fiber bundles, clumps, and aggregates.

Finally, suitable standard samples are necessary for interlaboratory tests of proposed

methods.

This work was supported in part by the U. S. Environmental Protection Agency, Environmental Monitoring and Support Laboratory. The authors wish to express appreciation to

Mr. Michael Beard of EMSL/RTP for air sample collection and advise. The amphibole fiber

analysis interlaboratory comparison of Minnesota Department of Health air samples was accomplished in cooperation with Mr. Kyle Bishop and Mr. Steven Ring of MDH.

References

[1] Chatfield, E. J. Measurement of asbestos fiber in the workplace and in the general

environment. In: Mineral Association of Canada Short Course in Mineralogical Techniques of Asbestos Determination. Mineralogical Association of Canada. Royal Ontario

Museum. Toronto, Canada; 1979. 111-163.

[2] Jaffe, M. S. Handling and washing of fragile replicas. J. Appl. Physics 19:1189;

1948.

[3] McCrone, W. C. ; Stewart, I. M. Asbestos. Amer. Lab. 6:10-18; 1974.

[4] Cook, P. M. ; Rubin, I. B. ; Maggiore, C. J.; Nicholson, W. J. X-ray diffraction and

electron beam analysis of asbestiform minerals in Lake Superior waters. In: Proc. Inter. Conf. on Environ. Sensing and Assessment. Institute of Electrical and Electronic Engineers. New York; 1976. 2, 34-41.

[5] Samuda, A. V. ; Harwood, C. F. ; Stockham, J. D. Electron microscope measurement of

airborne asbestos concentrations. A provisional methodology manual. EPA-600/2-77-1 78

revised June 1978; 1978. 49 pp.

[6] Ortiz, L. W. ; Isom, B. L. Transfer technique for electron microscopy of membrane

filter samples. Ind. Hyg. Assoc. J. 35:423-425; 1974.

[7] Zumwalde, R. D. ; Dement, J. M. Review and evaluation of analytical methods for

environmental studies of fibrous particulate exposures. DHEW (NIOSH) Publ. No. 77-204;

1977. 66 pp.

[8] Spiel, S. ; Leinweber, J. P. Asbestos minerals in modern technology. Environ. Res. 2: 166-208; 1969.

[9] Kramer, J. R. Fibrous commingtonite in Lake Superior. Can. Mineralogist 14:91-98;

1976.

66
[10] Nicholson, W. J.; Rohl , A. N. ; Ferrand, E. F. Air pollution in New York City. In: Proceedings of Second International Clean Air Congress. Englund, H. M; Berry, W. T.

,

eds. New York: Academi c Press ; 1971. 136-139.

[11] Cook, P. M. Preparation of extrapulmonary tissues and body fluids for quantitative

transmission electron microscope analysis of asbestos and other mineral particle

concentrations. Trans. N. Y. Acad. Sci. 330:711-724; 1979.

[12] Cook, P. M. ; Smith, P. L. ; Wilson, D. G. Amphibole fiber concentration determination

for a series of community air samples: use of X-ray diffraction to supplement electron

microscope analysis. In: Electron Microscopy and X-ray Applications to Environmental

and Occupational Health Analysis I. Russell, P. A.; Hutchings, A. E. , eds. Ann Arbor

Science Publishers, Inc.; 1978. 107-119.

[13] Peters, E. T. ; Doerfler, T. E. Amphibole mineral fiber analysis by electron

microscopy: Comparison of sample preparation procedures. In: Electron microscopy

and X-ray Applications to Environmental and Occupational Health Analysis I. Russell,

P. A.; Hutchings, A. E. , eds. Ann Arbor Science Publishers, Inc.; 1978. 189-203.

[14] Bishop, K. ; Ring, S. ; Suchanek, R. ; Gray, D. Preparation losses and size alterations

for fibrous mineral samples. Scanning Electron Microsc. 1:207-212; 1978.

[15] Anonymous. Montgomery County Asbestos Study. U. S. Environmental Protection Agency.

Environmental Monitoring and Support Laboratory. Environmental Monitoring Branch.

Research Triangle Park, NC 27711; 1977. 22 pp.

[16] Rohl, A. N. ; Langer, A. M. ; Selikoff, I. K. Environmental asbestos pollution related

to use of quarried serpentine rock. Science 196:1319-1322; 1977.

[17] Beard, M. E. U. S. Environmental Protection Agency. Environmental Monitoring and

Support Laboratory. Research Triangle Park, NC. personal communication; 1980.

67
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gaithersburg, Maryland, October 1-3, 1980. (Issued March

1982)

DEVELOPMENT OF AN ASBESTOS REFERENCE SUSPENSION

Richard S. Feldman 1

Physical Scientist

Drinking Water Research Division

U.S. Environmental Protection Agency

26 West St. Clair Street

Cincinnati , Ohio 45268

Abstract

The objectives were: consistent asbestos fiber' numbers and sizes

per vial, fiber sizes similar to those in field samples, fiber loading

easy to count, no clumps of fibers, no debris, no significant physical

and chemical deterioration. Methods included separation and suspension of fibers in 0.1 percent Aerosol 0T surfactant, fiber length

selection/production by high-power ultrasonication, and uniform dispersal of fibers in surfactant by reciprocal shaking. Vials were used

instead of ampoules for quantitative transfer of the 25 mL contents.

A filtration "buffer" promoted even dispersal of fibers on filters.

Filters were not dried, in order to prevent fiber losses before carbon

coating. Low-power ultrasonic baths may cause separation of fibrils.

The objectives were met.

1. Introduction

There has been growing interest in and concern about the occurrence of asbestos in

drinking water during the past several years. This has resulted in: 1) an increasing

number of laboratories analyzing water for asbestos, and 2) continuing evolution of the

analytical technique. As must be expected with such a relatively new technique that con- tinues to be adopted by more laboratories and undergo modifications in various laboratories,

uniform application of the technique does not exist between laboratories.

An asbestos reference suspension would provide a tool for determining the adequacy of

the presently accepted analytical techniques and for improving the techniques. A reference

sample would also provide a means to evaluate and compare laboratory capability.

The research presented here to develop an asbestos reference suspension, had several

objectives in the following general order of priority:

- consistent numbers of asbestos fibers per vial or ampoule

- consistent fiber size distribution per vial or ampoule

- fiber sizes similar to those found in field samples - fiber loading easy to count and analyze

- no clumps of fibers - no debris - no significant physical and chemical deterioration.

1 Present address: Department of Environmental Sciences, Clark Hall, University of Virginia

Charlottesville, Virginia 22903.

68
1 . 1 Initial Approach

Initially, the goal was to accomplish all of the objectives simultaneously. Because

most of the objectives, except for chemical stability of the fibers seemed to have been

achieved or could easily be achieved (based upon work by previous researchers in this laboratory), the primary emphasis was placed on finding the optimum pH to achieve stability

[l]2 .

The original procedure utilized 2 mg NIEHS #007C chrysotile that was stirred in 0.5 percent Aerosol OT® 3 for 16 hours to separate the fibers. This suspension was ul trasonicated

at 200-300 watts with a Biosonik IV (VWR Scientific) for 30 minutes to break the fibers. A portion of the sonicated suspension was diluted and its pH adjusted; a subset then buffered

at 10.0, 10.5, or 11.0. While the suspension was being stirred, 20 mL aliquots were withdrawn and placed in ampoules. Two sets of ampoules calculated to contain 36 or 40 ng of

chrysotile were prepared. Mercuric chloride was present throughout the procedure as 27.1

mg/L for bacteria control.

The results from using this procedure consisted of: (1) reduced fiber numbers and

increased fiber length when soni cation power was reduced from 300 to 200 watts; (2) the percent diffraction of the fibers being higher at pH 11 than at 10.5; (3) a wide variance in

fiber concentration among the ampoules; and (4) the suspensions containing clumped and

distorted fibers, and debris.

The most critical problems that arose were uneven distribution of fiber numbers and

sizes, and the occurrence of clumps. These problems eliminated the quantitative use of the

reference, and were much more significant problems than reduction of percent diffraction.

(This reduction could have been caused by the low pH Aerosol 0T® leaching magnesium from

chrysotile. Analysis of the filtered solution of one ampoule, five months after preparation,

gave a concentration of dissolved Mg that approximated 5.8 percent of the Mg that should

have been in the chrysotile.) The pH adjuster and buffer solution (NaOH and NaHC03 ) may have

been responsible for some of the problems observed since they change the charge on chrysotile

(the isoelectric point of chrysotile is pH 11.8).

Corrective approaches included the lowering of the pH to 9 and 10, using Teflon® vials

(to avoid possible dissolution of glass), finding an effective alkaline surfactant, and

reducing sonication power and time. Some of these approaches were applied simultaneously.

The Teflon® vials produced a new type of smooth, fibrous debris. The alkaline surfactants

tried were either incompatible with polycarbonate filters (sodium oleate) or ineffective in

separating and evenly suspending chrysotile (Triton RW-100®). Sonication was still too

powerful at 150 watts, since too many fibers were produced and too many were small fibers.

Debris and distorted fibers were also present. Additionally, deterioration of the sonic

probe released titanium particles into the suspension.

1.2 Redirection

Because of the inability to attain the project objectives with the above approaches,

plus the need to use a different model sonicator that had a probe in good condition, major

changes were made in the procedure. All aspects of the original procedure were evaluated

through experimentation, and eventually resulted in a new procedure that produced a satisfactory asbestos reference suspension.

2 Figures in brackets indicate the literature references at the end of this paper.

3Mention of trade names of commercial products does not imply endorsement by the U. S. Environmental Protection Agency.

69
2. Materials and Methods

2.1 Surfactant

Surfactant is used to separate asbestos fibers and to help keep the fibers suspended.

An early reference to the industrial use of surfactants for dispersing asbestos fibers shows

that Aerosol OT® is one of the most effective surfactants [2]. Using relatively concen- trated suspensions, Novak determined the minimum amount of Aerosol OT® necessary to maintain

a stable suspension of a known percentage of chrysotile in the surfactant solution. The

plot he presented can be described as:

Percent Aerosol OT® = (0.0376 x percent chrysotile) + 0.0343.

The reference suspensions contained on the order of 10~ 7 percent chrysotile, so the

first quantity can be ignored. For such dilute suspensions of chrysotile, the percent

Aerosol OT® needed to maintain a stable suspension is therefore 0.0343 percent.

Novak observed that it was necessary to add more surfactant solution to a chrysotile

mixture after several days to maintain fluidity. This indicated that more asbestos surface

was being exposed and interacting with the surfactant. With the possibility of this

increased surfactant demand with time, plus unknown errors in the determination of the

optimum amount of Aerosol 0T® for a dilute reference suspension of chrysotile (as opposed to the industrial slurries), 0.1 percent Aerosol 0T® solution was used throughout the procedure.

Sodium chloride has been used to make Aerosol 0T® more effective in preparing chrysotile suspensions that remained clump-free for at least three months; the solution was 0.1

percent Aerosol 0T® plus 0.1 percent NaCl [3]. The manufacturer of Aerosol 0T® gives data

showing that both interfacial and surface tension of 0.1 percent Aerosol 0T® (and other con- centrations) are reduced by the addition of NaCl at 0.25 and 0.50 percent with no data at

0.1 percent [4]. With Aerosol 0T® of 1.0 percent or greater, these NaCl additions caused

clouding, but it was not stated whether the clouding directly affected the ability or indicated a change in the ability of Aerosol 0T® to wet and suspend solids.

Chrysotile suspensions prepared with 0.1 percent Aerosol 0T® plus 0.1 percent NaCl were

not noticeably different than those without NaCl. A solution of 0.1 percent Aerosol 0T®

plus 0.1 percent NaCl not containing asbestos developed cloudy streaks after a few weeks.

With no clear benefit and the possibility of NaCl disrupting the Aerosol 0T®, NaCl was not

used with this reference.

Distilled, deionized filtered (0.22 urn) water was used for these solutions as well as for washing and rinsing of all equipment, and for all steps in the method that required

water.

2.2 Standard Chrysoti le

The chrysotile used was acquired from the National Institute of Environmental Health

Sciences, Research Triangle Park, North Carolina. This sample is labeled NIEHS #007C, and

is referred to as their short-range chrysotile. The source is Union Carbide Corporation's

mine in the New Idria serpentinite mass in California; the company labels the standard

chrysotile as C0F-25. A detailed description of this material has recently been published

[5].

Weighing the standard chrysotile was the most arduous step in the preparation of the

reference suspension, particularly because sub-milligram quantities were required. A Mettler M5 Microchemical Balance was used to weigh 0.1 mg chrysotile. Aluminum weighing

dishes were used instead of plastic ones to eliminate the problem of static charge

buildup. Weighing was done on days with low atmospheric humidity since chrysotile is

hydroscopic. To allow for a moisture equilibrium to be reached between the chrysotile and

the atmosphere, the vial of chrysotile was opened at least an hour before weighing the

0.1 mg portion. During this period, the stability of the balance was checked.

70
The weighed chrysotile was added to a screw-capped bottle containing 2 L of 0. 1 percent

Aerosol 0T® and a stirring bar. This stock suspension was stirred for 30 minutes, with a 2-3 cm deep vortex. To dilute this suspension to a desirable concentration, 2 percent of

the suspension was withdrawn and diluted. A stirring suspension may have an uneven distribution of fibers, and lead to the withdrawal of unrepresentative aliquots. Small scale

tests in this lab with powdered carbon, bottles of various shapes, and shakers (reciprocating and wrist-action), showed that bottles laid length-wise in a reciprocating shaker gave

the most random distribution of the visible carbon in water. Thus, the 2 percent withdrawal

of suspension was accomplished by shaking the bottle lengthwise in a reciprocating shaker,

withdrawing 20 mL, shaking again, and withdrawing another 20 mL. Shaking time was not

recorded, but was about one minute each time. The 20 mL aliquots were added to a 350 mL wide-mouth jar containing 200 mL of 0.1 percent Aerosol 0T®.

2.3 Soni cation

The 240 mL suspension in the wide-mouth jar was sonicated with a Fisher Sonic Dismembrator, Model 300. The intermediate probe was used, and the unit was operated for five

minutes at the 60 percent setting, after trying higher and lower settings. Calibration of

the sonicator was performed by operating at these same conditions, with the probe immersed

in 240 mL 0.1 percent Aerosol 0T® without chrysotile. The solution was at equilibrium

temperature before sonication, and the temperature was monitored with an electronic thermometer for the five minute sonication. During preparation of the asbestos reference, the

jar was placed in an ice bath during sonication, but during calibration the bath was not

used.

The output of the sonicator was determined by a relation between mechanical and thermal

energy. Assuming 100 percent transfer of the sonic energy to the solution as heat:

3 Increase in thermal energy = (4.2 x 10 joules/KgC°)mAT

3 where 4.2 x 10 joules/KgC° is the specific heat of water, AT is the change in temperature

of the water, and mis the mass of water [6]. Dividing this energy figure by the period of

sonication, in seconds, the power received by the solution is expressed in watts. Assuming

the 0.1 percent Aerosol 0T® has a specific heat that is nearly that of water [7], this

relation yields 15.5 watts or 0.065 watts/mL of 0.1 percent Aerosol 0T®. Using 240 mL of

distilled deionized filtered water, the values were 16.1 watts or 0.067 watts/mL. Not

enough individual calibrations were performed to indicate if the difference between the

values for Aerosol 0T® and water are significant. During the five minute calibrations, 4.5-

4.8 C° temperature changes occurred, with upper limits below 27.3 °C; air temperature

averaged 23.0 °C. Under these conditions there was probably negligible heat loss to the

surrounding environment [8], so the calibration should be reliable.

2.4 Reference Suspension

The 240 mL sonicated suspension was poured into a glass aspirator bottle. The widemouth jar was rinsed with measured volumes of 0.1 percent Aerosol 0T® which were added to the aspirator bottle. Then 0.1 percent Aerosol 0T® was added to create a 2 L volume of

suspension in the aspirator bottle. The bottle had a smooth glass neck that was stoppered

with an aluminum foil-covered rubber stopper. The aspiration port was cut and fitted with

a screw-top neck using epoxy. The cap for the neck had a Tef lon®-coated liner.

The bottle was fastened length-wise into the reciprocating shaker, with the screw-top

neck facing upward. This arrangement allowed for the easy withdrawal of aliquots after

shaking the suspension. During preliminary tests, 25 mL aliquots were withdrawn with a volumetric pipet and filtered immediately. During preparation of the final reference sus- pension, the 25 mL aliquots were placed in vials. The suspensions were shaken for about

one minute before each aliquot was withdrawn.

The vials, caps, and liners were washed in a sonic bath with Aerosol 0T® before using

them for the reference suspension. The liners were rubber-backed Teflon^ Ampoules had

been used for previous references, but they do not allow for the quantitative transfer of

the contents. With 25 mL ampoules, the contents must be shaken out (they cannot be poured),

some of the fluid is generally trapped in the cap, and rinsing is more difficult than with

71
vials. Ampoules are also often difficult to break open safely. The rubber-backed liners

for vial caps create a seal tight enough to replace the melted glass seal of ampoules.

Seventy-two vials were filled and four additional aliquots were taken throughout the

vial-filling procedure and filtered directly. The vials were labeled and numbered according

to the order of filling. Four vials had their contents filtered the day they were filled,

and the rest were boxed and refrigerated. Results from the four vials determined whether

the reference suspension was suitable to send to other laboratories.

2.5 Filtration

During preliminary tests, the 25 mL aliquots of reference suspension were filtered

directly from the pipet rather than from vials. With the final reference suspension the same was done with four aliquots, plus the filled vial to be filtered was sonicated in an ultrasonic bath (50 watts) for three minutes.

A Schleicher and Schuell glass filtration apparatus, holding 47 mm filters, was used. A cellulosic membrane filter is placed on the glass frit as a backing filter for the polycarbonate filter, which actually collects the fibers. The pore size of the backing filter,

is probably not important, though 0.45 urn is the smallest that would normally be used, just

for the convenience of not slowing the filtration. (Further study may reveal that there is an optimum filtration rate to achieve the most even distribution of particles on the

filter.) Another researcher at the Environmental Research Center [9] discovered that an effective way of placing the Nuclepore polycarbonate filter (0.1 urn) to avoid wrinkling and

folding, is to cover the backing filter on the frit with a bubble of water, lay the Nuclepore filter on the bubble, and apply vacuum. The glass funnel is attached firmly, and then

the vacuum valve closed (vacuum is still maintained in the filtration flask for some time).

Prior to adding the reference sample to the funnel, approximately 25 mL of water is

added to the funnel; the reference sample is then added and the vacuum is applied. This

technique was found to give more even distribution of fibers on filters than filtering the

reference without first adding the 25 mL "buffer". The water buffer was also slightly more effective than a 0.1 percent Aerosol 0T® buffer (25 mL). (See section 3.1.) Although this

is somewhat surprising it is more convenient for laboratories to add only a water buffer,

and not Aerosol 0T®.

The vials are rinsed three times with water, and the rinses are added to the filtering

reference. The funnel is covered with aluminum foil during filtration.

2.6 Filter and Grid Preparation

Once the reference sample has been filtered completely, the vacuum is closed, and the

filter is gently lifted to release the remaining vacuum pressure. The filter is not dried,

but is enclosed in a 50 mm diameter tight-fitting petri dish; a dried filter may lose

fibers easier. Petrographic glass slides are conveniently sized for attaching a quadrant of

the filter and a label. The edges of the quadrant are attached to the slide with tape or drops of chloroform. Care is taken to handle the filter gently and to keep it horizontal

to avoid loss of fibers. Two or three slides are placed in a petri dish for transport and

for later storage.

A Denton vacuum evaporator is used in this laboratory to carbon coat the filter quadrants. The evaporator is operated at a constant current of 30 amp and a potential of 60 V for no more than 10 seconds. The slides are not rotated during operation of the evaporator.

The arrangement used to prepare grids is the necessary number of specimen grids (200

mesh) lined up on a stainless steel screen (200 mesh, approximately 3x3 cm) that is laid on a cleaned glass slide. A section of carbon-coated filter measuring approximately 2x2

mm is cut with a cleaned razor blade, grasped on an edge with self-closing tweezers, passed

before a static charge eliminator, and then laid squarely on a grid with the coated side of

the section facing against the grid. A drop of chloroform is released onto the section and

grid with a microliter syringe to seal the section to the grid. This is repeated with all

7 2
the coated filters, usually preparing two grids per filter quadrant. The screen, with the

grids upon it, is then transferred to the chloroform wick.

The chloroform wick used is a modification of the Jaffe wick [10], replacing the glass

slides and filter papers with a polyurethane sponge. The screen supporting the grids is

placed on the sponge. Roughly 24 hours are needed to dissolve the filter material away from

the sections.

2.7 TEM Analysis

The initial analyses of the preliminary reference samples and of the first four reference vials were conducted to determine whether the primary objectives had been met. Therefore, the chrysotile was identified only by morphology, not by crystal structure or elemental composition. The data from these initial analyses also enabled determination of

a statistically suitable number of fibers or grid squares to count.

When analyzing a reference, three distantly spaced grid squares are observed on each of

two grids. A diffraction pattern is attempted on each fiber found. The fiber is sized and

listed as either diffracting chrysotile, non-diffracting chrysotile, or something else,

e.g., amphibole. If the width of the fibers are 0.1 urn or greater they are noted with the

lengths of the corresponding fibers. Irregular chrysotile and debris should be noted. Each

grid square observed is measured to give an accurate determination of fiber concentration.

This measurement is done at low magnification on the TEM. Observation of fibers is best

performed between 14,000 and 20,000 X.

3. Results

3.1 Prel imi nary Test

The preliminary test began with 0.077 mg of chrysotile, which was a quick random

measurement close enough to 0.100 mg. (With the standard chrysotile formed into varioussized small balls, it is difficult to weigh out the exact amount needed in such small quantities.) No vials were used; the aliquots were filtered from the pipet. Three series of

filtrations were carried out, each with three aliquots, i.e., three filters: 1) 25 mL water buffer; 2) 25 mL 0.1 percent Aerosol 0T® buffer, and 3) no buffer. Two grids were prepared and analyzed from each filter.

The data show that filtrations with the water buffer produced the most consistent

resul ts (Tabl e 1 )

.

Table 1. Variation in Chrysotile Count with Three Filtration Techniques

for an Asbestos Suspension.

25 mL 25 mL Water 0.1 percent Aerosol 0T® No Buffer Buffer Buffer

No. grid squares 23 20 25

Fibers/grid square, x 8.91 8.45 8.44

Standard deviation 2.83 3.44 4.27

Range 3-16 3-16 2-17

The fiber numbers per grid square were satisfactory and could be twice as numerous without creating difficulties in counting and analyzing. The fiber lengths ranged from

0.2 urn to about 12.0 urn, and there were adequate numbers of bundles of 0.1 urn and wider;

clumps were absent. The overall appearance of the grids was good. These results encouraged

the preparation of the reference suspension to be distributed to other laboratories.

73
3.2 Final Suspension

The final suspension was prepared with 0.100 mg chrysotile yielding 25 rig per vial or per 25 mL aliquot. During the filling of 72 vials, four additional 25 mL aliquots were

filtered directly from the pipet. Four vials (numbers 5, 23, 39, and 56) were also selected

from the 72 vials and filtered a few days later. Vial number 5 did not produce satisfactory

grid preparation; all the squares were empty, possibly because of a defective filter (the

filters and grids prepared for the four vials were prepared under identical conditions).

The results for the three vials and the direct filtrations are in Table 2.

Table 2. Chrysotile Count in the Final Asbestos Reference Suspension.

Direct Filtration From Vials

Number of filtrations 4 3

Total number of grid squares 28 24

Fibers/grid square, x 9.64 19.25

Standard deviation 3.14 6.97

Range 1-16 12-34

Within the two categories the results were consistent. The overall appearances were

good; only a few clumps occurred, but these had countable numbers of fibers. The vials

yielded twice as many fibers as the direct filtrations. More detailed data is available for

the preliminary test and the final suspension.

4. Discussion

Both the direct filtrations and the filtrations from vials exhibited the desired major

objectives. Debris was minimal, and physical and chemical deterioration of the fibers was

not evident. Only after several months of shelf-life can a real evaluation be made, however.

Realizing that different results may arise between direct and vial filtrations, direct

filtrations were repeated when the final reference sample was prepared to confirm the

results from the preliminary test. The results were similar with mean counts of 8.9 fibers/

grid square for the preliminary test (water buffer) and 9.6 fibers/grid square for the

reference. The difference can be partially attributed to the greater amount of chrysotile

used for the final reference (0.077 mg vs. 0.100 mg).

The discrepancy between the direct filtrations and the vial filtrations may be due to four factors: 1) unrepresentative sampling - this is unlikely because both the direct

filtration and vial filtration aliquots were taken at intervals throughout the filling of

72 vials; 2) contamination of vials - the vials, caps, and liners were scrupulously washed

with filtered Aerosol 0T®, distilled deionized, filtered water, and an ultrasonic bath and

protected, however; 3) time - the vial references were filtered two and a half days after

the direct filtrations; the extra time may have allowed for more separation of fibers in

the vials; 4) ultrasonic bath - the three minute sonication at 50 watts of the vials should

not be enough to break fibers, but may have caused separation of fibers.

During the development of this asbestos reference suspension, discrepancies in asbestos

counts between samples from the same suspension were always accompanied by differences in

fiber size distributions between the samples. This was evident between the direct filtrations and the vial filtrations for this reference, too (Figure 1). The direct filtrations

had more fibers in the longer classes than did the vial filtrations. The mean lengths were

2.6 and 1.8 urn, respectively. The direct filtrations had 23 percent of the fibers with

widths of 0.1 urn or greater, whereas the vial filtrations had a corresponding figure of

20 percent.

74
75
The most reasonable explanations for the size discrepancy is that during the low-level

sonication of the vials, fibers separated, creating the lower percentage of wide fibers.

Because many fibers include fibrils of different lengths, the separation of fibers probably

released many of the smaller fibrils that were attached to longer fibrils.

Despite the apparent changes caused by low-power sonication, it probably has the beneficial effect of resuspending fibers that may be adhering to the vial or may be adhering to other fibers. Unfortunately, there was not time to further test the use of ultrasonic

baths. However, any effect of ultrasonic baths should be constant, from vial to vial, as long as the sonication conditions are the same, e.g. , power output.

The vials were sent to over 20 laboratories for analysis. The decision to send the

vials was based upon the consistency of results within the groups of suspensions analyzed,

and upon the agreement between the preliminary suspension and the direct filtrations of the

final suspension.

A complete set of instructions was sent with each vial as a guide for filtration and

analysis. Data from vial filtrations in this study were used to determine how many grid

squares should be observed by each analyst. With approximately 2.7 x 10 6 fibers per vial 4 and a pooling of the variance for each vial, the analysis of six grid squares for each vial

resulted in a confidence interval of approximately 0.6 x 10 6 fibers per vial. Analysis of

six squares is not an overwhelming task and the confidence interval was acceptable.

The next stage of this study will be to evaluate the data from the laboratories that

analyzed the reference. Initial returns are encouraging.

References

[1] Snyder, P.; McFarren, E. Formerly of the Water Supply Research Division. U.S. Environmental Protection Agency, Cincinnati, OH.

[2] Novak, I. J. Asbestos Dispersion and Method of Forming Same. U.S. Patent 2,626,213;

1953.

[3] Stasny, J. T. ; Albright, F. R. The Separation of Asbestos in Foods, Drugs, and Talc

for Identification and Determination. Structure Probe, Inc. , 535 E. Gay Street,

West Chester, PA 19380; 1979.

[4] American Cyanamid Co. Aerosol" Surface Active Agents. Publication PRT 102, Wayne, NJ.

[5] Cambell, W. J.; et al . Chemical and Physical Characterization of Amosite, Chrysotile,

Crocidolite, and Non-fibrous Tremolite for Oral Ingestion Studies by the National

Institute for Environmental Health Sciences. Bureau of Mines Report 8452; 1980.

[6] Tilley, D. E. ; Thumm, W. Physics for College Students with Applications to the Life

Sciences. Cummings Publishing Co., Menlo Park, CA; 1976.

[7] Hougen, 0. A.; Watson, K. M. ; Ragatz, R. A. Chemical Process Principles. John Wiley

& Sons, Inc. , NY; 1954.

[8] Gerstman, B. Personal communication. Princeton University, Dept. of Physics; 1980.

[9] Pansing, M. F. Formerly of the Health Effects Research Laboratory. U.S. Environmental

Protection Agency, Cincinnati, OH.

[10] Anderson, C. H. ; Long, J. M. Interim Method for Determining Asbestos in Water. EPA- 600/4-80-005; 1980.

4 Filterable area of filter , ... , , =7—r-i r tt x number of fibers counted I(grid square areas observed;

6 2 1075 x 10 urn x 482 fibers = 2 7 x 1Q 6 fl

- bers/vial

1944 x 10 urn
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gai thersburg , Maryland, October 1-3, 1980. (Issued March

1982)

PREPARATION OF AIRBORNE ASBESTOS STANDARDS

David R. Jones and George Yamate

1 1 T Research Institute

10 West 35th Street

Chicago, Illinois 60616

Abstract

The methods used to generate a set of standard hi-vol and personal

samples for use in comparison of analytical methods is described.

Various problems of predicting the filter loading, minimizing external

contamination, and generation of aerosolized fibers are discussed.

1. The Preparation of Laboratory Filters of Controlled Asbestos Loading

1 . 1 Introduction

A statistically designed study to evaluate the electron microscope analytical methodology for determining asbestos required that filters be prepared under controlled

conditions to obtain three asbestos concentrations. Both polycarbonate (Nuclepore) and

cellulose ester (Millipore) filters, with pore diameters of 0.2, 0.4, and 0.8 pm, were

used and samples were collected using high-volume samplers (with 20 cm x 25 cm filters)

and personal samplers (with 3.7 cm diameter filters).

The filters could be prepared in several ways, although it is preferable to obtain

simultaneous sampling for different filter types, pore size, and sampler. The methods

available to prepare filters are:

taking samples close to a natural source, varying flow rate and sampling time;

preparing stock suspensions of known asbestos fiber concentration by

ultrasonic treatment of asbestos in water and filtering from the liquid

suspension(s)

aerosolyzing asbestos fibers and sampling from the aerosol cloud.

Sampling from a natural asbestos source (for example, an asbestos products factory),

would be the most convenient but, unfortunately, it has the serious disadvantage that the

concentration of the source is not known and significant quantities of extraneous particles are also present.

Filter samples can be prepared from liquid suspension(s) of known concentration of

asbestos minerals. The disadvantage of this method is that the deposition of the fibers

from water suspension onto a filter may not be equivalent to that obtained from an aerosol cloud.

Simultaneous sampling from an aerosol cloud of known concentration is the best technique since it simulates normal sampling conditions while allowing control of the aerosol

concentration and filter loading.

77
1.2 Experimental

The Aerosol Chamber--

The aerosol cloud was generated into a spherical chamber fabricated from welded steel

plate with a diameter of 5.5 m and a volume of 86 m

3

. The inside of the chamber is coated

with an epoxy-phenol ic material (Plasite 7122) to prevent corrosion and to provide a smoother finish. The chamber can be cleaned by a hot water spray to wash down the walls,

and a high volume extraction system purges the chamber air through an absolute filter

device at the rate of 12 air changes per hour.

Figure 1 shows the internals. Three high-volume samplers and six personal samplers were mounted on a catwalk located midway on the chamber walls. The aerosol cloud entered

the chamber from the generator located outside the chamber. A fan inside the chamber

circulated air to ensure a uniformly mixed aerosol.

Air Circulating

Figure 1. Top view and side view of aerosol chamber showing location of apparatus.

Ultrasonic Treatment to Break Fibers to a Sufficiently Fine Size--

The UICC asbestos minerals have a very coarse particle size, which is unsuitable for

charging in an aerosol cloud. Three ultrasonic devices were tested to determine their

efficiency in breaking up asbestos into fibers under 10 urn in length. They were:

Ultra-Sonic Industries - System Forty

80 Watts Bath Type;

• Polytron Cell Disruptor - PT10

5000 Watts with High Speed Agitator;

Branson Sonifier - W 185C

100 Watts Horn Type.

Tests were conducted by weighing out a small quantity of asbestos and suspending it

in distilled water to give an asbestos concentration of about 0.3 percent by weight.

Aerosol 0T was added as a dispersing aid at a concentration of about 0.2 percent by

78
weight. Ultrasonics were applied for time periods of 5, 10, 20, and 30 minutes using

each device. Each sample was then diluted to a concentration of 0.03 percent with filtered

distilled water.

The Branson Sonifier was the only unit found suitable for achieving small enough fiber

lengths in chrysotile asbestos. By varying the time of the ultrasonic treatment, the

chrysotile asbestos could be reduced to any fiber length desired. The most satisfactory

chrysotile dispersion was produced by a 45 minute treatment at 100 watts power to 250 mg

of asbestos suspended in 150 mL of water with 2 percent of Aerosol 0T added as a dispersing agent. The quality of the treatments were checked by both optical and electron

microscopes.

The Branson unit was found to be less effective with amosite asbestos and fiber

glass. A series of hand-grinding experiments were performed using an agate pestle and

mortar. A technique was developed which led to satisfactory dispersion of both amosite

and fiber glass. It consisted of wet hand-grinding a 100 mg of quantity of fiber in a few

drops of 1:1 solution of water and Aerosol 0T for 30 minutes.

Aerosol Generation

The Sierra Instrument Company's Model 133G Fluid Atomization Aerosol Generator utilizes

air-blast atomization and inertial impaction to produce aerosols, ii could produce droplets

at rates up to 109 particles per second. The droplet size was variable from 0.03 to 3 urn diameter.

The generator is schematically illustrated in figure 2. It consisted of a dryer, a pressure regulator, an absolute filter, an adjustable valve, two precision flowmeters, a fluid atomizer, an impactor, and an ionizer.

Test

Aerosol

Particle Charge Neutralizer

Filter

Val ve rtXH

Air at 45 T^Tf

Pressure

Regulator

Di lution

Fl owmeter

Atomizer

Atomizer Flowmeter

Impac tor

Figure 2. Flow diagram of aerosol monitor.

High pressure air is supplied to the generator at a minimum pressure of 45 psig. The

air passes through a chemical dryer and a pressure regulator which reduces the pressure to 35 psig. The air then flowed through an absolute filter and was subsequently divided into

two fractions: the atomizer air and the dilution air.

The atomizer air flows through a flowmeter and a Colli son-type atomizer. As the air

passed through the nozzles of the atomizer, it produces a spray of the suspension directed

against a baffle. The spray is then carried by the air through an impactor where large

droplets are removed, leaving an aerosol of a narrow size distribution. The remaining

droplets then flow to a mixing tee located upstream of the ionizer.

After flowing through the filter, the dilution air flows through a manually adjusted

valve. It then passes throug'h a flowmeter and into the mixing tee. From the mixing tee,

the solvent is evaporated and the diluted aerosol flows into the ionizer where it is mixed with bipolar ions. The aerosol then exhausts through the outlet located on the side

79
of the generator housing. Care was taken to adjust the fiber liquid concentration to a point where each droplet formed would contain 0 or 1 fiber the vast majority of the time.

This precaution is required to minimize agglomeration or clumping of the fibers in the

final aerosol. The ionizer employs a radioactive source (1 milli-curie of Krypton 85 gas)

to neutralize any static charge developed on the particles in the generator.

During preliminary runs, contamination of the aerosol by the high-volume samplers was

observed. The brushes of a hi-vol sampler wear rapidly and, as they wear, emit large

numbers of very small carbon particles. The carbon particles deposit on the sample

collection media and interfere with the analysis of the filters. There are two ways to solve this problem:

separate the hi-vol collectors from the motors and bring the motors outside

the chamber;

adjust the chamber concentration to bring the total operating time of the hivols to an acceptable level of brush wear particles.

For the particular set of samples, time and money dictated that we use the second

remedy.

The requirement that high-volume sampling time be kept below a total of one hour,

coupled with the failure of aerosol generators producing more droplets to provide an adequately dispersed aerosol, required a modification of the aerosol generator. Provision

was made to pump asbestos slurry, whose concentration was adjusted to compensate for the

fiber loss and evaporative water loss, into the atomizer unit. The Sierra Aomizer was

thus made operable for periods of 16 to 80 hours on a continuous basis using this make-up

system.

This method is useful when the fiber sizes are small enough to remain suspended

nearly indefinitely with adequate air circulation in the chamber. It produced excellent

relative loadings of single fibers when used with chrysotile asbestos fibers in the size

range found in ambient air. When working with the amphibole asbestiform minerals or fiberglass, the comminution procedures result in a higher level of non-fibrous particles and

the sedimentation rates are significantly higher.

There are two problems associated with the higher sedimentation rates. The higher

sedimentation rate gives rise to a greater uncertainty in the aerosol concentration of

the fibers and some fibers may settle onto filter collection surfaces.

We feel that independent generation of the amphibole asbestos aerosol could minimize

some of the difficulties experienced or a cleaner source of small amphibole fibers could

be used. However, efforts to use other aerosol atomizers of higher capacity to obtain

higher concentrations in a shorter time were ineffective since poor dispersion of individual fibers resulted. Such air samples would be unsuitable as standards for electron

microscopy work.

1.3 Details of Experimental Work in Samples Prepared in Chamber

In all, 27 samples were prepared as detailed in Table 1. Each sample was unique and

a filter was individually prepared.

80
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The filters were prepared in three chamber runs, collecting nine filters per run.

A detailed schedule for each run was prepared in advance using standard engineering techniques. While it is fairly reasonable to assume that removing 2 L/min with several

personal samplers will not produce a significant change in concentration in an 86,000 liter

aerosol chamber, it is also reasonable to assume that 60 min. of hi-vol operation at about

500 L/min, a total of 30,000 L will result in a drop in concentration.

During the study, we first assumed, then demonstrated, that a good estimate of the

concentration at any given time could be made by treating the chamber as a well-mixed

batch reactor. If the chamber is a well-mixed batch reactor, the hi-vol streams become a dilution stream entering the reactor. The mathematical expression describing the concen- tration in the situation is an exponential decay equation whose constant is proportional

to the total instantaneous flow of air through the hi-vol samplers. The flow rates through

the hi-vol samplers were dependent on both filter type and pore size, as seen in the

following measured flow rates, as shown in Table 2.

Table 2. Flow Rates of High-Volume Samplers

Flow Rate (L/min)

Nomi nal

Pore Size Polycarbonate Cellulose Acetate

0.2 651 396

0.4 708 453

0.8 764 679

aThese results are independent of the sampler

actually used, and remained constant for

every sampling run.

Combining the concentration-time equation with two other criteria permitted us to prepare the series of 27 filters in three runs. The two criteria which defined the experimental conditions were:

1. Less than 1 hour hi-vol operating time;

2. Usage of less than 75 percent of the asbestos aerosolized.

The resulting schedules can be seen in Table 3. Table 3 also represents the mass loadings and aerosol concentration in the chamber.

82
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83
The XRF was used as an independent check on the asbestos mass determination. The

results of the mass of chrysotile in a laboratory sample determined by both techniques were

in excellent agreement. EM = 2.725 ug/m3 vs. 2.452 ug/m3 for XRF analysis.

1.4 Problems

There are several problems associated with using aerosol techniques to prepare airborne asbestos standards. The first limitation is that we used a liquid suspension to prepare the aerosol. It is not a fatal objection, but direct suspension of the fibers is

a goal which would help assure minimal modification of the fibers.

Other difficulties in the experimental procedure are created by the broad range of

fiber sizes present. The broad fiber size range imposes limitations and requirements on the equipment. The presence of the asbestos fibrils, which are numerous and weigh little,

required very dilute suspensions and thus a long aerosol ization time. On the other hand,

the relatively huge amosite and fiber-glass fibers require much higher velocities to minimize fiber settling, and the loss of even a few of the largest significantly modify

the mass concentration and mass ratios on the collected filter sample.

When using aerosol techniques to prepare asbestos filter standards, a laboratory is obligated to protect personnel and monitor both personnel and the environment in which

the work is done. In addition, provision to exhaust the aerosol without adding asbestos

to the ambient air must be made. These necessary precautions add to the time and cost of

the preparation.

In our specific experiments, the health and emission problems were already solved,

as the aerosol chamber used was isolated in an area ventilated through a HEPA cleaning

system. The filter system was also piped to permit direct air purging of the chamber.

The design of the experiments was based on the specific goal of preparing filters of

uniform loading in the fiber count ranges needed for the evaluation of Electron Microscopy

Analysis of asbestos fibers. Thus the potential loss of large fibers affecting the precision and accuracy of mass estimates was accepted as an unavoidable risk.

During the development of the technique, it was learned that placing the hi-vol pumps, or at least their motors, directly in the chamber limited our available operating severely.

A better solution to the problem would have been to bring the motors outside the chamber.

This was considered, but due to time, cost, and the ability to the alternate short sample

period to achieve our goals was not done.

The aerosol technique we used provided samples with primarily mono-dispersed fibers

which could be collected on filters. The relative fiber loading and the mass loading were controllable to the extent required by the study. The reproducibility of the filter

loadings was good, and the aerosol chamber was found to approximate a well-mixed reactor

in its concentration behavior through time. Since our quality judgements were based on fiber dispersions and numerical loading, the uncertainty in the mass loading was not as critical a defect as it could have been.

If we were asked to reprepare standards using this basic procedure, the only significant change we would strongly recommend would be to aerosolize any amphibole asbestos

fibers or glass fibers independently using an alternative generator of higher capacity.

The result obtained with chrysotile fibers in the size range suspended in ambient air was quite satisfactory. The actual analyses were obtained using numerous preparatory and

methodology variations, which makes direct comparison of the results meaningless.

84
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gaithersburg, Maryland, October 1-3, 1980. (Issued March

1982)

CONCENTRATION AND SEPARATION OF CHRYSOTILE BY TWO-PHASE LIQUID SEPARATION

Carl W. Melton, Sandra J. Anderson, Carolyn F. Dye, and W. Eugene Chase

BATTELLE

Columbus Laboratories

505 Ki ng Avenue

Columbus, Ohio 43201

and

Charles H. Anderson

Environmental Research Laboratory

Office of Research and Development

U.S. Environmental Protection Agency

Athens, Georgia 30605

Abstract

The development of a rapid analytical method for determining

chrysotile asbestos in water that requires substantially less time per

analysis than electron microscopy methods will be described. Based on the proposition that separation of chrysotile from other waterborne

particulate would greatly simplify the task of detection, the research

effort was directed toward establishing separation and concentration

techniques. This investigation led to the development of a separation

procedure whereby chrysotile is extracted from a water sample into an immiscible organic liquid phase. The procedure is called two-phase

liquid separation (TPLS).

TPLS extracts chrysotile from water into isooctane after the

chrysotile surface has been rendered hydrophobic by reaction with an anionic surfactant (dioctyl sodium sul f osucci nate) . Extraction of the

chrysotile from the water phase into the isooctane phase occurs as the

two liquids are shaken in a separatory funnel. Agitation creates an emulsion that is broken by adding sodium chloride solution. The

isooctane is then filtered to deposit the chrysotile on a filter where

its concentration is analyzed by light microscopy or spot test

procedures

During the course of work to develop a rapid analytical method for waterborne

asbestos, a separation procedure for chrysotile was devised. This makes it possible to concentrate the chrysotile fraction and thus facilitate analysis by eliminating much of the

interfering extraneous particulate. This method selectively extracts chrysotile from the

water suspension into an immiscible organic liquid phase. The procedure is called two-phase

liquid separation (TPLS).

Essentially, TPLS selectively extracts chrysotile from a water sample into a waterimmiscible oil phase, such as mineral spirits, or isooctane, after the surface of

chrysotile fibers is made hydrophobic through a reaction with an anionic surfactant. TPLS

selectivity is based on the difference between the zeta potential of chrysotile and the

zeta potentials of most other waterborne particulate. The surface charge of chrysotile is

positive over a pH range from 2 to 11.4, whereas most other waterborne particulate is

negative over this pH range. The chrysotile, therefore, selectively reacts with the

85
negatively charged anionic surfactant, becomes hydrophobic, and is transferred from the

water phase to the oil phase.

Extraction of the chrysotile from the water phase into the isooctane phase occurs as the two liquids are shaken in a separatory funnel. Agitation creates an emulsion that is

broken by adding sodium chloride solution. The isooctane is then filtered to deposit the

chrysotile on a filter.

The logic behind the approach taken in the development of a rapid analytical method

was guided by an awareness of problems arising from gross amounts of interfering particulate prevalent in most water samples. The sample could be rid of interfering organic matter

by low- temperature ashing, but the inorganic particulate could not easily be eliminated to leave only the asbestos. Analysis either had to be carried out in the presence of both

organic and inorganic particulate or in the presence of inorganic particulate or the

asbestos had to be separated from the interfering particulate. Analysis could be performed

for separated and concentrated asbestos with much less difficulty than for asbestos in the

presence of interfering particulate. Accordingly, our research effort emphasized the

development of a separation method for asbestos in order to facilitate its subsequent

detection. The separation of chrysotile was investigated first because chrysotile is the

most prevalent form of asbestos found in most sampling locations. Also, its surface

chemistry is sufficiently different than most other waterborne particulate to make its

selective separation possible.

1. Effects of Critical TPLS Factors on Selectivity and Percent Recovery

Ths most critical factors in TPLS proved to be (1) mode of agitation, (2) type of

surfactant, (3) concentration of surfactant, (4) number of extractions from a single water

aliquot, (5) ratio of oil to water phase, (6) pH, and (7) the promotion of the reaction

between the chrysotile surface and the surfactant. Although the effects of some factors

overshadowed others, all were found to be important in the optimization of recovery and

selectivity.

The problems of recovery and selectivity were dealt with separately during experimentation primarily because there was no satisfactory procedure to evaluate recovery when

there were large amounts of interfering extraneous particulate in the original water

suspension. Consequently, all recovery experiments were run on standard suspensions of

pure chrysotile.

The general TPLS procedure in its stage of development at the beginning of these

optimization studies consisted of adding anionic surfactant to the water phase, agitating

the water phase and oil phase together, salting out the emulsion that formed upon agitation, and after phase separation, filtering the oil phase to concentrate the extracted

chrysotile on a filter.

2. TPLS Recovery

Effects of Agitation . The mode of agitation is perhaps the most critical of all the

investigated factors. When all other factors are optimized, if agitation is inadequate,

low recoveries are inevitable.

First success was realized using ultrasonic treatment. Later, hand shaking in a separatory funnel produced good results. After consulting the literature, it was decided

to attempt to obtain better control of agitation by substituting mechanical stirring for

shaking by hand. Lai and Fuerstenau reported ^95 percent recovery of alumina from water

suspension into isooctane using mechanical stirring and they were able to use it while

varying other parameters to work toward optimum recovery. The effects of stirring appeared

to be quite reproducible.

86
After switching to stirring, varying recoveries were obtained; some were quite high

but others were low. This variability was attributed to other, possibly unidentified,

parameters when the manipulation of identified parameters did not consistently improve

results.

Mechanical stirring was performed at speeds of 600, 1200, and 2200 rpm and recovery was found to increase with stirring speed. However, stirring speeds of 2200 rpm and above were

impractical. We settled on 1600 rpm, ran 18 experiments under identical conditions, and

analyzed recovery by atomic absorption analysis for magnesium. A mean recovery of 28

percent ±14 was obtained. This degree of recovery was considered to be too low and to have

unacceptable reproducibility. Consequently, a superior means of agitation was sought.

At this point, shaking by hand was tried again and better recoveries were found. A mechanical shaker was used and consistently higher recoveries were obtained than by stirring. Then differences were observed as factors other than agitation were varied. Ultimately, a recovery of 74 percent ±9.6 was obtained using shaking at 150 cycles per minute

for five minutes. This was considered adequate for the purposes of the rapid analytical

method.

Effect of pH . Hydrogen ion activity is not extremely critical in the range from pH

3 to 7, and there is no problem in maintaining the pH in this range during TPLS. However,

to attain the highest positive charge on the chrysotile surface to promote maximum reaction

with the anionic surfactant, the pH of the water phase was adjusted to 3.5 where the zeta

potential has the highest positive value. This is accomplished usually by addition of 0.1 N HC1. In all cases, the pH of water samples has been found to be above 3.5.

Effects of Surfactant Compound and Surfactant Concentration . Only one type of surfactant has been investigated to any extent, this is the anionic alkyl sodium sul f osucci nate. However, two analogous compounds of this type have been investigated. Because of initial success with these compounds, dioctyl sodium sul f osucci nate (Aerosol 0T and M0-70) and

ditridecyl sodium sul f osucci nate (MT-70), no others were sought.

There was some evidence that MT-70 promoted slightly higher TPLS yields than Aerosol

0T, but later experiments indicated comparable recoveries with MT-70 and M0-70, the dioctyl

sodium sul fosucci nate supplied by Mona Industries, Inc., Paterson, New Jersey 07524.

Greater TPLS selectivity was observed while using M0-70. Therefore, M0-70 has been recom- mended in the finalized TPLS procedure.

Surfactant concentration is critical in TPLS, not only from the standpoint of recovery

but also in that too high concentration produces emulsion stability and phase inversion.

Therefore, concentration must be carefully adjusted to obtain acceptable TPLS recovery and

to avoid emul si f ication problems.

While employing MT-70, optimized recoveries were observed to occur at concentrations

ranging from 1 x 10 -3 M to 2.5 x 10 -3

M. No phase inversion was detected at 1 x 10 -3

M, but

it was occasionally seen at the 2.5 x 10" 3 M concentration. Recoveries were determined by

AAS analysis for magnesium at these two surfactant concentrations to determine whether there

was a significant difference in recovery. Twenty-nine determinations were made at

1 x 10~3 M MT-70 concentration and the recovery was measured to be 63 percent ±11. At

2.5 x 10~3 M concentration, it was measured to be 74 percent ±9.6.

Because no further problems with phase inversion were encountered, the higher surfactant concentration is recommended in the description of the TPLS method.

Ratio of Oil to Water Phase . The ratio of the volume of oil phase to the volume of

water phase is important with respect to phase inversion. The oil volume was varied from

25 percent of the total liquid volume to 50 percent (1:1 water-oil). We finally settled on

100 mL of water phase and 50 mL of isooctane. Under these conditions satisfactory interfacial contact occurs and phase inversion is, in most cases, avoided.

87
Promotion of the Reaction Between Chrysotile and Surfactant . It was found that TPLS

recovery was improved by heating the water phase after the addition of the surfactant.

Presumably, heating promotes the reaction between the chrysotile surface and surfactant.

Perhaps one of the mechanisms by which this occurs is the desorption of interfering species

from the chrysotile surface.

Heating at 60 °C is recommended in the routine TPLS method.

3. TPLS Selectivity

Preliminary experiments to investigate TPLS selectivity were run on synthetic mixtures

of chrysotile and diatomaceous earth. Diatomaceous earth was chosen because of the

prevalence of diatoms in water samples. The recommended method is quite selective for

chrysotile in the presence of diatoms.

It is impossible to predict the interferences that might be encountered when applying

TPLS to water samples; however, TPLS has been run on water samples from 26 different

sources. Some interferences were observed, but they were not extremely serious. All were related to lack of TPLS selectivity. Interferences that could give both high and low

results were observed.

High results were obtained on some samples from which organic filaments were extracted

by TPLS. Nevertheless, these filaments can be identified as not asbestos with a little

experience. These filamentous structures are approximately 0.1 urn in diameter and have

varying lengths. One of their identifying characteristics is that along their lengths they

have swellings 0.2 pm diameter and approximately 0.5 pm long. These swellings can be seen

in the light microscope with vertical illumination at 500x magnification after the filtered

TPLS deposit has been carbon coated. No other interferences giving high results were

discovered.

Two types of interferences produce low results. One is an extracted platy mineral

that could obscure chrysotile fibers. These plates yield a diffraction pattern the same as that of talc. In no case was a sufficient quantity extracted to prevent the detection of

chrysoti le.

The other type of interference that will give low results is related to the nature of

the chrysotile surface. If magnesium has been leached from the fiber surface, the zeta

potential becomes negative, hence, it will not react with the surfactant. Also, coatings

may form on the fiber and thus prevent access of the surfactant. Observed in the TEM,

these coatings have low electron densities indicating that they are probably organic.

In the cases in which interference from organic material is a problem, heating after

surfactant addition improves recoveries of coated fibers, but removal of organic fractions

by a chemical treatment possibly would result in further improvement. Because of the time

required, 1 ow- temperature ashing was not considered during the development of the rapid

analytical method. Neither were "wet ashing" techniques explored. It is believed that any

future work should include the investigation of "wet ashing".

4. TPLS Recovery as a Function of Particle Size and Aspect Ratio

Experiments were conducted to determine whether TPLS favors the extraction of certain

chrysotile fiber sizes and aspect ratios. Several groups of experiments have been run investigating TPLS particle size recovery; however, the set of experiments that used

the finalized TPLS conditions are the most pertinent. This last set of experiments also was designed to determine the effects on particle size recovery as a function of zeta

potential

Four samples of chrysotile having different zeta potentials were supplied by Mr. George

Reimschussel of Johns Mansville Research and Engineering Center. Mr. Reimschussel measured

their electrophoretic mobilities as listed below:

88
Sample

96-1

El ectrophoretic Mobility at pH 10

+3.9 MV

96-2 +0.8 MV

96-3 -1.2 MV 96-4 -2.6 MV

No electrophoretic mobilities were measured at lower pH conditions. Nevertheless, they all

have positive zeta potentials at pH 3.5-4. Good TPLS recoveries were obtained in this pH

range. The existence of zeta potential variations among chrysotile types makes it important

to control the pH during TPLS.

Standard chrysotile suspensions were prepared from samples 96-1, 96-2, 96-3, and 96-4,

and equal (50 mL) aliquots were taken from the same suspension. One aliquot was merely

filtered onto a 25-mm, 0. 2-unrpore-si ze Nuclepore filter; three other aliquots from the

same suspension were subjected to TPLS and the extracted chrysotile was also deposited on the same type Nuclepore filters. Specimens for TEM were prepared from the filtered

deposits by carbon coating followed by dissolution of the filter material by the Jaffe Wick

Method.

Specimens were prepared for electron microscopy by carbon coating the filtered chrysotile deposits and dissolving away the Nuclepore filter by the Jaffe technique. TEM fiber

counts and measurements of fiber lengths and widths were made of at least 100 fibers in each

preparation. A computer was used to assign lengths, widths, and aspect ratios to size

classes and to plot each versus cumulative number percent.

The results showed that TPLS favors the extraction of slightly longer fibers with

slightly larger aspect ratios. In this respect, these results are consistent with those

previously obtained under different TPLS conditions.

(2) Seventy-five percent of chrysotile in water suspension can be extracted by TPLS.

(3) TPLS favors the extraction of longer fibers with larger aspect ratios.

(4) Filtration and resuspension is a practical means of concentrating particulate from

large water volumes for TPLS. It is not as feasible to scale up TPLS to handle

large water volumes as it is to filter and resuspend particulate in smaller, more manageable water volumes.

5. Summary of Results

Major findings were as follows:

(1) Percent chrysotile recovery by TPLS is optimized:

~ at pH 3.5 to 5.0

when the oil /water volume ratio is 1:2

when shaking is the mode of agitation

when 2.5 x 10"3 M anionic surfactant concentration is used

when five extractions are made from the same water aliquot.

89
6. Steps in the TPLS Procedure

The following detailed procedure presents step-by-step the instructions for carrying

out TPLS.

Step 1: Shake the bottom of water sample with a mechanical shaker for approximately five

minutes at M80 cycles per minute to create a uniform particulate suspension.

Step 2: Take an aliquot either directly from the water sample, or alternatively, filter

an aliquot into a Metricel DM450 filter.

Alternative 2A : Take a 75-mL aliquot directly from the uniform water sample,

adjust the pH to 3.5 ± 0.1 with 0.1 N HC1 , add 25 mL of 0.01 M MO-70, heat to

140 °F while stirring, and allow to cool to room temperature. If acid additions

overshoot the correct pH value, correct by adding 0.1 N NaOH. Heat at least to

140 °F; heating as high as the boiling point has no adverse effect except to extend the time required for cooling. Cooling may be accomplished by refrigeration or in an ice bath. Also, other than 75-mL aliquots may be taken; then the

amount of surfactant added must be adjusted to maintain the 2.5 x 10"3 M concentration, and the amount of isooctane must be 1/3 of the total liquid volume.

Alternative 2B : Filter a relatively large aliquot (1 liter or more) of water

samples to deposit the particulate on 47-mm Metricel DM450 (0.45 urn pore size)

filter(s) (change filters if necessary as they become loaded to the degree that

the water flow stops). Before the filter deposite have dried, ul trasonically

remove the deposited particulate from the filter(s) in a beaker containing 75 mL

of water to which 25 mL of 0.01 M M0-70 was added to make a total volume of

100 mL. Adjust the pH to 3.5 ± 0.1 with 0.1 N HC1 , heat to 140 F, and allow to

cool to room temperature.

Step 3: Combine the water phase from Step 2A or 2B with 50 mL of isooctane in a 250-mL

separatory funnel equipped with a Teflon stopcock.

Step 4: Attach the separatory funnel to a mechanical shaker and shake at M50 ± 25

cycles per minute for five minutes.

Step 5: Add 10 mL of 10 percent NaCl to break the emulsion. Shake lightly to distribute

the NaCl in the emulsion. Occasionally the isooctane layer will form under the

aqueous layer. Further gentle shaking will eliminate this condition.

Step 6: Remove the separatory funnel stopper during separation of the two liquid layers.

Drain off the water phase through the stopcock. Allow time for the water droplets

to settle and drain them off before pouring the isooctane from the top of the

separatory funnel into the filtering apparatus. It is extremely important to take care that no water is filtered along with the isooctane.

Step 7: Deposit the separated chrysotile onto the filter by aspirating the isooctane

through the f i 1 ter.

Step 8: Combine the same water phase again with 50 mL of isooctane in the separatory

funnel and repeat the extraction procedure without adding additional sodium

chloride, and deposit the extracted chrysotile on the same filter. Repeat the

extraction procedure three more times to make a total of five extractions from

the same water aliquot.

90
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gaithersburg, Maryland, October 1-3, 1980. (Issued March

1982)

ANALYTICAL PROCEDURES AND STANDARDIZATION FOR ASBESTOS FIBER COUNTING IN AIR,

WATER, AND SOLID SAMPLES

Eric J. Chatfield

Ontario Research Foundation

Sheridan Park Research Community

Mississauga, Ontario

L5K 1B3

Abstract

Interlaboratory analyses of air, water and mineral samples for

asbestos fibers have shown much variability. Sources of error in this

type of analysis include fiber losses or size modification during sample

preparation, contamination by extraneous fibers, non-uniform deposition

on analytical filters, differences between operators in fiber counting

philosophy, and use of different criteria for fiber identification. The

lack of suitable reliable standard samples has also confused efforts to incorporate good controls when analytical work has been split between

several laboratories. Interlaboratory distribution of aqueous fiber

dispersions for analysis has been found to be particularly difficult, and

in several studies has resulted in a very wide range of reported

concentrations from the same sample.

The published EPA interim procedures for determination of asbestos

in air and water samples do not specify in detail the topics of fiber

identification or fiber counting philosophy. Morphology, selected area electron diffraction, and energy dispersive x-ray analysis, used either

separately or in combination can provide adequate fiber identification,

depending on prior knowledge about the sample. However, economic

considerations usually prevent classification of every fiber into its

precise mineralogical species. A fiber classification system is proposed which provides a basis for uniform reporting of fiber counting

data; some aspects of specimen preparation and fiber counting techniques

area also discussed.

Key Words: Aqueous standard fiber dispersions; asbestos analysis variability;

fiber identification criteria; interlaboratory calibration;

preparation techniques.

1. Introduction

The control of asbestos in both the workplace and the environment requires reliable

methods of measurement. The precise methods chosen depend upon the particular application. In the atmospheres of workplaces where asbestos is being processed or used, the

membrane filter method [I] 1

, which incorporates fiber counting by phase contrast optical

microscopy, can be used to provide an index of the airborne fiber concentrations. It is,

however, important to recognize the limitations of this approach: the technique is non- specific in that all particles having aspect ratios exceeding 3:1, rather than just

asbestos fibers, are counted, and the resolution is inadequate to detect fibers having

diameters less than about 0.2 pm, whatever their lengths may be.

1 Figures in brackets indicate the literature references at the end of this paper.

91
Detection and identification of asbestos fibers in the environment outside of the

workplace is a rather more difficult problem. A typical air or water sample contains a variety of minerals and fibrous organic debris from which asbestos fibers must be discriminated. Many minerals have good cleavage; elongated fragments of these will certainly

be assigned as asbestos fibers in phase contrast fiber counts, and in some cases their

precise identification may even present problems for electron microscopy techniques.

The transmission electron microscope (TEM) has been applied to measurements on environmental samples, using morphology, selected area electron diffraction (SAED) and energy

dispersive x-ray analysis (EDXA) for fiber identification. Where an absolutely rigorous

demonstration must be made of the precise mineral species, all of these techniques must be

used, and analysis of electron diffraction data from several crystal orientations is

required. Chrysotile is an exception to this rule in that the morphological characteristics

combined with some supporting data by either SAED or EDXA are generally adequate for identification by experienced observers.

The scanning electron microscope (SEM) has limitations of both resolution and contrast

[2], and fiber identification is confined to observations by EDXA of the chemical composition. The SEM-EDXA combination has therefore found little application to the identification

and measurement of asbestos fibers in environmental samples, where there are numerous possibilities of confusion by other mineral species of similar chemical composition [3,4].

2. Sample Preparation Techniques for the TEM

The carbon-coated Nuclepore technique forms the basis of both the Environmental Protection Agency (EPA) interim procedure for water samples [5] and the EPA provisional method for

air samples [6]. The steps in the technique are illustrated in figure 1. After collection

of the sample on a Nuclepore filter a carbon film is applied by vacuum evaporation which

envelops and traps all the particles on the filter surface. A portion of the coated filter

is then placed on a support mesh (no carbon coating), and extracted in a Jaffe washer using

chloroform as the solvent. The sample obtained is a copy, or replica, of the filter

surface, with all of the original particles retained in position. The solvent extraction

does not apparently cause loss of particles [2,7] since absence of a particle would usually

be indicated by a replicated region with no particle inside it. The Nuclepore filter is

selected for this technique, since its surface is relatively featureless apart from the

cylinderical pores. Table 1 shows a summary of results obtained using this technique

compared with those from other preparation methods. The filters analyzed were all prepared

from identical loadings of the same fiber dispersion. It is evident that methods based on solvent washing of unfixed particles on membrane filters were all unsatisfactory, leading

to large particle losses and non-uniform samples.

SOLVENT DISSOLUTION

Figure 1. Steps in the carbon coated

Nuclepore procedure for TEM

specimen preparation.

92
Table 1. Comparison of TEM Sample Preparation Techniques

Chrysoti 1

Preparation

Technique

Carbon-coated

0.1 urn Nucl epore

Jaffe Washer

Uncoated

0. 1 urn Nuclepore

Jaffe Washer

0.45 urn Mi 1 1 ipore

Condensation

Washer

0.45 um

Mi 1 1 ipore

Jaffe Washer

0.22 um

Mi 1 1 ipore

As hi ng/

Coated

Nuclepore

Number of

Repl icate

Samples

26

Mean

Value

10 6 Fibers/L

23.4

12

14

12

11

10

18.5

4.7

9.6

10.0

25.3

95% Conf.

Interval

10 6 Fibers/L

20.0-26.9

13.9-23.2

2.6- 6.

5.9-13.3

7.8-12.3

18.4-32.2

Percentage of

Samples with

Uniform Fiber

Di stri buti on

85

90

50

25

10

100

Fiber

Loss

%

21

80

59

57

Filter uniformity demonstrated at 0.1 percent significance.

^Fiber loss calculated assuming carbon-coated Nuclepore technique to have zero loss.

Assuming that a transfer of the particulate from the original filter to the TEM

specimen has been achieved without losses, and that the material has been retained in more

or less the position it occupied on the original filter, errors in the measurement can occur

in a number of different ways.

(a) The deposit may not have been uniform on the original filter.

(b) Contamination may have been introduced.

(c) Microscopists may differ in the way they count particular arrangements of fibers.

(d) Microscopists may accept different criteria for classification of fibers as asbestos.

In addition, depending on the type of sample, other errors may have been introduced

at earlier stages in the preparation. For example, fibers on an air sample may have been

moved or lost during transportation, and fibers in water samples may have flocculated or been scavenged from suspension by the container surfaces.

Uniformity of the filter deposit can be ensured by use of backing filters [7,8], to provide an even distribution of flow. However, these techniques cannot correct a basic

aggregation or flocculation which may already be present prior to sampling. It has often

been the practice to assume that fibers are deposited on the filter surface according

93
to a Poissonian distribution, and to compute confidence intervals based on this assumption.

Table 2 shows that particularly in the case of chrysotile in water, the assumption of a Poissonian distribution could not be justified, and that a normal distribution of larger

standard deviation was a better fit to the data. Fundamentally, unless it can be demonstrated that there are no interactions between fibers during filtration, there is no reason

to expect a Poissonian distribution. The Poissonian is the most favorable distribution

which can be achieved at low filter loadings in ideal samples.

Table 2. Chrysotile in Water Samples: Goodness of Fit Test for Fiber Distribition on

Fi 1 ters

Chrysoti 1

e

Crocidol ite

(total fibers)

Taconite

(total fibers)

Crocidol ite

(SAED)

X

2 (Poisson) x2 (Normal

)

1189

32. 5

17.2

6.57

15.9

6.8

4.03

8.0

Degrees of Hypothesis

Freedom Accepted

Normal

Normal

Normal

Ei ther

Taconite 1.53 1.78 3 Either

(SAED)

Contamination of samples by extraneous fibers has always been a problem in this type

of analysis. Blank measurements are extremely important.

Fiber counting rules have not yet been defined precisely. This will not be possible

until a decision has been made as to the treatment of fiber bundles and fiber aggregates.

Unfortunately, particularly in the case of air samples, there are often many more fibers

contained in a single loosely held aggregate than the total number of individual fibers

that have been counted in the whole sample. The parallel fiber bundle is likely to be

unopened fiber, and can be treated logically as a single fiber of larger width. The loose

fiber aggregates can be dispersed in water using ultrasonic agitation, but if surfactants

are added, chrysotile fiber bundles will also be dispersed into individual fibrils. The

question of which measurement should be made, or which is biologically relevant, must be

addressed before a solution to the fiber aggregate problem can be found. This aspect is extremely important, since some air samples collected on membrane filters are processed

by ashing, ultrasonic redispersal in water and followed by the carbon-coated Nuclepore

procedure. It would hardly be expected that these would yield the same results as those

of the direct preparation, particularly if the fibers are highly aggregated prior to

col lection.

3. Other Preparation Techniques

There is only one current TEM preparation method which can be realistically compared

in performance with the carbon-coated Nuclepore procedure. The "collapsing membrane"

method [9] is illustrated in figure 2. Essentially the particulate is collected on a conventional mixed-esters sponge-type membrane, after which the filter structure is

collapsed into a continuous film by exposure to solvent vapor. The filter is then carbon

coated, and processed exactly as for the Nuclepore procedure, except that acetone is used

as the solvent.

94
UNUSED MILLIPORE

2 AFTER FILTRATION

3 . COLLAPSED MILLIPORE Figure 2. Steps in the "collapsing Millipore" technique of filter preparation for the SEM

and TEM.

4 . CARBON COATED AFTER^COLLAPSE

5 . CARBON FILM AFTER DISSOLUTION

Methods for selective concentration of asbestos fibers include ashing to remove organic debris [6,7,8] liquid density separation [10] and two phase liquid separation [11,

12]. More advanced techniques based on magnetic separation, ozone-UV oxidation [13] and

high pressure oxygen treatment are under development.

Interlaboratory calibration studies have been compromised by a number of factors,

which are common to both air and water samples.

(a) No consistently reliable air or water standards have been available. In the case

of air samples it is difficult to generate duplicate standards on which the

fiber loadings are uniform. In the case of water standards it has been found

difficult to produce a liquid standard dispersion which yields the same result

after storage for even short periods of time.

(b) Until recently, different preparation techniques were in use in different labora-

(c) No systematic criteria have yet been established for counting of fiber bundles

and aggregates.

(d) Analysts have differed in their interpretation of what criteria constitute

satisfactory identification as asbestos.

Some progress can now be claimed in all of these areas, but (a) and (d) will be

considered in detail.

It has been established that within a single laboratory, replicate filters prepared

from the same dispersion of a single fiber type can yield replicate results using the

carbon-coated Nuclepore procedure [7]. Figure 3 shows an example of the results obtained

when ampoules of an aqueous dispersion of purified chrysotile in double distilled water

were distributed to a number of laboratories experienced in asbestos fiber counting work.

It is clear that in addition to an unacceptable degree of interlaboratory variability, there

was also a significant trend towards lower concentrations when there were longer time

intervals between issue of the samples and preparation of the filters. The results from

the author's laboratory are identified, and these indicate that use of analytical

4. Interlaboratory Calibration and Standardization

tories.

5. Aqueous Standard Fiber Dispersions

95
10

as n

COO

•z. o

J—<S

5

O

2

O

u

ccLUO

u.

O

to>

orX

A O

o

O—* 190 doys

O-*- 190 doys

O-*- 190 doys

J.50 100

INTERVAL BETWEEN AMPOULE PREPARATION AND ANALYSIS , doys

Figure 3. Results of interlaboratory analyses using aqueous dispersion

of Union Carbide chrysotile fibers (Sample 11).

techniques identical in all respects did not allow reproduction of the initial fiber counts

after an extended period of storage.

A large number of these ampoules were originally prepared, and they were studied more closely to determine the reasons for their failure as standards. Ampoules were opened

and the contents filtered directly without the use of any ultrasonic treatment. No chrysotile fibers were detected on the filters, indicating that the chrysotile was no longer in suspension. The glass ampoule was cut open so that the interior surfaces could

be gold coated and examined in the SEM. An interesting feature then emerged from this

examination: many chrysotile asbestos fibers were found attached to the glass surface,

and al 1 of them were associated with some organic material of a gelatinous appearance.

This effect is shown in figure 4. It was quite obvious that the asbestos fibers were cemented to the interior surfaces of the glass ampoule by this organic material, and that

attempts to redisperse them using ultrasonic treatment of ampoules before they were opened were only partially successful. The origin of the organic material in these samples was

at this point unclear.

96
Figure 4. SEM micrograph showing

chrysotile asbestos fibers

attached by organic material to the inside surface

of a glass container.

In a separate study, experiments on the dispersal of chrysotile in very dilute sus- pensions were initiated in order to assess the effects of pH and polyvalent ions which had

been reported by Naumann and Dresher [14]. In the initial experiment, a chrysotile

asbestos dispersion was divided into nine equal volumes in polyethylene bottles. Samples

from three of these were filtered and analyzed immediately. The following treatments of

samples prior to filtration and analysis were studied using two bottles for each treatment:

(a) continuous shaking in a laboratory shaker;

(b) continuous ultrasonic treatment in a bath; and

(c) static storage, followed by treatment for 15 minutes in an ultrasonic bath.

One bottle from each treatment was removed for filtration and analysis after 24 hours, and

the second one after 7 days. The results are shown in Table 3. It is immediately evident

that continuous shaking removed all of the fibers from suspension within 24 hours, whereas

after 7 days in an ultrasonic bath the results were essentially unchanged. In static

storage, followed by ultrasonic treatment for 15 minutes prior to filtration, the results

fell by about a factor of 2 after 7 days. The curious and unexpected results for continuous shaking were thought to be due to collection of fibers by the inside surfaces of the

containers, and re-suspension might be possible by ultrasonic treatment. The bottles

which had been continuously shaken for 24 hours and 7 days were placed in an ultrasonic

bath for a period of 30 minutes, and it was found that the re-suspended fiber concentrations were less than 10 percent of the initial value.

Table 3. Stability of Very Dilute Chrysotile Fiber Dispersions

(Fiber Concentrations in 106 Fibers/Liter)

Treatment

Time Continuous Shaking Continuous Ultrasonic

Static Storage

with Routine

Preparation

Initial 107 100 117

24 hours <0.6 Not Analyzed

78

75

7 days <0.6 51

Experiments were performed with polyethylene bottles containing equal

volumes of the same dispersion, which contained approximately

3000 ng/liter of Union Carbide chrysotile.

97
Throughout this work, no particular effort had been made to maintain sterility in the

suspensions, and it was at this point that the very important part which bacteria and

their decomposition products play in the stability of dilute dispersions of chrysotile

asbestos fiber was recognized. The importance of the electro-kinetic effects of chrysotile in removal of polysaccharides is well known in the beverage and pharmaceutical

industry [15,16] and this lends support to the view that these organic materials may

scavenge chrysotile fibers from suspension and adhere to the container walls.

A new dispersion of Union Carbide chrysotile was prepared, using sterilized glass

containers and double-distilled water taken directly from the condenser outlet of a glass

still. The dispersion was immediately transferred to 50 mL glass ampoules, and these were flame sealed. The ampoules were then autoclaved at 121 °C for 30 minutes in order to sterilize the contents. All of the preparation was completed within one day. Using this

procedure no bacteria or their decomposition products should be present in the dispersion,

and any organisms which may have accumulated during handling would be destroyed, so that

no biological activity should exist in the final ampoules. Table 4 shows the results which

have been obtained using 10 mL samples from these ampoules of the standard dispersion. It can be seen that even after storage for a period of 60 days, the fiber count remained

unchanged, and that constant results were obtained without the use of ultrasonic treatment. The dispersions also showed no change after continuous shaking for periods up to

7 days. However, when an ampoule was opened and the contents contaminated with unsterile

distilled water before being re-sealed, it was found that very variable fiber counting

results were obtained after shaking. The nature and concentration of the organisms added,

however, were not under good control. The results of similar work using UICC chrysotile

are shown in Table 5. These sterile suspensions remained unchanged for long periods, but

when they were contaminated with water containing active organisms the results were again

unsatisfactory. In particular, the unsterile ampoule shaken for 7 days yielded a very

inhomogeneous filter, and the chrysotile fibers were found to be strongly aggregated and

attached to organic debris. In this case a large mass of organic material containing

20 fibers was found on one grid opening, resulting in a very large confidence interval for

the calculated concentration value. Table 6 shows the results obtained to date using

UICC crocidolite. Although this work is incomplete, it appears that stable dispersions

of crocidolite can also be prepared.

Table 4. Stability and Storage of Sterile Union Carbide Chrysotile Asbestos

Fiber Dispersions

(All Concentration Values in 106 Fibers/Liter)

Mean 95% Confidence Interval

Analysis at initial ampoule preparation 82. 6 44. 5

- 121 After continuous shaking for 24 hours 73. 5 47. 1 - 99. 9 After continuous shaking for 7 days 69. 5 51. 5

- 87. 5 After 60 days storage, no ultrasonic) 76. 8 55. 2

- 98. 5 treatment used ) 80. 0 61. 3

- 98. 7 Unsterile ampoule shaken for 24 hours 29. 4 15. 0

- 43. 8 Unsterile ampoule shaken for 7 days 16. 7 4. 4

- 29. 0 Unsterile ampoule shaken for 4 days 62. 4 38. 8

- 86. 4

98
Table 5. Stability and Storage of Sterile UICC Chrysotile Asbestos Fiber Dispersions

(All Concentration Values in 10 6 Fibers/Liter)

Mean 95% Confidence Interval

1Q 1

\ 3 1 11.2- 27. nU 27.9 18.2 - 37. 6 25.

1

15.9 - 34. 3 24.8 12.5 - 37. 0 22.4 15.9 - 28. 9 29.4 21.0 - 37. 8

38.8 0

- 77. 7 6.4 3.3 - 9. 6

Analysis at initial ampoule preparation

After continuous shaking for 24 hours

After continuous shaking for 7 days

After 52 days storage, no ultrasonic \ treatment used ' Unsterile ampoule shaken for 24 hours

Unsterile ampoule shaken for 7 days

Unsterile ampoule shaken for 4 days

Table 6. Stability and Storage of Sterile UICC Crocidolite Fiber Dispersions

(All Concentration Values in 10 6 Fibers/Liter)

Mean 95% Confidence Interval

Analysis at initial ampoule preparation 28.9 15.4 - 42.4

}

After 30 days storage, no ultrasonic I 34.7 26.9 - 42.5

treatment used ) 29.8 23.2 - 36.5

It has now become clear that much of the variability associated with interlaboratory

analyses of liquid chrysotile fiber dispersions was due to the presence of biological

organisms, the decomposition products of which scavenged the fibers and subsequently

adhered to the surface of the containers. The result obtained was then a function of the

extent to which this process had already occurred and the degree to which any ultrasonic

treatment was able to detach and redisperse the fibers. If biological organisms are excluded, and absolute sterility is subsequently maintained, standard dispersions of

chrysotile and crocidolite can now be prepared which appear to be stable for long periods

of time.

6. Proposed Criteria for Identification of Asbestos Fibers

6. 1 Chrysoti 1

e

Identification of chrysotile fibers can be achieved using various combinations of

morphology, SAED and EDXA.

The scrolled structure of chrysotile gives rise to a characteristic morphology in the

TEM, which is seen only in a few other minerals. This morphology is often referred to as "tubular", and in many cases observation of this may be an adequate identification. This

internal morphology of the fiber is not seen in secondary electron SEM images. Although an experienced TEM operator can often distinguish chrysotile fibers from other confusing

materials using morphological observations alone, the technique is too subjective for

routine use without some other means of confirmation.

In the TEM, selected area electron diffraction is capable of precise identification of

chrysotile fibers. Unfortunately, for reasons of instrumental variations, operator techniques, sample history, fiber crystal 1 i ni ty and size, a diffraction pattern on which a confident identification can be made is not always obtained. In many cases, no diffraction

pattern is obtained at all. The frequency with which satisfactory SAED patterns are obtained from chrysotile fibers may be very low; cases are on record where one operatorinstrument combination achieved 90 percent, where another obtained only 10 percent, usi ng

the same sample .

99
In those cases where a satisfactory SAED pattern is obtained, the identification is usually rather simple. Because of the scroll structure of the fiber, there is no requirement to tilt it to a precise angle. The pattern is always the same, and can usually be

identified by inspection at the microscope. Moreover, bundles of parallel fibers give more

or less the same pattern as that from single fibrils.

In the TEM the internal fiber morphology can be used as the primary identification,

with the EDXA spectrum as the confirmatory technique. However, as in the case of SAED,

there are a number of effects which may prevent satisfactory identification of an individual

fiber. The magnesium-silicon ratio varies in fibers obtained from different sources, and

subsequent treatment of the fibers is also capable of changing this ratio. Some of the

magnesium is more loosely bound. In aqueous suspensions, and particularly if these are acidic, magnesium ions may be removed either partially or completely from the structure.

Even in those cases where the magnesium has been completely removed, a silica shell remains

which still has the characteristic internal morphology. The degree of dissolution of

magnesium from chrysotile fibers in aqueous suspensions may be variable from fiber to fiber

in the same sample. Acid leaching studies have shown that the magnesium-silicon ratio of some fibers remained unchanged after exposure to pH values of 3.0 for seven days, whereas

others were not resistant in this way. Moreover, it has been found that a low magnesium

content in an individual fiber is not necessarily related to an inability to obtain a satisfactory SAED pattern. Thus a fiber identified crystallographically as chrysotile by

SAED may contain only a small proportion of the stoichiometric value of magnesium. The

observation that fibers having the crystal structure of chrysotile may have a variable

magnesium content indicates that a magnesium-silicon ratio approximately equivalent to the

published composition cannot be used as an identification criterion for chrysotile.

Chrysotile Fiber Identification Criteria in the TEM

Fibers of chrysotile will be identified primarily by morphology, with supporting

evidence from either SAED patterns or EDXA spectra obtained on a proportion of those

fibers reported. Morphological characteristics required will be:

(a) the i ndi vidua! f ibri 1should have high aspect ratios exceeding 10:1 and be about

40 nm in diameter;

(b) there should be some evidence of internal structure suggesting a tubular appearance similar to that shown in figure 5; (c) the electron scattering power of the fiber at 60-100 kV accelerating potential

should be sufficiently low for internal structure to be visible.

Figure 5. High magnification TEM micrograph of UICC Canadian chrysotile fiber. Note the

symmetrical tubular appearance.

'

1 0.05 um

100
The fiber morphology should be inspected closely, since other minerals such as vermiculite exhibit scrolls (figure 6) which are similar in appearance.

Figure 6. High magnification TEM micrograph showing scroll of

vermiculite which could be

incorrectly classified as chrysotile.

0.05 ym

In order to report the presence of chrysotile, confirmation of the identity of typical

fibers is required either by SAED or EDXA. The requirements for satisfactory identification by these techniques are discussed below.

SAED Confirmation of Identification . An SAED pattern should be obtained which appears

similar to that shown in figure 7. The relevant criteria for identification of this pattern

are indicated on figure 7. Most of these criteria can be viewed directly on the fluorescent

screen of the microscope. Where a satisfactory pattern is obtained, identification can be

made on the basis of visual observation only. However, a photographic record of a typical

pattern should be made, which should also carry a gold ring calibration. The necessary

identification criteria are as follows:

(a) a 7.3 A spacing for the (002) reflections;

(b) a 5.3 A spacing for the layer line repeat distance;

(c) a characteristic "streaking" of the (110) and (130) reflections.

A corresponding SAED pattern from scrolls of vermiculite is shown in figure 8. Although

this might be classified as chrysotile in a cursory examination, it is clear that in addition to other relevant differences the 7.3 A spacing is not present.

Figure 7. Selected Area Electron diffraction pattern

from fiber of chrysotile asbestos. Identification should be made on the basis of 7.3 & 002 spacings, streaking of the 110 and 130

reflections and a 5.3 A layer line repeat

spacing.

101
Figure 8 Selected area electron diffraction pattern from

scrolls of vermicul ite. Measurement of largest

d spacing corresponds to 10.2 A.

EDXA Confirmation of Identification . An EDXA spectrum from a typical fiber showing

peaks from magnesium, silicon and iron only may be satisfactory confirmation of the presence

of chrysotile. However, in many cases peaks from aluminum and calcium also appear in the

spectrum, and the magnesium peak may be of variable magnitude for the reasons previously

discussed. Confirmation by EDXA must therefore be on the basis of quantitative peak area

ratios. Because of differences in detector efficiency and window thicknesses, the values

of peak area ratios obtained from the same material will be different from instrument to instrument. A method of calibration is therefore required, and for simplicity this should

be based on silicate samples of known composition. A substantial simplification of the

procedure will result if these standards are selected to have compositions close to those

of the asbestos varieties.

It is important to determine what values of the Mg/Si ratio should be accepted as con- firmation of the presence of chrysotile. The work of Barbeau [17] indicates that two of

the three magnesium ions in chrysotile are more easily removed by acids than is the third

one. Dissolution of the third magnesium ion requires very drastic treatment. It seems possible that complete destruction of the crystal structure is not achieved until the third

magnesium ion is attacked. Certainly, the variation of magnesium concentration from fiber

to fiber observed in the TEM would support this view. Since it is now necessary to define

a minimum magnesium concentration for classification of a fiber as chrysotile, it would be

logical to establish a permissible range corresponding to more than one magnesium ion in

the structure. The lowest permissible value could be provisionally set as 1/3 of the Mg/Si

ratio obtained using UICC chrysotile.

The concentration of iron in chrysotile is not diagnostic, since a large proportion

is in the form of associated magnetite. Limits on the Fe/Si , Ca/Si and Al/Si ratios must

also be established, if other mineral fragments are to be excluded from the fiber count. Palygorskite, and particularly the Attapulgite variety, can be mistakenly classified as chrysotile on the basis of morphology and a superficial examination of the SAED pattern

at the instrument. This mineral, however, can be discriminated from chrysotile by use of the Al/Si ratio, and also by careful measurements on the recorded SAED pattern.

As a provisional measure, it is suggested that the EDXA confirmation criteria should

be as fol lows:

i) Mg/Si ratio greater than 1/3 of that from UICC chrysotile;

ii) Al/Si and Ca/Si ratios smaller than 0.05, unless it can be shown that the aluminum

and calcium originate from extraneous particulate close to the fiber being analyzed;

iii) Fe/Si ratio not more than double that obtained from UICC chrysotile.

In a sample about which nothing is known, suspected chrysotile fibers can be classified as in Table 7.

102
Table 7. Classification of Fibers with Tubular Morphology

TM Tubular morphology, similar to but insufficiently

characteristic for classification as chrysotile

CM Chrysotile by morphology only

CD Chrysotile by SAED

CX Chrysotile by EDXA

CDX Chrysotile by both SAED and EDXA

6.2 Amphiboles

Unlike chrysotile, the amphiboles are of very varied chemical composition [18,19].

The lattice parameters are also variable within the species, depending on the composition.

Amphibole is a very common constituent of the earth's crust and particles of it will be

encountered frequently in environmental samples.

Amphibole has a pronounced cleavage parallel to (110). Because of this, crushed

fragments of amphibole tend to be elongated, needle-shaped particles. These cleavage

fragments are brittle and do not display the flexible fiber and strength characteristics

of asbestos. Amphibole asbestos does not display the prominent (110) cleavage, and

generally the fibers have higher aspect ratios than the cleavage fragments of the non- fibrous varieties. Studies on methods for discriminating between individual small particles

of the fibrous and non-fibrous forms are in progress, but no simple method yet exists,

apart from the observation that the cleavage fragment will often lie on a (110) face. It may be possible to distinguish a popul ation of particles from a population of fibers on the basis of the distribution of aspect ratios. Proposals to increase the aspect ratio

from the current 3:1 to 10:1 for definition of a fiber are based on the observation that

the mean aspect ratio is much higher for fibers than for cleavage fragments. In the

absence of any other method of discrimination, this proposal has some merit and is

supported by many mineralogists.

The aspect ratio to be chosen should preferably have some mineralogical or biological

relevance, rather than be another arbitrary value such as that currently in use. Unfortunately, any change from the current 3:1 value would receive substantial opposition from

many legislative bodies, whose past records are based on this. Moreover, there seems to be inadequate evidence that cleavage fragments in the aspect ratio range of 3:1 to 10:1

do not have health effects similar to those of the fibrous mineral in the same aspect

ratio range. The model of Pott [20] logically postulates that the carcinogenic potential

is a continuous function of fiber size, and although the health effects of low aspect

ratio fibers may be assigned in this model to be lower than those of the high aspect ratio

fibers, they are not considered to be actually zero. In summary, a recommendation to increase the aspect ratio to 10:1 for definition of a fiber would currently have to be

justified on the basis of analytical expediency alone. Selection of any other value

between 3:1 and 10:1 would be an arbitrary compromise.

It is recognized that confidence in classification of a particle as a fibrous variety

increases as the aspect ratio increases. The most convenient approach which is currently

available to specify a fiber population to be either "asbestos" or "non-asbestos" is probably to examine the aspect ratio distribution according to the technique of Wylie

Proposed Amphibole Identification Criteria

Amphibole particles and fibers viewed in the SEM or TEM display no characteristic

morphology on which discrimination from most other minerals can be based. Morphological

selection for further examination can only be made on the basis of aspect ratio, combined

with observations of cleavage. Any additional characteristics such as evidence of flexibility and fibrillated ends add confidence to the classification as a fibrous mineral, and

[21].

103
should be noted. Without any further identification, an individual elongated particle should

be classified as an "unidentified mineral fiber". Because of the large amount of time

required to perform an unequivocal identification, several degrees of identification will be

defined in order to permit the analysis to be economically viable. It would obviously be

desirable to identify each fiber completely, but this can only be achieved by some hours

of labor expenditure on each fiber. Moreover, not all of the equipment or identification

techniques may be available, and it is important that the method of identification be

stated. However, if identification terminology is defined, there will be a minimum of

confusion.

Terms for the identification techniques are defined below:

i) Routine SAED

SAED patterns obtained at random fiber orientations, without tilting of the

specimen to align a crystal lographic zone axis parallel to the microscope optic axis.

Figure 9 shows a typical amphibole pattern obtained.

o

5.3A

Figure 9. Amphibole SAED pattern (crocidol ite)

obtained from a fiber without precise

orientation on to a zone axis.

ii) Routine EDXA

Examination of the EDXA spectrum for the presence or absence of peaks, without

detailed quantitative interpretation.

iii) Zone Axis SAED

Precise tilting of the fiber using a goniometer stage to obtain an SAED pattern

along a crystal lographic zone axis. Dual Zone Axis SAED can also be performed, in

which two different

between the patterns

ture. Some typical

shown in figure 10. calibration standard.

zone axis patterns are obtained from the fiber, and the angle

is also examined for consistency with the test amphibole strucsatisfactory zone axis patterns for this type of analyses are These also include rings from a thin film of gold as an internal

Figure 10. Selected area electron diffraction patterns from

two different zone axes in anthophyl 1 ite.

104
Amphibole Fiber Identification Criteria in the TEM

As previously discussed, morphology should be used to select mineral fragments for

further study. A search of the sample should be conducted of the sample for particles

having aspect ratios greater than 3:1, with parallel or stepped sides. These should, in

the absence of other background information, be classified as "unidentified mineral

fibers". Obvious material of biological origin is excluded at this stage.

Further identification using either routine SAED or EDXA may not be possible. No recognizable SAED pattern may be obtained because of unsuitable fiber orientation. For

example, it has been shown that only about 50 percent of amphibole fibers in random axial

orientations yield a pattern which the operator can recognize as similar to that in figure 9. Using EDXA, the x-ray emission may for some reason not be able to reach the

detector. In general, however, one or other of the techniques will be useful. The proposed

classification scheme is shown in Table 8, for the compl etely unknown sampl e situation .

Table 8. Classification of Fibers Showing No "Tubular" Morphology

Identification Criteria

Aspect ratio > 73 : 1 and

parallel or stepped sides

Routine SAED pattern

obtai ned

EDXA spectrum obtained

similar to that from

standard amphibole

asbestos samples

Quantitative interpretation of EDXA spectrum,

data available as mass percentages

Routine SAED and

EDXA spectrum

One zone axis SAED

pattern consistent

with amphibole, and

EDXA spectrum

Two zone axis SAED

patterns, consistent

with amphibole, and

angle between them is

also consistent

Code Classification

UF (Unidentified Mineral Fiber)

AD (Amphibole by routine SAED)

AX (Amphibole by routine EDXA)

AQX

(Amphibole by quantitative EDXA)

ADX (Amphibole by both SAED and

EDXA)

AZX (Amphibole by zone axis SAED

and EDXA)

AZZ (Amphibole by dual zone

axis SAED)

Unidentified mineral fiber

(Suspected amphibole)

Possible amphibole

Possible amphibole

Probable amphibole

Probable amphibole

Confirmed amphibole

Confirmed amphibole

In a sample about which nothing is known, the classifications in Table 8 are the most

definitive identifications that can be made. The routine SAED pattern showing a 5.3 A spacing shows only that a chain silicate may be present, or it may possibly be a layer

silicate. Some clay minerals also give this spacing, e.g., palygorskite. Use of the rou- tine SAED technique by itself is more a rejection technique than an actual identification,

since obviously non-amphibole patterns are often obtained. Similarly, the EDXA spectrum

showing a peak from silicon, with various combinations of peaks from sodium, magnesium,

iron, and calcium is not sufficiently specific for identification of amphibole; a number of

other minerals have these approximate compositions. Combined routine SAED and EDXA

105
observations, or quantitative compositions calculated from EDXA spectra, give some additional confidence that the fiber is amphibole, but it is not absolutely certain. The use

of zone axis SAED patterns, with computer analysis to determine their consistency with an amphibole structure, is the only way in which the presence of amphibole can be positively

conf i rmed . Either one pattern with a routine EDXA spectrum, or two patterns with a known

rotation between them along the fiber axis, should be the minimum criteria for positive

confirmation. This also allows some discrimination to be made between individual amphiboles. Complete discrimination between the individual amphiboles requires matrix-corrected

EDXA measurements so that accurate compositional data are available, in addition to the

crystal 1 ographic data.

When amphibole is known to be present in the sample, either from previous knowledge of

the source or from a zone axis SAED measurement, this gives much more confidence that the

remaining morphologically and compositional ly similar fibers initially classified as "possible" and "probable" amphibole are actually amphibole. The same situation applies if the

sample being analyzed is one of a series from the same experiment or location, in which

amphibole has been completely identified in some of the samples, and in which there is no reason to suspect a change of mineral type between samples. Under these circumstances the

"possible" and "probable" categories should be reclassified as "confirmed amphibole". The

unidentified mineral fibers, i.e., those which have not been rejected either by observation of an obviously non-amphi bol e SAED pattern, or by an obviously non-amphibole EDXA

spectrum should then be reclassified as "probable amphibole".

This work was supported by the U.S. Environmental Protection Agency, Analytical Chemistry Branch, Contract Number 68-03-2717, and by the Ontario Ministry of Industry and

Tourism. The author wishes to express appreciation for the financial support of both

sponsors

.

References

[1] Leidel, N. A.; Bayer, S. G. ; Zumwalde, R. D. ; Busch, K. A. USPHS/NIOSH Membrane Fi Iter

Method for Evaluating Airborne Asbestos Fibers. U.S. Dept. of Health, Education,

and Welfare, National Institute for Occupational Safety and Health, Cincinnati, Ohio;

1979 February.

[2] Chatfield, E. J.; Dillon, M. J. Some Aspects of Specimen Preparation and Limitations

of Precision in Particulate Analysis by SEM and TEM. SEM/1 978/1 , 0. Johari , ed. SEM

Inc., AMF O'Hare, Chicago, IL 60666; p. 487-496.

[3] Champness, P. E. ; Cliff, G. ; Lorimer, G. W. The Identification of Asbestos. J. Microsc. 108:231-249; 1976.

[4] Ruud, C. 0.; Barrett, C. S. ; Russell, P. A.; Clark, R. L. Selected Area Electron

Diffraction and Energy Dispersive X-ray Analysis for the Identification of Asbestos

Fibres, a Comparison. Micron 7(2): 1 15-132; 1976.

[5] Anderson, C. H. ; Long, J. M. Interim Method for Determining Asbestos in Water. Report

EPA-600/4-80-005, National Technical Information Service, Springfield, Virginia 22161;

1980 January.

[6] Samudra, A. V.; Harwood, C. F. ; Stockham, J. D. Electron Microscope Measurement of

Airborne Asbestos Concentrations. Report EPA-600/2-77-178. U.S. Environmental

Protection Agency. Research Triangle Park, NC 27711; revised 1978 June.

[7] Chatfield, E. J.; Glass, R. W. ; Dillon, M. J. Preparation of Water Samples for Asbestos Fiber Counting by Electron Microscopy. Report EPA-600/4-78-011 . National Technical Information Service. Springfield, Virginia 22161; 1978 January.

106
[8] Chatfield, E. J. Measurement of Asbestos Fibres in the Workplace and in the General

Environment. Short Course Handbook. Mineralogical Techniques of Asbestos Determination, Vol. 4. R. L. Ledoux, ed. Mineralogical Association of Canada, c/o Department

of Mineralogy, Royal Ontario Museum. 100 Queen's Park, Toronto, Canada M5S 2C6.

p. 111-163; 1979 May.

[9] Ortiz, L. W. ; Isom, B. L. Transfer Technique for Electron Microscopy of Membrane

Filter Samples. American Industrial Hygiene Assoc. J. 35(7):423-425; 1974 July.

[10] Bonner, W. P.; Bustamante, R. B. ; Leimer, H. W. ; Scardina, J. T. Identification of

Asbestos-type Materials in Suspended Solids. Research Report 61. Water Resources

Research Center. University of Tennessee, Knoxville, TN 37916; 1977.

[11] Melton, C. W. Progress in the Development of Rapid Extraction Methods for Chyrostile

Asbestos in Water. Symposium on Electron Microscopy of Microfibers, F.D.A. U.S. Government Printing Office. Washington, D. C. 20402. p. 166-169; 1976 August.

[12] Melton, C. W. ; Anderson, S. J.; Dye, C. F. ; Chase, W. E. ; Hef f el f i nger , R. E. Development of a Rapid Analytical Method for Determining Asbestos in Water. Report EPA-600/

4-78-066. National Technical Information Service. Springfield, Virginia 22161;

1978 December.

[13] Chatfield, E. J. Preparation and Analysis of Particulate Samples by Electron Microscopy, with Special Reference to Asbestos. SEM/1979/I, 0. Johari , ed. SEM Inc., AMF 0'Hare, Chicago, IL 60666; p. 563-578.

[14] Naumann, A. W. ; Dresher, W. H. Colloidal Suspensions of Chrysotile Asbestos: Surface

Charge Enhancement. J. Coll. Interface Sci . 27(1 ): 133-140; 1968.

[15] Wilke, H. ; Voss, H. E. The Removal of Pyrogenic Substances from Injection Solutions by . Filtration. Arzneim-Forsch. 4:8-14; 1954.

[16] Kienholz, M. Experiments with a Plant for Sterilization, De-mineralization and De- pyrogenization of Water. Arzneim-Forsch. 14:1259-1262; 1964.

[17] Barbeau, C. Evaluation of Chrysotile by Chemical Methods. Short Course Handbook.

Mineralogical Techniques of Asbestos Determination, Vol. 4. R. L. Ledoux, ed. Mineralogical Association of Canada, c/o Department of Mineralogy, Royal Ontario Museum.

100 Queen's Park, Toronto, Canada M5S 2C6. p. 197-211; 1979 May.

[18] Deer. W. A.; Howie, R. A.; Zussman, J. Rock Forming Minerals. Vol. 2. Chain Silicates. Longmans, London, England, p. 379; 1963.

[19] International Mineralogical Association. Nomenclature of Amphiboles. (Compiled by

B. E. Leake). Can. Mineral. 16:501-520; 1978.

[20] Pott, F. Some Aspects on the Dosimetry of Carcinogenic Potency of Asbestos and Other

Fibrous Dusts. Staub-Reinhalt, Luft 38:486-490; 1978.

[21] Wylie, Ann G. Fiber Length and Aspect Ratio of Some Selected Asbestos Samples. Health

Hazards of Asbestos Exposure. Irving J. Selikoff; E. Cuyler Hammond, eds. Annals of

the New York Academy of Science 330:605-610; 1979.

107
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gaithersburg , Maryland, October 1-3, 1980. (Issued March

1982)

DEVELOPMENT OF A RAPID SURVEY TECHNIQUE FOR THE DETECTION OF ASBESTOS FIBERS

Peter Riis and Eric J. Chatfield

Ontario Research Foundation

Sheridan Park Research Community

Mississauga, Ontario

Abstract

Amphibole and chrysotile asbestos fibers adopt preferred alignment

directions when suspended in a strong magnetic field. The alignment

direction of the fiber may be parallel to, normal to, or at a specific

angle to the flux lines. When an aligned distribution of fibers is

illuminated by a collimated beam of light, scattering occurs in which

maxima of intensity are observed in all directions normal to the fiber

lengths.

The magnetic alignment and light scattering technique is being

developed for the detection of asbestos fibers. The current procedure

consists of filtration of a liquid sample in a magnetic field of 1.0

Tesla, during which the fibers become aligned before they come into

contact with the filter. The mixed cellulose ester filter is mounted on

a glass slide and its structure is collapsed by exposure to acetone

vapor. The cleared membrane filter is removed from the slide and

illuminated by a normal incidence laser beam. The scattered light

intensity is measured as a function of the angular position of the

original magnetic field direction, and the peaks which occur allow the

concentration of aligned fibers to be deduced. This method shows much

promise as a rapid survey technique for the presence of asbestos fibers

in water samples. It is also possible to distinguish between chrysotile

and amphibole asbestos fibers, and even between some specific amphibole

varieties on the basis of differences in the profiles of their scattered

1 ight di stributions.

Key Words: Asbestos fibers; magnetic alignment; magnetic filtration;

light scattering; rapid fiber analysis.

1. Background

Current methods for the measurement of asbestos in water samples require the use of an electron microscope by a skilled operator for a time of typically three hours per sample.

This procedure has the disadvantages of being slow, requiring expensive equipment, and it is

also subject to operator subjectivity. A method which could be used to screen samples rapidly for asbestos content would be attractive. Thus if samples were shown to have asbestos

concentrations below a specified threshold value they could be eliminated from further

analysis. This would result in substantial economies in the effort required for monitoring

of water supplies. Desired features of such a method would include the following:

(a) a requirement for substantially less labor than is demanded by current electron

microscopy techniques;

(b) routine analysis without requirement for a high degree of skill;

(c) it should not require expensive equipment or a clean room environment;

108
(d) it should not rely on fiber counting techniques, either by electron microscopy

or by light microscopy;

(e) it should be sufficiently sensitive to detect 2 x 10 s fibers/liter or 1 nanogram/

1 iter of asbestos

;

(f) it should be able to distinguish between chrysotile and amphibole asbestos.

This paper describes the results of progress to date by the Electron Optical Laboratory of

the Ontario Research Foundation to develop such a method.

Previous attempts have been made to develop a rapid measurement technique for

asbestos. Birks et al . [I] 1 used electric fields to align chrysotile fibers and then

measured the quantity by x-ray diffraction analysis. The sensitivity of the technique was

0.2 ug in the absence of other particulate. Melton et al. [2] developed the technique of

two phase liquid separation for the separation of chrysotile asbestos from water samples.

While this method removed other particulate giving a much cleaner sample, it still relied

on light microscope counting techniques to obtain a 1 ng/L sensitivity. Another method

developed by Diehl et al. [3] was based on single particle light scattering using a focused laser beam. This method is specific for distinguishing asbestos fibers from other

particulate, but it requires a complete initial characterization of the water source by

electron microscopy for calibration of the output. Any fluctuation in the relative

proportions of different types of particulate may lead to an erroneous result. The

possibility of error, and the requirement for prior TEM characterization of the sample make

it unsuitable as a rapid screening technique. The approach being developed at ORF follows

that described by Timbrel 1 [4], namely the alignment of asbestos fibers by magnetic fields,

followed by analysis of the scattered light from the aligned fibers.

2. Magnetic Alignment of Asbestos Fibers

When asbestos fibers are placed in strong magnetic fields (about 1.0 Tesla(T)) they

become aligned in three possible modes as illustrated in figure 1. Depending on the type

of fibers, and in some cases their origin, they may align parallel to the field direction

(P-type), normal to field (N-type), or transversely to the field at a constant angle

(T-type). Figure 2a is a phase contrast optical micrograph of a dispersion of Union Internationale Contre le Cancer (UICC) crocidolite; figure 2b shows the same dispersion but in

this case the sample was prepared in a magnetic field of 1.0 T. In the case of crocidolite,

the majority of the fibers align parallel to the magnetic field and a smaller number align

in directions perpendicular to the field. Figure 2c shows a phase contrast optical micrograph of an aligned UICC amosite dispersion, which illustrates that for this material there

is a greater proportion of N-type fibers. Figure 2d shows the alignment effect observed

with UICC Canadian chrysotile. The fibers are P-type, but the fibers are curved, and there

are larger deviations from precise alignment than is the case with the previous two

materials. New Zealand cummi ngtoni te which contains T-type fibers with their alignment

directions symmetrically disposed about the field direction are shown in figure 2e.

1 Figures in brackets refer to the literature references at the end of this paper.

109
Figure 2. Phase contrast optical

micrographs of:

a) unaligned UICC crocidolite;

b) aligned UICC crocidolite;

c) aligned UICC amosite;

d) aligned UICC Canadian

chrysoti le;

e) aligned New Zealand

"cummingtonite.

(The magnetic field direction

is indicated on the figures).
The alignment of asbestos fibers in magnetic fields is due to the paramagnetism or dilute ferromagneti sm of the fibers. In amphiboles, the P-fiber alignment mode is a consequence of the direction of maximum magnetic susceptibility being parallel to the length

of the fiber; for N-fibers it is normal to the length of the fibers and for T-fibers it is at a constant angle to the fiber length. It has also been suggested that the alignment

behavior of iron-containing amphiboles is due to inclusions of magnetite [5]. Transmission

electron microscope (TEM) examination of amosite fibers by Cressey and Whittaker [6] has

not revealed any such inclusions. This suggests that the explanation of the alignment

effect probably lies within the structure itself. They have studied the crystal lographic

orientation of aligned amphibole fibers by selected area electron diffraction. In the case

of N-type UICC amosite fibers they found that the crystal y-axis is oriented within ±20°

of the field direction. This is explained if the axis of greatest magnetic susceptibility

is parallel to the y-axis and further supports the idea that the alignment effect is due

to the crystal 1 ographic structure of the fiber. Angular restrictions about the z-axis of

P-type fibers were also found but were of a much broader range. No explanation for the

differences between P-type and N-type amosite fibers have so far been found. Analyses of

their chemical compositions by energy dispersive x-ray analysis have shown no significant

differences. In the case of chrysotile, Timbrell [4] has suggested that the fibers align

in a direction parallel to the field because of magnetite particles in the fiber. This has

not been investigated.

3. Preparation of Aligned Fiber Samples

A simple technique for the preparation of aligned fiber samples from aqueous suspensions is to add one percent by weight of agar to the dispersion and gently heat until the

agar has dissolved. A small volume of the dispersion is then placed on the surface of a glass microscope slide which is located in a magnetic field of approximately 1.0 T. The

liquid dispersion is left to solidify in the magnetic field, resulting in a permanent film

which contains aligned asbestos fibers. This technique is useful for demonstration of the

alignment effect, and was used for preparing the samples shown in figures 2 a, b, and c. It is not suitable, however, for the development of a rapid screening detection method

since it requires the sample in the form of a highly concentrated dispersion.

The sample preparation method currently under investigation at ORF is based on "magnetic filtration". In this technique a 2.5 cm diameter glass Millipore filtration

assembly is located between the poles of an electromagnet, and a non-magnetic clamp is used

to attach the filter reservoir. The filtration apparatus is illustrated in figure 3. Fibers become aligned as the liquid passes through the magnetic field and when collected on the filter surface they retain their orientation. This technique has the advantage of

allowing concentration of the fibers from a known volume of liquid onto the active area of

the filter. The complete procedure is as follows.

(a) With a 0.22 urn pore size type GS Millipore filter mounted in the filtration

assembly, the aspirator is turned on and the magnetic field adjusted to the

desired value (typically 0.8 to 1.0 T).

(b) The desired volume of liquid is filtered through the assembly. For small

volumes of liquid the filtration rate can be adjusted by changing the applied

vacuum. A 20 ml_ volume of suspension is usually filtered in about two

minutes. This ensures that the fibers have had adequate time to become

aligned before contacting the filter surface.

(c) The magnetic field is then turned off.

(d) A mark is made on the edge of the filter nearest to one pole piece of the

magnet. This provides an indication of the field direction after the filter

has been removed.

(e) The filter is removed and dried for approximately 15 minutes at 70 °C.

Ill
Figure 3. Equipment used for magnetic filtration.

(f) A clean glass microscope slide is used as a substrate for the filter. This

is first dipped in a 20 percent solution of collodion in ethanol and allowed

to stand for about 30 seconds, after which the surface film has become

viscous. The filter is placed on to this surface film with a rolling

action.

(g) The filter is then collapsed by exposure to acetone vapor, either by the Ortiz

and Isom [7] technique or by that of the Asbestos International Association

(AIA) [8].

If the sample is to be permanently mounted onto the slide, the edges of the filter should be

sealed to the slide using clear nail polish or a similar lacquer. If the filter sample is to be used for scattered light analysis (described in the following section), the filter is carefully removed from the slide after cutting one edge to release it. This results in a sample consisting of a clear plastic membrane which contains aligned fibers, and which can

be supported in a holder for scattered light analysis. Although the sample can be removed

from the glass slide immediately after clearing, it generally peels from the surface easier

if it is stored for about 24 hours after it has been mounted.

If aligned samples are to be prepared for TEM analysis, dispersions are magnetically

filtered using 0.1 urn pore size Nuclepore filters, and TEM grids are prepared by the

conventional carbon-coating Jaffe washer technique.

112
4. Analysis cf Aligned Asbestos Fibers by Light Scattering

When a beam of light is used to illuminate a fiber, light is scattered perf erenti ally

in directions perpendicular to the length of the fiber. For a distribution of randomlyoriented fibers this results in random light scattering in all directions. This is illustrated in figure 4a, which shows the scattered light distribution for the unaligned

dispersion of UICC crocidolite fibers of figure 2a. Figure 4a was obtained by photographing

the scattered light when a laser beam was used to illuminate the sample. When a fiber

distribution has been aligned in a magnetic field, the scattered light distributions from

individual fibers are also aligned, and this results in a maximum in the scattered light

intensity in directions perpendicular to the length of the fibers. This is illustrated in figures 4b to 4e for magnetically-aligned distributions of crocidolite, amosite, chrysotile,

and New Zealand cummi ngtoni te respectively. Most of the crocidolite fibers align parallel

to the magnetic field, resulting in sharp scattering peaks perpendicular to the field

direction, but small N-fiber peaks are also visible. Amosite contains large quantities of

both P-type and N-type fibers, resulting in scattering maxima both perpendicular and

parallel to the magnetic field. Chrysotile fibers align parallel to the field direction

but because the fibers are often curved, there is incomplete alignment and the peak in the

scattered intensity is much broader. New Zealand cummi ngtoni te contains fibers which align

at a constant angle to the field (T-type), giving rise to the "X" pattern of figure 4e.

Figure 5 shows schematically the equipment used for the analysis of the scattered

light from aligned fibers. A beam of light from a laser is used to illuminate the aligned

fiber dispersion. The fiber dispersion is rotated about an axis coincident with the center

of the beam. A photomultipl ier detector is mounted at an angle 0 to the incident beam with

its center of rotation at the center of the fiber dispersion. The angle 0 can be varied.

For a particular value of 0, the aligned fiber sample can be rotated through 360° (i.e. -can

be varied from 0° to 360°). The detector output of scattered light intensity is then

available as a function of the original magnetic field direction.

The actual instrumentation now in use is shown in figure 6. The laser beam of 514.5 nm wavelength from an argon ion laser is expanded to a diameter of approximately 1.5 cm. The

diameter of the beam illuminating the sample can be varied using an iris. This arrangement

allows a large area of the filter sample to be illuminated with light of constant intensity

per unit area. The cleared filter membrane is mounted in a holder which is inserted at the

center of the turntable. This is shown in figure 7. The scattered light is detected by a photomul tipl i er assembly mounted on an arm which can be rotated about the center of the

membrane filter sample. The photomultipl ier signal is displayed by an oscilloscope for

initial evaluation of the samples, and by an x-y recorder which is used to obtain permanent

records of the scattered light distributions. The sample turntable can be rotated at different speeds to accommodate this dual display technique.

Figures 8 a, b, and c show the outputs obtained respectively from crocidolite, amosite,

and chrysotile dispersions. The quantity of asbestos fiber per square millimeter of

filter area is indicated on the figures. These plots should be compared with the scattered

light distributions shown in figure 4. Chrysotile ha c very broad output peaks, making it

easily distinguishable from the scattered light outputs of the amphiboles. Furthermore the

differences between amosite and crocidolite shows that differentiation between certain

amphibole fiber types is possible.

113
Figure 4. Scattered light distributions

from:

a) unaligned UICC crocidolite;

b) aligned UICC crocidolite;

c) aligned UICC amosite;

d) aligned UICC Canadian

chrysotile;

e) aligned New Zealand

cummingtonite.

(The magnetic field direction

is indicated on the figures).

114
115

CROCIDOLITE ( UICC ) : I5ng /mm 2

°° 180° 360"

Figure 8(a).

AMOSITE (UICC TRANSVAAL) : 16 ng/mrn 2

0° 180° 360° Figure 8(b).

CHRYSOTILE (UICC CANADIAN) I2ng/mm 2

0° 180° 360

Figure 8(c)

5. Results

Initial work was performed with cleared filter samples which were permanently mounted

on glass microscope slides. In an effort to maximize the sensitivity of the method, possible noise sources were investigated. It was found that improperly cleaned glass slides

resulted in substantial scattering, and even a cleaned blank slide had small symmetrical

scattering peaks 180° apart which could be misinterpreted as being due to small concentrations of aligned fibers. Furthermore, samples mounted on slides showed spurious sharp

117
scattering peaks which were due to scratches in either the collodion film or the glass

slide. Samples were subsequently removed from the glass slide and followed by analysis of

the cleared filter alone. The results no longer incorporated these spurious peaks, and

the slide mounted technique was therefore abandoned in favor of examination of the cleared

filter without the slide substrate.

Examination of the cleared filters by phase contrast optical microscopy has shown that

the filter surface texture still remained when using the clearing technique described by

Ortiz and Isom [7]. This structure would lead to some background scattered light which

would limit the sensitivity. Clearing of filters using the AIA method [8] resulted in

samples without this surface texture and is now the preferred technique.

Figure 9 shows the scattered light distributions from dilute dispersions for which

distinct scattering patterns were obtained using crocidolite, amosite, and chrysotile. The

chrysotile sample was sputter-coated with a gold film of approximately 100 nm thickness.

This technique is currently under investigation as a means of enhancing the scattered light

intensity from the filter samples. It is interesting to note that the volume of dispersion

filtered for the crocidolite and amosite dispersions is 1/2 and 1/5 respectively of that

which would have been used in preparing TEM samples to have approximately 10 fibers per

grid square. Estimation of the detection limit as 1/10 of the peak height of these graphs

would correspond to a detection limit of 0.02 ng of amosite per square millimeter of filter

area illuminated by the laser beam, 0.04 ng/mm 2 for crocidolite, and 0.03 ng/mm2 for

chrysoti le.

CROCIDOLITE ( UICC) 0"4ng/mm 2 850 Fib /mm2

( L m = 0 6 pm )

118
CHRYSOTILE (UICC CANADIAN) ! O- 3 ng / mm 2

Figure 9(c).

To investigate the effects of non-fibrous particulate, various volumes of city water

containing 0.3 ug/mL of total insoluble solids were mixed with 5 mL of aliquots of a 0.2

ng/mL dispersion of amosite, and the resultant dispersions were magnetically filtered. The

effect of the non-fibrous particulate was to increase the value of the background intensity

between the peaks on the plots. The sizes of the peaks remained approximately constant.

Figure 10 shows the output for which the filter loading was approximately 50 ng/mm2 of total

insoluble solids and 1 ng/mm2 of amosite. The experiments so far conducted indicate that

approximately 0.1 percent by weight of amosite as a proportion of total insoluble solids

was detectable.

The current sensitivity of the magnetic filtration and light scattering technique is

approximately 0.03 ng of asbestos fiber per square millimeter of filter area. Improvements

in sample preparation technique and light scattering instrumentation are currently under

investigation and it is likely that sensitivities will be further improved. The presence of

non-fibrous particulate in a sample does not interfere with the detection of aligned

asbestos fibers. Good reproducabi 1 i ty has been obtained between samples containing the same concentration of amosite and a range of different concentrations of non-fibrous particulate. The sensitivity of the technique as an index of fiber concentration has also been

demonstrated. In particular for three different filter loadings of amosite 0.2 ng/mm2

, 1.0 ng/mm2

, and 16 ng/mm2, the corresponding peak heights were 9 mV, 17.5 mv , and 220 mV

(figures 9b, 10, and 8b respectively). The experimental results show that magnetic alignment, combined with analysis of light scattering, shows great promise as a rapid screening

method for asbestos detection.

119
This work was supported by the U.S. Environmental Protection Agency, Analytical

Chemistry Branch, Contract Number 68-03-2717. The authors wish to express appreciation for

this financial support.

References

[1] Birks, L. S. ; Fatemi , M. ; Gilfrich, J. V.; Johnson, E. T. Quantitative Analysis of

Airborne Asbestos by X-ray Diffraction, Final Report on Feasibility Study, EPA Report

EPA-650/2-75-004, January 1975.

[2] Melton, C. W. ; Anderson, S. J.; Dye, C. F. ; Chase, W. E. ; Hef fel f i nger , R. E. Development of a Rapid Analytical Method for Determining Asbestos in Water, EPA Report

EPA-600/4-78-066, December 1978.

[3] Diehl, S. R. ; Smith, D. T. ; Sydor, M. Optical Detection of Fiber Particles in Water,

EPA-600/2-79-127, August 1979.

[4] Timbrell, V. Alignment of Respirable Asbestos Fibers by Magnetic Fields, Ann. Occ.

Hyg. , Vol. 18 (1975), 299-311.

[5] Timbrell, V. Magnetic Properties of Amphibole Fibers (Abstract), Mineral. Soc. Bull.,

Vol 43 (1979), 3.

[6] Personal communication with Cressey, B. A. and Whittaker, J. W. , Department of Geology

and Mineralogy, University of Oxford, Oxford, England.

[7] Ortiz, L. A.; Isom, B. L. Transfer Technigue for Electron Microscopy of Membrane

Filter Samples, Am. Ind. , Hyg. J., Vol 35,(1977), 423-425.

[8] Reference Method for the Determination of Airborne Asbestos Fiber Concentrations at Workplaces by Light Microscopy (Membrane Filter Method), Asbestos International

Association, 68 Gloucester Place, London, England, September 1979.

120
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gai thersburg , Maryland, October 1-3, 1980. (Issued March

1982)

INTERLABORATORY MEASUREMENTS OF THE CHRYSOTILE ASBESTOS FIBER AND MASS CONCENTRATIONS

IN WATER SAMPLES

K. S. Chopra

Union Carbide Corporation - Metals Division

P.O. Box 579

Niagara Falls, New York 14302

D. Beaman

Dow Chemical

Midland, Michigan 48640

and

P. Cook

National Water Quality Laboratory

620 Congdon Boulevard

Duluth, Minnesota 55804

Abstract

In i nterl aboratory determinations of the chrysotile asbestos con- centrations in water, it was found that:

1. The accuracy in the determination of mass in ug/L is about

50 percent, and is systematically low, i.e., the average recovery

is 50 percent.

2. The precision in the determination of mass and fiber concentration

is about ±50 percent.

3. The precision of the analyses did not improve during the three years

of analyses involving six different chrysoti le-containing samples.

4. Water samples retained in glass tend to produce lower mass determinations as the length of storage is increased.

5. Selected area electron diffraction provides highly variable

results. In samples where the only fibers were chrysotile, the

portion of fibers reported as to giving positive SAED patterns

varied from 0-97 percent. The average fraction of chrysotile

fibers with positive SAED for all investigators and investigations

was 47 percent.

1. Introduction

Our initial work [l] 1 in measuring asbestos concentrations in water showed that interlaboratory reproducibility was ±50 percent, using the analytical transmission electron

microscope (ATEM). The earlier work involved the measurement of amphibole asbestos,

whereas this study deals with the chrysotile form. Moreover, this report presents, for

the first time, data on the accuracy of the ATEM method of measurement.

Figures in brackets indicate the literature references at the end of this paper.

121
A serious limitation of such i nterl aboratory comparisons involving complex equipment

and technology is that some of the participating laboratories do not provide high quality

data, primarily as a result of inexperience. This tends to bias the results negatively

and to make the method appear less effective than it really is. Problems still exist in

that uniform criteria for fiber identification have not been established. The laboratories, to varying degrees, base fiber identification on fiber morphology and/or fiber

structure (using electron diffraction) and/or fiber chemistry (using energy dispersive

spectrometry). In spite of these problems, it is clear that ATEM enables the asbestos

content of water samples to be determined with accuracy and precision adquate for most

environmental and toxicological studies being presently conducted.

2. Experimental Results and Discussion

In all published work, the precision of i nterl aboratory analyses has been reported.

The ASTM Committee has attempted to determine the accuracy of the method in i nterl aboratory

measurements, using clean water samples spiked with known quantities of chrysotile [2].

The results of this work are presented in Tables 1-4. Table 5 gives the results for an environmental sample, namely, water from a lake near a closed asbestos mine in Canada.

The precision and accuracy achieved by the group are summarized in Tables 6 and 7 for the

last six chrysotile samples studied by the group. The ASTM committee has analyzed seven chrysotile containing samples. The first group analysis is omitted from the present evaluation because of poor results [1] which were correctly attributed to a lack of experience

with chrysotile. At the time of the original chrysotile analysis, most group members had

worked only with amphibole asbestos.

Table 1. Results of Interl aboratory Measurement of Water Sample

Spiked with 6.3 ug/Liter of Chrysotile.

Fiber (mfl.)

a Mean Fiber Mass in Percent'3 Concentration Length urn ug/£ SAED

700 1.76 45.

C 35

918 1.69 10.9 70

562 1.20 2.6 5

510 1.29 3.7 30

509 1.71 4.4 10

1310 0.87 3.0

1100 1.26 9.5 30

1040 1.38 4.9 15

1290 1.31 3.0 55

830 1.70 4.6

Mean 877 1.42 5.2 31

± la ±305 ±0.3 ±3.0 ±22

mfl - means millions of Fibers/Litre.

^Percentage of chrysotile fibers giving positive selected area electron

di ff raction.

Q Excluded from mean.

122
Table 2. Results of Interl aboratory Measurement of Water Sample

Spiked with 0.63 pg/Liter of Chrysotile.

Fiber (mfl.)

a Mean Fiber Mass in Percent'

3 Concentration Length pm pg/£ SAED

30 1.35 0.21 75

29 1.08 0.15 97

22 1.09 0.12 5

23 1.21 0.13 26

29 1.28 0.18 29

27 1.41 0.63 30

22 0.96 0.07 15

51 1.38 0.13 53

22 1.21 0.08

Mean 28 1.22 0.19 41

± la ±9 ±.15 ±.17 ±31

amfl - means millions of Fibers/Litre,

b Percentage of chrysotile fibers giving positive selected area electron

di f fraction.

123
Table 3. Results of Interlaboratory Measurement of Water Sample

Spiked with 0.127 ug/Liter of Chrysotile.

Fiber (mfl.)

a Mean Fiber Mass in Percent^

Concentration Length urn ug/£ SAED

5. 3 0. 94 0. 084 40

4. 3 0. 77 0. 032 88

4. 8 1. 60 0. 120 90

9. 1 1. 30 10

14. 0 1. 10 0. 046

8. 6 1

30 0. 026

1. 4 106 0. 106 13

1. 6 0, 54 0. 004C

4. 7 2. 10 0. 110 36

5. 0 167 0. 066 20

5. 3 1. 47 0. 032 26

5. 4 1

24 0. 041 65

3. 1 1. 72 0. 036 69

0 2 0, 002

C 50

5. 0 1. 25 0. 029 60

6. 0 0. 085 75

5. 9 1. 3 0. 061 49

±3. 0 ±0. 4 ±0. 034 ±28

mfl - means millions of Fibers/Litre.

Percentage of chrysotile fibers giving positive selected area electron

diffraction.

Excluded from mean (delayed filtration).

124
Table 4. Results of Inter laboratory Measurement of Water Sample

Spiked with 0.56 ug/Liter of Chrysotile.

Mean

± la

• (rnfl.)

3 mtration

Mean Fiber

Length urn Mass inug/£

Percent

SAED

40 1. 00 0. 40 40

13 1. 13 0. 17 93

12 1. 9 0. 36 74

15 2. 1 0

9 1. 4 0. 15 38

3 2. 3 0. 03

C

19 1. 71 0. 31 84

29 1. 94 0. 47 34

24 1. 97 0. 53 41

14 U

U. 1 01 5 50

10 1. 25 0. 088 80

6 1 . 34 0. 0/b 65

<1 0. 099 C

91

17 2. 45 0. 13 50

23 0. 21 88

18 1. 6 0. 25 59

±9 ±0. 5 ±0. 16 ±27

a mfl - means millions of Fibers/Litre.

^Percentage of chrysotile fibers giving positive selected area electron

di f f racti on. Excluded from mean (delayed filtration).

125
Table 5. Results of Interl aboratory Measurements for an Environmental

Sample from a Lake in Canada Near a Closed Asbestos Mine.

Mean

± la

Fiber8 ConcentrationMean Fiber

Lenqth urn Mass in

ug/je

Percei

SAED

66 0. 83 0. 20 35

50 1.46 2. 06 96

7 1.40 0.05

C 72

52 1.60 2

32

12 1.62 0. 17

C

69 1.05 0. 70 26

75 1.54 1 . 70 39

111 0.75 0.54 28

41 1.19 0.43

9 1.78 0. 16 18

<1 1.75 0.04

C 70

17 1.75 0.09 15

56 0.47 64

53 1.3 0.7 42

±29 ±0.4 ±0.7 ±29

b

a mfl - means millions of Fibers/Litre.

Percentage of chrysotile fibers giving positive selected area electron

di f fraction.

Excluded from mean (delayed filtration).

126
Table 6. Interl aboratory Accuracy for Clean Water Samples Spiked with a Known

Quantity of Chrysotile Asbestos.

Sample:

Added

Chrysoti 1

e

in ug/£

6.3

0.63

0. 127

0.56

Mean ± la

Measured Mass

in ug/£

Group Mean

± la

5.2 ± 3.0

0. 19 ± 0. 17

0.06j± 0.03 4

0.25 ± 0.16

Accuracy

Measured-Added Mass

Added Mass Recovery

in % in %

- 17 83

- 70 30

- 52 48

- 55 45

48 +22 52 ± 22

Measured Mass

in ug/£-Calculated

using d = 41 5^

group mean ± la

5.1 ± 1.6

0.15 ± 0.05

Table 7. Compilation of Precision and Accuracy Data for Six Samples Containing

Chrysotile Asbestos.

Mass Determination Concentration Determination

Sample:

Added Measured Mass

Chrysotile pg/£

in ug/£ Mean ±la

6.3

0.63

0. 127

0.56

Canadian

Lake Sample

12/6/76

Mean ±la

5.2 ± 3.0

0.19 ± 0.17

0.06

] ± 0.034

0.25 ± 0.16

0.7 ± 0.7

Percent of

Concentration Fibers Giving

Precision in MFL: 106 Precision Positive SAED:

Accuracy ±a/mean Fibers/liter to/mean Mean ±la

in % Relative a Mean ±la Relative a ( range)

17

70

52

55

58, 31'

89, 33

56

64

unknown

not spiked

100

not reported not

measured

877 ± 305

28 ± 9

6 ± 3

18 ± 9

53 + 29

59 ± 24

48 ±22 46 ± 17

35

32

50

50

55

41

44 ± 9

31 ± 22

(5 - 70)

41 ± 31 (5 - 97)

49 ± 28

(10 - 90)

59 ± 27

(0 - 93)

42 ± 29

(2 - 96)

not reported

Mass based on f ibri 1 concentration and fibril diameter of 415 A. For spiked samples only using footnote a values.

127
The results in Table 6 show that the measured mass was always less than the added mass, with recoveries ranging from 30 to 83 percent and averaging about 50 percent. The accuracy,

defined as 100 (measured mass - true mass)/true mass, ranged from 17 to 70 percent and

averaged about 50 percent. The actual accuracy is probably somewhat better than this

because of the loss of fibers associated with delays in filtration. Figure 1 is a plot

of the mass determined for the 0.127 pg/L sample as a function of the days of storage prior

to filtration. It is apparent, notwithstanding the scatter, that the measured mass tends

to decrease after long storage times. The 0.63 ug/L sample was prepared from an aliquot of

the 6.3 ug/L sample and was filtered after storage. This may account for the poor accuracy

(70%) in the case of the 0.63 ug/L sample.

A significant problem in reporting mass is the need to measure the fiber diameter.

This task cannot be accomplished by visual determination on the fluorescent screen of the

microscope. A careful determination of the fibril diameter characteristic of the addition

o was made by one laboratory and found to be 415 A. Using this value and reported fibril

(not fiber) concentrations to calculate the measured mass does not significantly alter the mean value of the reported mass. However, it does markedly reduce the scatter, i.e., it

improves the precision (see Table 6, column 5).

Table 7 summarizes the precision and accuracy values for the chrysotile samples. From

these limited data, it appears that the precision in the determination of fiber concentration and chrysotile mass is about ±50 percent. The accuracy is also about 50 percent and

is systematically low. There does not appear to be a significant correlation between

accuracy or precision and sample mass or fiber concentration within the range examined.

Thus, the precision and accuracy of the ATEM technique remains at about 50 percent for the

determination of chrysotile asbestos in water. There was no improvement during the 3 years

of analysis (August 1976 to December 1979), as shown in figure 2. These results were achieved on relatively clean water samples, i.e., spiked samples and lake samples. The

interlaboratory reproducibility would not be as good on relatively unclean water samples,

such as plant effluents or most rivers. On the other hand, if the only data included in

the evaluation were from laboratories making measurements routinely under carefully con- trolled conditions, the interlaboratory reproducibility would be improved.

There are many reasons for what appears to be mediocre precision and accuracy and the

reliance on electron diffraction is a major contributor to the problem. In these samples,

where the fibers were known to be chrysotile, only 47 ± 27 percent of the fibers gave

identifiable selected area electron diffraction (SAED) patterns, and the percentage of

fibers so identified varied greatly between the different laboratories (0-97%). This is primarily due to differences in equipment and to the criteria used to classify a SAED

pattern as positive. Improvement in the diffraction portion of the analysis has been

reported [3].

Refinements and improvements in the methodology are being pursued and these will

undoubtedly lead to better interlaboratory comparisons. In the meantime, accuracy and

precision of 50 percent would appear to be sufficient to address any meaningful water com- tamination problem involving asbestos.

The Committee has studied some problems associated with the analysis on a limited basis

and found that leaching of MgO from chrysotile is rapid in HC1 , but does not occur after

7 days in 30 percent NaOH [4]. The fiber contamination of new Nuclepore and Millipore

filters and the fiber loss associated with storage in glass are still under investigation.

128
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130
Following is a list of the members of the Task Group of Subcommittee E04.ll on Electron

Metallography under Committee E-4 on Metallography.

C. Anderson, Environmental Protection Agency

D. Beaman, Dow Chemical-Midland

D. Benefiel, Dow Chemical -Freeport

K. Bishop, Minnesota Department of Health

E. Chatfield, Ontario Research Foundation

K. S. Chopra (Chairman), Union Carbide Corp.

P. Cook, EPA (ERLO)-Duluth

R. Feldman, EPA, Cincinnati

P. Giles, Bethlehem Steel

H. Bohmer, PPG Industries

R. Lee, U. S. Steel

J. MacArthur Long, USEPA, Athens, GA

C. Melton, Battel le

J. Millette, Environmental Protection Agency

J. Mothersill, Lakehead University

E. Peters, A. D. Little Co.

S. Ring, Minnesota Department of Health

I. Stewart, McCrone Associates

G. Surrus, Dow Chemical-Sarnia

A. Szirmae, U. S. Steel

G. Yamate, Illinois Institute of Technology Research Institute

References

[1] Chopra, K. S. J. of Testing and Evaluation 6(4): 241-247; 1978.

[2] The spiked samples were graciously prepared and distributed by E. J. Chatfield and

J. Dillon of the Ontario Research Foundation.

[3] Beaman, D. R. ; Baker, H. M. Anal. Chem. 52: 1983-1984; 1980.

[4] C. Melton of Battelle performed the analysis of the leached fibers for the ASTM

committee.

131
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gaithersburg, Maryland, October 1-3, 1980. (Issued March

1982)

CONSIDERATIONS IN THE ANALYSIS AND DEFINITION OF ASBESTOS USING ELECTRON MICROSCOPY

R. J. Lee, J. F. Kelly, and J. S. Walker

U. S. Steel Corporation

Research Laboratory

Monroeville, Pennsylvania 15146

Abstract

The most widely used working definition for asbestos in electron

microscopy analysis was established on neither a physical nor health

related basis. It derived from the definition applied to occupational

monitoring using optical microscopy, without regard for the differences

between the two types of instruments. The definition has no lower limit

on fiber length and an inappropriate lower limit on aspect ratio. This,

plus the reporting of data with poor counting statistics, has detracted

from the effective use of electron microscopy in quantifying nonoccupational exposures to asbestos. Thus, much of the time and effort expended

so far on this type of analysis has no long term value. The purpose of

this paper is not to present new data, but rather to summarize some difficulties with current analyses. Further, we propose for

consideration some operational definitions and procedures that are more physically meaningful and cost effective than those presently in use.

Introduction

Until recently, optical microscopy has been the only method used for monitoring

concentrations of asbestos in the work place. Because of the limited resolution of the

optical microscope, the transmission electron microscope (TEM) is now widely used to measure concentrations of sub-microscopic asbestos particles in nonoccupational samples

[l] 1

. Even though the TEM is a powerful instrument, rigorous measurement of fiber concen- tration is a difficult problem because of a large number of variables involved in the

procedure [2].

This symposium is being held to review and identify those aspects of the analysis that

can be quantified and to propose the development of a set of standards. Therefore, this is an ideal forum to address those questions which left unresolved will continue to affect

results in spite of attempts to standardize.

This paper is divided into two sections. In the first part, some statistical con- siderations are reviewed; in the second part, implications of the working definition of

asbestos used in electron microscopy are discussed.

1. Analytical Considerations

Asbestos analysis involves several separate phases, from sample collection to data

interpretation. The goal is to determine the asbestos concentration from a microscopic

count, and so the ultimate result is critically dependent on the validity of the assumptions

and procedures used in each phase. The basic assumptions are: (1) representative samples

1 Figures in brackets refer to the literature references at the end of this paper.

132
collected; (2) particulates are uniformly deposited on the filter substrate; (3) no con- tamination introduced during sample preparation and analysis; (4) no instrument or operator

influence on the analysis and no misidentif ication of nonasbestos fibers; (5) collection

and analysis of all asbestos fibers in the sample; (6) no effect from sample dilution or repreparation steps.

The first four assumptions have been extensively examined and are now being rigorously

studied in the NBS/EPA program (Steel et al . , [3]). Since the collection efficiency of

any filter is dependent on the size of the particles, the fifth assumption cannot be

valid. Further, as will be discussed in the following the sixth assumption can lead to erroneous results if not carefully and systematically validated.

The sixth assumption is required because of constraints on the total mass per unit area

that can be analyzed. Normally, the electron microscope analysis of asbestos in air or water samples requires dilution of the original sample whenever the expected loading will

exceed 5 to 20 ug/cm2 [4]. A small portion of the results of a recent large-scale study of

mine water effluents are presented in Table 1. Noted here are calculated fiber concentrations ranging from 106 to 10 11 f/l, based on the detection of only one or two fibers per

sample. Examination of the data reveals several inadequacies that have important implications. By dividing the reported concentration by the number of fibers counted, the unit

fiber concentration can be back-calculated. The report also indicated that 20 grid openings were analyzed on each sample. Further, it was stated that 100 mL of water were filtered

each time. With this information the effective sample dilution and blank levels can be

calculated.

Table 1. Electron Micoscope-Determi ned Concentration of

Chrysotile Asbestos in Mine Water Samples.

Chrysotile*

Concentration Mass Fiber

Ore (f/l) (gm/l) Count

Ag 107 10- 7 4

Ag 1011 10" 4 2

Au 109 10" 6

1

Cu 10 11 10" 3 30

Fe 1010 10" 4 7

Hg 1011 10" 4 11

Mo 1010 10" 5

1

Pb/Zn 1010 10" 5 4

Ti 109 10- 3 4

u 106 10- 8

1

W 1011 10" 3 19

Data extracted from consultant's report to EPA.

Figure 1 shows the calculated concentration in fibers per litre and fibers per gram

compared with the calculated dilution factor. It can be seen that the concentration is

directly proportional to the dilution and independent of sample type over some six orders

of magnitude. Thus, the effective blank level is not significantly different than the

calculated concentration for the vast majority of samples.

133
Dilution Factor

g/liter f/liter

Figure 1. Comparison of calculated dilution factor and

reported chrysotile concentrations.

This example analysis illustrates several problems common to all asbestos analysis.

There is no uniform method for reporting results, and inadequate regard is given to the

effects of sample dilution and the meaning of terms such as "minimum detectable limit" and

"effective blank levels". These inadequacies have resulted from the unavailability of a coherent statistical treatment of the data.

To eliminate these deficiencies, we propose several procedures for consideration.

Every report of asbestos analysis should include the following:

1. A statement of the population statistics assumed; binomial, negative binomial,

Poisson, or other [5].

2. An estimated uncertainty range for the calculated concentration [2]. The uncer- tainty range associated with the unit fiber concentration and the effective blank level

should also be stated.

3. Only those cases where the lower confidence level for the measured concentration

exceeds the upper confidence level for the unit fiber concentration or effective blank

level (whichever is larger) should be reported as a positive result. For example, assuming

the Poisson distribution, the lower confidence level for unit fiber concentration is 0.1

(with a = 0.05) and the upper confidence level is 5.6. In order to report a positive

result, the analysis would continue until 12 or more fibers were detected. If the analysis

is terminated before this point, the results should be reported as indeterminate.

4. In the preparation of samples, the total suspended solids (TSS) for water or total

suspended particulates (TSP) for air should be determined and used to estimate the concen- tration pg/cm2 of particulates on the final filter substrate. Those samples that must be

diluted in order to achieve the optimal 5 pg/cm2 loading should be prepared twice, once at

an estimated loading of 10 pg/cm2 and another at about 2 pg/cm2

. Two 200 mesh TEM grids

should be prepared for each sample and five openings on each grid analyzed. If the measured

concentrations are not statistically equivalent, the heavily loaded sample should be

repeated or the result reported as indeterminate. Another advantage of measuring TSP or TSS before the analysis is that, using simple assumptions, a lower limit on the measurable

concentration can be estimated as a function of three parameters. The result is a so-called

"minimum measurable concentration" for water analysis, as shown in Table 2. The data

134
shown are based on the following assumptions: (1) 5 ug/cm2 loading on the final filter;

(2) analysis of 100 fibers in 20 to 200 mesh grid openings (1.6 x 10-3 cm2 ); (3) mean particle mass of 10~ 12

g.

Table 2. Suspended Solid Concentration and Required Sample Aliquot vs. Measurable Asbestos Concentration.

Measurable

Concentration

(fibers/1 iter)

10

2

10

3

10

4

10'

10

l

10'

10'

10"

10

10

Maximum Concentration

of Suspended Solids

(mg/L)

10

10"

10

10"

1

10

10

2

10*

-4

-2

Sample Aliquot

500 liters (^130 gal.)

50 liters

5.0 liters

500 mL )

50

5

mL J

mL

.5 mL

.05 mL

clean drinking water

Iver water

While these assumptions are not the only considerations in obtaining a good analysis,

they do represent a major obstacle when analyzing environmental specimens where the

expected absolute fiber count will be small. Even though these recommended procedures may

require further development, they do at least provide a starting point for discussion of an otherwise neglected topic.

2. Working Definition of Asbestos

Presently, the most common working definition of asbestos for electron microscope

analysis calls for only a 3:1 length-to-width ratio, no minimum length, and the identification of a particle as an amphibole or chrysotile mineral. The shortcomings and ambiguities

of such a working definition have been discussed in detail elsewhere [5-8]. It is our intent here to illustrate the effect of this definition on the accuracy of asbestos

analysis rather than to argue its i nappropriateness alone.

The work of the ASTM Task Force on Asbestos was discussed by Chopra [9], and that of

the NBS discussed by Steel et al., [3]. Methods for identifying asbestos were presented

by Ring [7] and Yamate [10] at this meeting. From these discussions, it is apparent that

techniques are now well developed for mineral identification. Under very carefully controlled conditions, an analysis costing from $1000 to $2000 yields a result with a variability

from 50 to 100 percent. Even with such very expensive analyses, significant problems

remain because of two factors - the aspect ratio and length criteria used in the analysis.

Using the 3:1 aspect ratio fiber definition is responsible for a major fraction of the

large analytical cost, mainly because detailed analyses are necessary in order to eliminate

the interference by nonasbestos particles. Alternatively, if low-cost analyses are used,

large uncertainties are introduced because of misidentification.

The absence of a minimum length in the definition also has several effects:

1. The analyst spends most of this time on the smallest size particles; this results

in significant uncertainties in the estimated concentrations of the longer fibers.

135
2. As shown by Steel et al., [3], under the most carefully controlled conditions, two

analysts may arrive at the same average count by counting different objects.

3. Steel et al., [3] also report that a significant fraction of fibers shorter than

2 urn are missed in the analysis. Thus, even though we strive to produce an absolute fiber

count, the count is only a fraction of the total concentration and that fraction is clearly

dependent on the instrument and operator employed.

4. The reported concentration is not a measure of a known health hazard. Instead, it

is a mixed index which includes the concentration of both known and potential hazards. This

ultimately limits our ability to identify the individual significance of either group.

The problems listed here are not strictly analytical but rather derive from the

definition of asbestos. What is reguired for a meaningful analysis of fiber concentration

is an operational definition which includes bounds on the aspect ratio, length, and width.

Clearly, a given analysis will measure only a portion of a distribution. If desired, it is

possible to calculate the total concentration from the measurement of a subset of a distribution if the form of the distribution is known. Problems in asbestos analysis arise

because subset concentrations are reported implicitly as total concentrations and are compared with results from other laboratories which measured different subsets.

As the basis for the formulation of a meaningful working definition of asbestos, we propose the following:

1. For a routine method, a minimum aspect ratio of 10:1 should be used in a screening

analysis or survey. Existing data indicate that this would not affect the chrysotile

analysis at all and amphibole analysis only when the sample contains a significant percentage of acicular nonasbestos particles [11-16]. While this would undoubtedly result in

missing 5 to 20 percent of the short asbestos particles, it would eliminate 70 to 80 percent

of the nonasbestos particles from consideration.

2. A lower length limit for routine electron microscope analysis should be adopted.

On the basis of available information, a reasonable limit would be somewhere between 0.75

and 2.0 microns [3].

3. Asbestos analyses should be grouped into at least three size fractions and acceptable uncertainty levels defined for each range. For example, the length categories less

than 2, 2 to 5, and greater than 5 pm might be chosen, and a 50 percent relative error defined as the minimum level of acceptance for each size range.

Summary

Analysts need to examine the objectives in performing analyses for asbestos. Is it to provide an absolute measure of the concentration, or is it to produce a reliable,

reproducible measure or index which allows comparison of exposures?

In the past we have attempted to obtain answers to two questions simultaneously.

First, a research-type question, "What is the significance of small, low-aspect ratio

particles?" Second, "What are the concentrations of known hazards in environmental samples,

and by what procedures can they be measured?" Electron microscope analysts must treat

these as separate questions.

Analysts have been waiting for the medical community to define the size and shapes of

significance. Unfortunately, answers to the above questions may not be forthcoming if

analysts cannot provide a meaningful data base.

Now, at this session on standardization, it is time to define the statistical requirements and analytically based working definition of asbestos that will permit reliable and

effective analyses by electron microscopy.

136
References

[1] Zumwalde, R. D. ; Dement, J. M. Review and Evaluation of Analytical Methods for

Environmental Studies of Particulate Exposure. Proceedings 1st FDA Office of Science

Summer Symposium on Electron Microscopy of Microfibers; 1976. 139-150 (Available

from Supt. of Documents, U.S. Government Printing Office, Washington, D.C. 20402).

[2] Chatfield, E. J.; Dillon, M. J. Some Aspects of Specimen Preparation and Limitations

of Precision in Particulate Analysis by SEM and TEM. Proceedings SEM/78, Vol. 1; 1978. 487.

[3] Steel, E. B. ; Small, J. A.; Sheridan, P. Analytical Errors in Asbestos Analysis by

Analytical Electron Microscopy. These proceedings, p. 162.

[4] Anderson, C. H. ; Long, J. M. Preliminary Interim Procedure for Fibrous Asbestos.

U.S. EPA; 1976. p. 30.

[5] Chatfield, E. J.; Lewis, G. M. Development and Application of an Analytical Technique

for Measurement of Asbestos Fibers in Vermiculite. Proceedings SEM/80, Vol. 1; 1980.

329.

[6] Lee, R. J. Basic Concepts of Electron Diffraction and Asbestos Identification Using

SAD. Proceedings SEM/78, Vol. 1; 1978. 677.

[7] Ring, S. ; Suchanek, R. Fiber Identification and Blank Contamination Problems in the

EPA Provisional Method for Asbestos Analysis, these proceedings.

[8] Zoltai, T. Asbestiform and Acicular Mineral Fragments. Health Hazards of Asbestos

Exposure. Annals of the New York Academy of Sciences. I. J. Selikoff; E. C. Hammond,

eds. ; 1979. p. 621.

[9] Chopra, K. S. Interlaboratory Measurements of Amphibole and Chrysotile Fibers in

Water, these proceedings.

[10] Yamate, G. ; Beard, M. E. Refinements in the EPA Provisional Methodology, these

proceedi ngs.

[11] Pooley, F. D. ; Clark, N. Fiber Dimensions and Aspect Ratio of Crocidolite, Chrysotile and Amosite Particles Detected in Lung Tissue Specimens. Health Hazards of

Asbestos Exposure. Annals of the New York Academy of Sciences, Vol. 330, I. J. Selikoff; E. C. Hammond, eds.; 1979. p. 711.

[12] Wylie, A. G. Fiber Length and Aspect Ratio of Some Selected Asbestos Samples. Health

Hazards of Asbestos Exposure. Annals of the New York Academy of Sciences, Vol. 330,

I. J. Selikoff; E. C. Hammond, eds.; 1979. p. 605.

[13] Winer, A. A.; Cossette, M. The Effect of Aspect Ratio on Fiber Counts: A Preliminary Study. Health Hazards of Asbestos Exposure. Annals of the New York Academy

of Sciences, Vol. 330, I. J. Selikoff; E. C. Hammond, eds.; 1979. p. 661.

[14] Stevenson, R. J. Mineral Fiber Investigations of the Regional Copper-Nickel Study on Northeastern Minnesota. Health Hazards of Asbestos Exposure. Annals of the New

York Academy of Sciences, Vol. 330, I. J. Selikoff; E. C. Hammond; 1979. p. 705.

[15] Bignon, J.; Sebastien, P.; Gaudichet, A. Measurement of Asbestos Retention in the

Human Respiratory System Related to Health Effects. Nat. Bur. Stand. (U.S.) Spec.

Publ. 506; 1978 November, p. 95.

[16] Campbell, W. J.; Blake, R. L. ; Brown, L. L. ; Cather, E. E. ; Jobery, J. J. Selected

Silicate Minerals and Their Asbestiform Analogs. Bureau of Mines Information Circular 8751 ; 1977. p. 46.

137
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gaithersburg, Maryland, October 1-3, 1980. (Issued March

1982)

COMMENTS ON THE ACHIEVABILITY OF A VALID ASBESTOS STANDARD FOR TEM COUNTINGI. M. Stewart

Walter C. McCrone Associates, Inc. Chicago, Illinois 60616

1. Introduction

The validity of any standard reference material is only as good as the validity of the

sample preparation and analytical methodology used to characterize the standard. In the case of standards for the determination of fiber counts by electron microscopy, the literature is somewhat vague on the validity and the reproducibility of specimen counting

procedures. Most electron microscopi sts , when questioned on the subject, would generally

agree that results are valid within an order of magnitude but to a certain extent this has

been a gut-feeling or an inspired guess rather than the results of any tests performed to confirm this. Such round-robin tests as have been performed have generally been confounded

statistically by a confusing medley of comparisons between sample preparation methodologies,

different instrumentation used for the analyses, and variability in operator experience both

on these instruments and of asbestos identification. It is not the object of this paper to further cloud the issue but, instead, to provide some gleam of hope that standards may,

i ndeed, be val id.

1.1 Background

The work described in this report was performed in late 1974 and early 1975 with the

object of establishing whether or not a particular sample methodology would give accurate

and reproducible results for the determination of asbestos in water samples.

Walter C. McCrone Associates, Inc. had been contracted by the EPA to perform a sampling

of the impact of point and non-point sources of asbestos on waterborne levels of asbestos.

This program was to be conducted nationwide, sampling and filtration were to be carried out

on-site and the filters were to be returned to our Chicago laboratory for subsequent

analysis. The filter medium chosen for these samples was the Mi 1 1 ipore® 0. 45 urn pore size,

47 mm diameter mixed ester filter and samples were collected by vacuum filtration using the

standard Mi 11 ipore glass funnel and flask arrangement. Sample preparation was to be by the

condensation washing method in a Soxhlet condenser. Without discussing in detail the pros

and cons of Millipore vs. Nuclepore®, condensation washing vs. Jaffe Wick, the rationale for

the selection of this procedure can be briefly summarized as follows:

1. Sampling was to be performed in a mobile laboratory and samples were to be retained in

the laboratory until its return to home base. In many instances this meant three to four weeks of travel on the road in the laboratory. Although there was no hard data at that time 1 to support this assumption, it was felt by all the investigators concerned

that Millipore filters would have a better fiber retention than the smoother Nuclepore

filters under such circumstances, bearing in mind that facilities to carbon coat the

Nuclepore filters in the field were non-existent.

2. Previous work carried out independently by McCrone Associates and by Jack Murchio of

the University of California, Berkeley, had indicated that, in the case of glass fibers

at least, there was a tendency for large fibers to be lost during Jaffe Wick preparation of Nuclepore filters. As most of the samples on the contract were to be taken

close to point sources, it was felt that there was a real danger of the loss of large

fibers if Nuclepore filters and the Jaffe Wick techniques were used.

*A limited sample performed at a later date tended to confirm this finding (Stewart, I. M. , Proc. F.D.A. Office of Science Symp. on Electron Microscopy of Microfibers, 1976, p. 96

and Table II.

138
3. Comparative studies performed by the EPA during the Reserve Mining case in which

samples had been split between McCrone Associates and Jack Murchio, the latter using a Jaffe Wick technique, had shown good comparability of fiber counts between the Millipore condensation washing technique used by McCrone Associates and the Jaffe Wick as used by Murchi o

2

.

Taking these criteria into consideration therefore, the decision was made to use Millipore filters and the condensation washing technique.

The series of tests described in this paper were, therefore, devised to establish some

of the parameters which might affect the reproducibility and accuracy of the analysis of

Millipore filters, condensation washed.

Three test series were conducted:

1. An extensive examination of samples taken from one individual filter designed to test

the reproducibility of a single operator, the variation between different operators

and the evenness of distribution on the filter.

2. A series of tests aimed at testing the reproducibility between different filters

produced from different aliquots of the same sample.

3. A series of dilution tests aimed at assessing the accuracy of the count. The tests

are described in the following section.

1.2 Materials and Methods for Conducting Tests

In all groups of tests considered, the sample preparation method was identical and was

a direct transfer method using a Soxhlet extractor with acetone as a solvent. All the

sample measurements were made on a JEOL JEM-200 transmission electron microscope operating

at 200 kV. A standard suspension was prepared of chrysotile asbestos from the Jeffrey

Mine in particle-free water. We had worked extensively with this suspension in connection

with a sub-contract from an FDA contract on animal studies and, therefore, felt that we knew the suspension quite well. The suspension was prepared in water that had been filtered

three times, the last filtration being through a 0.2 urn pore size membrane filter. Three

filters were prepared in succession from this suspension; the remaining unfiltered suspension was then diluted by a factor of 10 and a fourth filter prepared, then by a further

factor of 10 and a fifth filter prepared. These five filters formed the basis for the

three test series. Measurements made on these filters are listed in Table 1.

1 . 3 Test Series I

The first of the five filters was the subject of a detailed statistical analysis

(performed by Dr. Neill of the Illinois State Water Survey), the object of which was to determine the variation in particle distribution on the filter. Thirteen test locations were established on the filter as shown in figure 1. At each of these 13 test locations an electron microscope grid was prepared and examined. The sample was prepared on finder's

grids which enable the precise re-location of the area examined on the grid. In this way

it was possible for more than one observer to measure the asbestos content on identical

areas. Three grid squares on each grid were examined by the first observer to give a total

of 39 observations from the one filter. Each grid square on the finder's grid is equivalent

in area to approximately three grid squares on a regular 200-mesh TEM grid.

2 A preliminary report on asbestos in Duluth, Minnesota area, Office of Technical Analyses,

Office of Enforcement and General Counsel, U.S. EPA, January 1974, Appendix III.

139
Table 1. Fiber Counts on Standard Chrysotile Suspension.

Filter Grid Location Observer 1 Observer 2

1 DELTA 1 0. 31 X 10

8

1 X2 0. 37 X 10

8

1 M-0 0. 32 X 810B

2 El 0. 32 X 10

8

2 Y2 0. 36 X 10

8

2 DELTA 1 0. 33 X 8 10°

3 1-2 0. 38 X 108

3 F-l 0. 31 X 10

8

3 Rl 0. 32 X 10°

4 c-o 0 30 X 10

8

4 A2 0. 31 X 10

8

4 6-3 0. 28 X

8

10

B

5 X3 0 34 X 108

5 H2 0 34 X 10

8

5 R0 0 26 X 8 10°

6 G3 0 25 X 10

8

0. 25 X 108

6 2-1 0 37 X 10

8

0. 38 X 10

8

6 7-2 0 35 X 8 10° 0. 44 X 10

8

7 X-3 0 28 X 10

8

10

8

7 P-3 0 32 X

o 108

0. 30 X

7 Kl 0 31 X

8 10°

8 DELTA 2 0 35 X 10

8

10

8

8 1-2 0 39 X 10

8

0. 37 X

8 D-3 0 36 X

8 10°

9 M-l 0 39 X 10

8

9 7-0 0 36 X 10

8

9 4-1 0 37 X

ft 10

S

10 1-1 0 33 X 108

0 35 X 10

8

10 H-2 0 35 X 10

8

8

10 0-2 0 32 X 10° 8io

a

0 34 X

11 L-0 0 34 X 10

8

10

8

11 5-1 0 33 X 10

8

0 24 X

11 W-2 0 37 X 10

8

0 42 X 10

8

12 K-0 0 35 X 10

8

12 s-o 0 35 X 10

8

12 6-1 0 35 X 10

8

140
Table 1 continued.

Filter Grid Location Observer 1 Observer 2

IB 01

Q-0 U. 0 1 X 1

U

13 T-l 0.34 x 10

8

13 Y-l 0.36 x 10

8

DELTA 2 0.23 x 10°

01 7-1 p 0.27 x 10

01 G3

Q

0.22 x 10

02 E3 0.25 x 10°

02 SI

o 0.27 x 10

02 5-1

Q

0.27 x 10

03 Nl 0.30 x 10°

03 DELTA 0

Q

0.22 x 10

03 Q0

o 0.26 x 10

04 L-0 0.23 x 10°

04 1-1 0.27 x 10

8

04 R-l 0.24 x 10

8

n£uo /i _ n n oo v i U . £.0 X 1 U 05 F3 0.27 x 108

U3 p-n nU ?q v ~\rfi

CO X 1 u

A3

p 0.32 x 10

01 DELTA 1 0.29 x 10

8

01 6-0 0.26 x 10

8

02 Tl p 0.29 x 10

02 w-o 0.26 x 10

8

02 L-l 0.27 x 10

8

03 F-2 0.26 x 10°

03 Fl p 0.26 x 10°

03 4-0

p 0.27 x 10°

04 N-i 0.27 x 10

8

04 6-1 0.27 x 10

8

04 Q-0 0.24 x 10

8

05 J3 0.30 x 10

8

05 H2 0.28 x 10

8

05 DELTA 2 0.29 x 108

141
continued.

Filter Grid Location Observer 1 Observer 2

2 01 0.23 x 10

02 0 25 X 10

7

03 0 20 X 10

7

04 0 28 X 10

7

05 0 23 X 10

7

01 0 31 X 10

6

02

03 0 24 X 10

6

04 0 70 X 10

6

05 0 35 X 10

6

Figure 1. Sampling locations on filter surface.

142
Summarizing the data from Table 1 in Tables 2 and 3, it can be stated that good reproducibility was obtained from different locations on the same filter indicating a uniformity

of distribution which, in fact, exceeded our expectations, bearing in mind that the

approximate 10 percent standard deviation in fiber counts for Observer 1 represents both

the variation of the observer's interpretation of what constituted a fiber plus the variation of distribution on the grids and filter. Additionally, the statistical treatment of

the length and width distributions by Dr. Neil 1 showed no systematic variations in these

parameters which would have been indicative of size segregation effects on the filter.

Table 2. Statistical Data Derived from Table 1 for Filter 1

Filter

Observer 1

Observer 2

Grid

Mean Fibers/Liter

Observer 1 Three readings

Observer 2 (No. of readings in

per location parenthesi s)

1 0 333 X

ft 10°

2 0. 337 X

, ~8 10°

3 0. 337 X 10

8

4 0. 297 X 10

8

5 0. 313 x 10 8

357 x 10

8

6 0 323 X 10 (3)

8

0.

7 0. 303 X 10

8

0 30 x 10

8

(1)

8 0. 367 X 10

8

0 37 x 108

(1)

9 0. 373 X 10

8

345 x 108

10 0. 333 X 10 (2)

8

0.

11 0. 347 X 10

8

0. 33 x 108

(2)

12 0. 35 X 10

8

13 0. 34 X 108

Mean Value Std. Deviation No/Readings

0.335 x 10

8 9.88% 39

0.3433 x 10

8 19.08% 9

143
Table 3.

No. of grid

Run No. Squares Examined Fibers/Liter Rel

.

S.D.

la 39

a 33.6 x 106 9 75%

lb 15

b 25.7 x 106 8 96%

lc 15

b 27.4 x 10

6

6 85%

2 20

C 2.38 x 10

6

11 09%

3 30

d 0.36 x 106 48 62%

Three grid squares on each of 13 different grids prepared from

the same filter, freshly prepared ul trasonerated suspension.

Three grid squares on each of five different grids prepared

from the same suspension as la, allowed to settle then manually

shaken to redisperse. lb and lc represent two separate filters.

Four grid squares on each of five different grids prepared from

suspension of la, b, and c diluted to 1/1 0th concentration.

'six grid squares on each of five different grids prepared from

suspension of la, b, and c diluted to 1/1 00th concentration.

When a second observer measured areas which had previously been observed by the first

one we again get good reproducibility in terms of the average fiber count, although in this

instance the standard deviation has in fact risen to approximately 20 percent.

Fiber levels measured by Observer 1 on the second and third filters prepared from the same standard chrysotile suspension did not differ significantly from each other but did

show a significant decrease in fiber count compared to the first filtration. Though both

filters did show decreases in length, width, and mass, the decrease could not be regarded

as statistically significant, it was therefore impossible to determine whether the decrease

in fiber count was due to failure to re-suspend all the large settled particles, to entrapment of smaller particles on the container walls, or whether both mechanisms might have

been operative.

On examining filters representing dilutions from the original standard suspension,

Table 3, there was indeed an order of magnitude decrease in fiber count observed between

the successive filtrations. One should note that when the data for the 100-fold dilution

is considered the standard deviation has increased markedly, reflecting now the effect of

probability statistics in terms of whether or not a fiber would be observed in a particular

grid square (at the 100 dilution level the fiber count should have averaged one to two

fibers per finder's grid square).

2. Summary

To summarize the results presented in this paper then, it seems that there is good

hope that a true, accurate, standard can be produced and can be made reproducible for the

analysis of asbestos by TEM, provided we ensure a sufficiently high loading for statistically valid counts to be made from it. This work suggests that a minimum loading of the

order of 10 fibers per 1000 square micrometers of filter is desirable.

144
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS , Gaithersburg, Maryland, October 1-3, 1980. (Issued March

1982)

MEMBRANE FILTER METHOD: STATISTICAL CONSIDERATIONS

Gerald R. Chase

Johns-Manvi lie Corporation

P.O. Box 5108

Denver, Colorado 80439

Abstract

The historical evolution of the understanding of errors associated

with workplace sampling for airborne asbestos using the membrane filter

method is presented. Statistical considerations of the membrane filter

method are illustrated using analytical and empirical results. Possible

sources of error and those which should be considered are discussed. The

importance of the form of the distribution used to quantify the

reliability of the method is shown using theoretical and empirical

findings. In particular, it is shown that Gaussian assumptions can be

very misleading when the error distributions are skewed. Finally, the

characterization of the reliability of the method is discussed.

Most of the early work in measuring workplace asbestos levels was carried out using

techniques which yielded results in the form of total particulate concentrations. The mem- brane filter method, which yields results in the form of the number of fibers of specified

dimensions, was pioneered by a British textile company in the 1 960 ' s . However, the

widespread use of the membrane filter method did not occur until the late 1 960 1 s and early

1970's.

The evolution of workplace monitoring of asbestos has been the source of a number of

interesting theoretical and analytical statistical problems, some of which are still under

i nvestigation.

Some studies have investigated the relationship between different monitoring techniques. One of those investigations, reported by Ayer, Lynch, and Fanney (1965) [l] 1

, used

a study design involving side-by-side samples of the membrane filter and impinger (which

yield particle counts) methods. Investigations of the relationship between the two methods

in the mining and milling industry has been carried out by Gibbs and LaChance (1974) [2]

and Dagbert (1976) [3]. The results of those studies did not support the use of a single

conversion factor. It is predictable that there is interest in the relationship between

any two techniques that are used to measure asbestos levels. However, the estimation

methodology for a conversion factor between two observations, both of which are subject to error, must take those errors into account. Thus, a classical least squares regression

1 ine is not suitable.

Any attempt at a characterization of the membrane filter method is complicated by the

fact that the technique known as the membrane filter method has differed between countries

and, even within countries, has actually changed over the years. Part of the explanation

is that sampling and analytical equipment and methodology have undergone change. The effect

has been substantial. For example, it was estimated by Steel (1979) [4] that a British

workplace measurement of 2 f/cm3 in the 1 960 1 s would likely have been on the order of

10 f/cm3 if it had been measured using today's membrane filter sampling and analytical

techniques.

figures in brackets indicate the literature references at the end of this paper.

145
To assess the uncertainty of a workplace measurement for a single day, it is necessary

to characterize the errors inherent in the entire sampling/analytic procedure. However,

when evaluating the uncertainty of an assessment of longer term (i.e., over days, months, or years) exposure, temporal and spatial variation of the dust cloud must be considered in

addition to the error inherent in the sampling/analytic procedures used for a single day.

Leidel, Busch, and Lynch (1977) [5] made some thoughtful contributions on these considerations in the NIOSH (1977) [5] publication on sampling strategies.

The remainder of this discussion will deal just with issues concerning the characterization of the variability of the membrane filter method for a determination on a given

day using only one sample. Much of the investigation of the inter- and intralaboratory

reliability of the method has focused on the analytical equipment and methodology rather

than the entire sampling/analytic procedure. Considerable attention has been given to whether or not a Poisson distribution adequately describes the distribution of fibers on a filter. Both intra-wedge and inter-wedge variability have been investigated. An exhaustive

study of three filters by six counters was reported by Conway and Holland (1973) [6] from

a study for the asbestos industry. That study found more inter-wedge variation on a single

filter than would be predicted by the Poisson distribution. Other studies, such as an intercounter-intermicroscope-intrafi Iter study by Leidel and Busch (1974) [7] and an intralaboratory-intrafilter-intracounter study by Lynch, Kronoveter, and Leidel (1968) [8]

clearly support the hypothesis that empirical variability is much greater than would be

predicted by the Poisson distribution alone. It should be emphasized that the appropriateness of a Poisson assumption for intrafilter variation is not of major importance in the

"bottom- 1 i ne" assessment of uncertainty. To characterize the inherent variability of the

complete process, factors such as intercounter variability and sampling error must be

added to the intrafilter variation. In addition, different laboratories, equipment, and

methods can have a pronounced effect.

In 1976 some of the theoretical problems of the counting procedure were explored and

it was found that the NIOSH counting rules produced a bias on the high side (Cooper et al.,

(1978) [9]). That is, the longer the fiber, the more likely it was to be counted. That

type of bias has implications when fiber size distributions are characterized, although it

may be negligible in many situations. Another interesting result that was found by analytical investigation is the theoretical lower bound on the variation if the distribution of

fibers is actually uniform. By changing the counting rule, the counting error was actually

reduced. In other words, the theoretical lower bound based on the Poisson distribution is

actually lower than 1/E(N), where N is the total number of fibers counted. There is also a very small bias due to the stopping rule (Johns-Manvi 1 le (1976) [10]).

The bottom line for all of these investigations is what sort of uncertainty should be

given to empirical results? It is known that a repeat measurement by the same counter using

the same slide and equipment is subject to error. It is also known that another counter

using the same materials and equipment will introduce more variability than a single counter. If another laboratory is brought into the picture, additional uncertainty is introduced. Additional variability can be attributed to sample replication. There is a miriad

of combinations of variables which can be investigated for attributable variability.

The precision of the method has been characterized using the coefficient of variation. The coefficient of variation has generally been given as a function of the expected

total number of fibers counted. Even through the Poisson does not describe total variation,

the number of fibers counted is still an important factor. Therefore, it is resonable to take the total number of fibers into account in order to estimate coefficients of variation.

The estimation of the coefficient of variation can be very difficult, since large

samples from the distribution of interest are often not available or even possible with

environmental samples. If many observations are available, they often are taken only two

or three at a time. The number of simultaneous personal samples which can be taken on a single worker has obvious limitations. Thus, the statistical comfort that statisticians

derive from large samples is simply not available.

146
The underlying distribution can be important in many applications of statistics. The

underlying distribution is crucial when the variability is expressed as the coefficient

of variation. Normal (or Gaussian) assumptions can be very misleading when the true distribution is skewed to the high values. Nonsymmetrical distributions are likely to be

encountered, especially when they are non-negative and "near zero". In other words, if the

true value is near zero, and negative values are known to be impossible, large negative

errors are ruled out. However, large positive errors are not ruled out.

To illustrate, consider a symmetrical Normal distribution with a mean of 0.2 and a coefficient of variation of 0.4, and a lognormal (i.e., a positively skewed distribution)

with a mean of 0.2 and a coefficient of variation of 0.6. These two distributions are quite different. For example, the middle 95 percent interval for the Normal is from 0.12

to 0.28 (i.e., only 5% of the values will be observed outside this range). The corresponding interval for the lognormal distribution runs from 0.06 to 0.51.

What about sample coefficients of variation from such distributions? Since it is

often difficult to obtain more than two replicate samples, consider samples of size two. A Monte Carlo study of 1000 independent sample coefficients of variation from these two distributions was undertaken and the results are given in figure 1. The sample coefficients of

variation from the two distributions are similar. In other words, it is difficult to distinguish between the two distributions by looking at the sample coefficients of variation

since the range and likelihood of the possible sample coefficients are similar. Of course,

this does not prove anything, it is intended only to illustrate that estimation schemes

based on Gaussian distributional assumptions are not appropriate.

In late 1975, early 1976, J-M conducted an in-house simultaneous sampling replicate

counting study (Johns-Manvi lie (1976) [10]). Ten workplace locations were selected to give

a variety of airborne asbestos fiber concentrations and other contaminants. A variety of

sampling times were also selected. Six simultaneous samples were taken at every sampling

location for each sample time period. The filters were fixed in a generally circular

pattern and separated by 4 to 6 inches. One hundred ninety-two filters were collected.

Those data were submitted to 0SHA as a part of comments with respect to an October,

1975 proposed rulemaking (Johns-Manvi 1 le (1976) [10]). The information submitted eventually led to a change in the counting rules in the NI0SH procedure. Also, NI0SH used those

data to estimate a coefficient of variation curve, expressing the CV as a function of the

total fibers counted in 100 fields. Figure 2 shows the resulting CV curve from Leidel

et al. , (1979) [11].

There are two points that should be made regarding the estimates:

1. Only within-filter and within-laboratory replicates were used.

2. It was assumed that the underlying distribution of errors was normal.

147
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i I I I T i i I I 0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9

VALUES OF SAMPLE COEFFICIENTS OF VARIATION

Figure 1. Cumulative distributions of sample coefficients of variation for samples of size

two from a Monte Carlo study:

1 ,000 Repetitions

Loqnormal Distribution Mean = 0.2 • Normal Distribution Mean = 0.2 -

CV = 0.6 CV = 0.4

148
0.4 5 n

0.05 4

TOTAL FIBER COUNT

Figure 2. Total coefficient of variation as a function of total fiber count

(including pump error). This figure appears as figure 3 on page 81 of

Leidel et al . , (1979).

The curve published and currently used by NIOSH, and the theoretical minimum based

just on Poisson error in counting are shown in figure 3 with a scale change. Analysis of

those same data using a technique which does not rely on a normal assumption is shown in

figure 4, along with the NIOSH curve. (If X and Y are a pair of observations and S

2 is the

sample variance for X and Y, then the coefficient of variation is estimated by the square

root of IS2 /IXY, where the summation is over all possible pairs.)

The data which were not utilized in the NIOSH analysis, between laboratories, are shown on figure 5. It is apparent that the variability between laboratories and filters is

substantially greater than the variability within laboratories and filters.

The curves are, of course, estimates and subject to uncertainty. They have not been

smoothed out, the individual points are presented here. The CV curves have been shown and

calculated as associated with the total number of fibers counted. That point was discussed

earlier. It is also known that there are other factors.

149
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152
This brief discussion of some of the statistical aspects of workplace asbestos measure- ments has emphasized one aspect; the importance of methodology which is not sensitive to assumptions which cannot be. verified. Other important statistical considerations, which

necessarily follow those considered here, such as reliable levels of detection and lowest

levels of reliable measurement, have not been discussed.

References

[I] Ayer, H. E. ; Lynch, J. R. ; Fanney, J. H. A comparison of impinger and membrane filter

techniques for evaluating air samples in asbestos plants, Ann. N.Y. Acad. Sci., Vol.

132 (1965), 274.

[2] Gibbs, W. G. ; LaChance, M. Dust/Fibre relationships in the Quebec Chrysoti le Industry,

Arch. Environ. Health, Vol. 28 (1974), 69-71.

[3] Dagbert, M. Etude le correlation des measures d

1 Empoussierage dans 1' Industrie de

1' Ami ante, Document 5, Beaudry Report (Quebec Committee of Enquiry of Health in the

Asbestos Industry), 1976.

[4] Steel, J. Asbestos Volume 2: final report of the advisory committee, Appendix B, published by Her Majesty's Stationery Office: London, 1978.

[5] Leidel, N. A.; Busch, K. A.; Lynch, J. R. Occupational Exposure Sampling Strategy

Manual, NIOSH Technical Publication 76-131, 1977.

[6] Conway, R. E. ; Holland, W. D. Statistical evaluation of the procedure for counting

asbestos fibers on membrane filters, prepared for AIA/NA, 1973.

[7] Liedel, N. A.; Busch, K. A. An evaluation of phase contrast microscopes for asbestos

counting, NIOSH unpublished in-house report TR-92, 1974.

[8] Lynch, J. R. ; Kronoveter, K. H. ; Leidel, N. A. Validity of the Poisson distribution in dust counting, NIOSH unpublished in-house report TR-83, 1968.

[9] Cooper, D. W. ; Feldman, H. A.; Chase, G. R. Fiber counting: a source of error cor- rected, Am. Ind. Hyg. Assoc. J., Vol. 39 (1978), 362-367.

[10] Johns-Manville. Comments of the Johns-Manvi lie Corporation with respect to the Notice

of Proposed Rulemaking: Occupational Exposure to Asbestos, Federal Register, October

9, 1975, submitted to the public record at the U.S. Department of Labor, Occupational

Safety and Health Administration (April 1976), 1976.

[II] Leidel, N. A.; Bayen, S. G. ; Zumwalde, R. D. ; Busch, K. A. USPHS/NIOSH membrane filter

method for evaluating airborne asbestos fibers, DHEW (NIOSH) Publication No. 79-127,

1979.

153
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gaithersburg, Maryland, October 1-3, 1980. (Issued March

1982)

SIMULATION OF THE EPA PROVISIONAL METHOD FOR AIRBORNE ASBESTOS CONCENTRATIONS

Terence Fitz-Simons

Data Management and Analysis Division

and

Michael E. Beard

Quality Assurance Division

Environmental Monitoring Systems Laboratory

U.S. Environmental Protection Agency

Research Triangle Park, North Carolina 27711

Abstract

The most expensive segment of the EPA provisional method for mea- suring asbestos in the ambient air and the segment that has the greatest

potential for error is transmission electron microscopy. Consequently

transmission electron microscope analysis is simulated. A hypothetical

filter is loaded with fibers. The fiber lengths and widths are generated

as a bivariate distribution. The location on the filter and orientation

of the filter are generated as uniform random variables. The EPA provisional method is then followed to estimate fiber counts, mass, and size

distribution. A plug is taken from the filter, a grid is defined, the

grid openings are sampled according to the method. The simulation is

presented graphically. Assumptions used in the simulation and the random

and systematic sampling errors are discussed.

1. Introduction

The unique physical properties of asbestos have encouraged widespread use of this

mineral for centuries in a variety of applications. Unfortunately, prolonged exposure to airborne asbestos fibers adversely affects the respiratory system by reducing lung capacity

and recent studies have related various forms of lung cancer to asbestos exposure. The

Environmental Protection Agency (EPA) has moved to control emissions of asbestos because of

its widespread use and hazardous nature [l] 1

. The need to monitor airborne asbestos exists

because of the difficulty in controlling all sources of emissions.

Respirable asbestos fibers range in length from a few micrometers (urn) down to submicrometer sizes. Median airborne asbestos fiber lengths reported in the literature range

from about 0.5 to 5.0 urn, [2,4] and are best measured using electron microscopy. The EPA provisional methodology for airborne asbestos measurements [5] employs transmission electron

microscopy (TEM) for identification and measurement of fibers collected on membrane

filters. The method is an uneasy marriage of statistical sampling and TEM microscopy. The

fibers are distinguished as asbestos by their morphology, crystal structure as determined by

selected area electron diffraction (SAED), and their chemical composition as determined by

x-ray fluorescence (XRF). Fiber count, length, diameter, and calculated mass are reported.

Samplers ranging from high volume (hi vol) with 8 x 10" (406.5 cm2

) filters to personal samplers with 37 mm (8.55 cm2

) diameter filters have been used to collect airborne

asbestos. A 3 mm diameter circle is taken from these filters for analysis by TEM. The

3 mm circle is placed on a 200 mesh grid and up to 10 grid openings are examined according

1 Figures in brackets refer to the literature references at the end of this paper.

154
to a strict counting protocol. The grid openings are approximately 75 to 100 um square

(10 4 cm2) and to relate the fibers on the grid to the original filter size multiplication

factors between 10 2 and 10 5 are needed. The importance of a well defined counting protocol

is obvious.

Testing the counting protocol has been accomplished by repeated experimental observations by TEM which are tedious, time consuming, and expensive. This paper describes a computer simulation of sampling asbestos fibers and tests various protocols for counting and

determining mass of the fibers. The simulation program was developed on the UNIVAC 1100 at

the EPA NCC. The program uses IMSL [3] and Tektronix software and Tektronix hardware.

Steps i n the simulation process :

1. Input all parameters pertinent to the simulation.

a. Parameters for the bi-variate distribution of length and width of the filters.

The program has been set up to handle four distribution types: normal, lognormal, three-parameter lognormal, and four-parameter lognormal (Johnson S^).

b. Fiber density and variation. The actual number of fibers on the filter is con- sidered to be a normally distributed random number. The input fiber density is

the mean of this number and the variation is the variance of this distribution.

c. Filter size and shape. The shape of filter can be either circular or rectangular. The dimensions are defined by length and width or diameter. Also a limit is defined as to how close to the edge a plug can be taken for sampling

purposes.

d. Size of TEM grid openings and dimensions of the TEM field of view. The grid

openings are considered to be square, and the field of view is considered to be

rectangul ar

2. Construct a seed for the various random number generators used in the simulation.

This seed is based on the time of day the program is run and is therefore changed for

every run since identical seeds produce identical sequences of random numbers.

3. Print out all parameters input to start the simulation.

4. Locate the 3.0 mm plug on the filter surface at a random point on the filter. The

plug is located so that it will not be too close to the edge of the filter.

5. Orient grid openings on the plug, the grid is oriented at a random angle. The grid

is simply defined as a series of coordinates corresponding to the grid opening size.

There is no space between grid openings as there is on real TEM grids.

6. Select 10 grid openings at random. The 10 grid openings are selected at this point so that fiber information may be stored for sampling purposes later on. Only 10 are selected because no more than 10 can be sampled according to the provisional method.

7. Generate fibers for the entire filter.

a. Generate length and width.

b. Generate location and orientation on the filter. The fiber location is generated

as a uniform random placement on the filter. The orientation is a random

angle.

c. Compare location with selected grid openings and store if necessary. The fiber

location is generated at a uniform random placement on the filter. The orientation is a random angle.

d. Accumulate statistics for fibers on the filter.

155
8. Print out statistics for fibers on the filter.

9. Make an initial count on 1st selected grid opening. Based on this count, follow

procedures for medium or low loading (high loadings are not counted).

10. Print out statistics for fibers in the samples.

11. Compute errors of estimation in sample. Example of output can be found in

Appendix A.

2. Conclusions

1. The aspect ratio cutoff of 3.0 causes bias in fiber count estimation and mass estimation. For six runs at low loading, the average percent difference between estimated

and true fiber count was -34.44. The average percent difference for mass was 1.96.

The large bias in count is expected due to the aspect ratios criteria. The lack of

bias was not expected for the estimates of mass. One possible explanation for this

is that most of the mass is concentrated in the higher fibers which have a higher

probability of falling in the sample, whereas the smaller fibers and the larger fibers

are counted the same.

For five runs at medium loading, the average percent difference for estimated

mass was -30.13 and the average percent difference for estimated count was -34.93.

The mass has just as high a bias in this case. One possible explanation for this is

that when sampling via the smaller field of view, the larger fibers will have a high

probability of being truncated by the field of view; therefore, the estimate loses

mass.

2. The border line between low and medium loading levels causes problems because not enough random fields of view can be found to get the required number of fibers in the

sample.

3. We found that the smaller fields of view (< 5x5 urn) caused serious truncation of fiber

length distribution, and on border line medium loadings the same problems of not being

able to define enough random fields of view to take a large enough sample occurred.

2.1 Future Problems to be Addressed

The simulation program can be used to test different sampling protocols for increased

precision, to characterize the effects of a clumped fiber distribution across the filter.

The simulation will also generalize assumptions such as non-bending fibers and no operator

error.

References

[1] Code of Federal Register, Title 40, Part 61, Subpart B, "National Emission Standard

for Asbestos," U.S. Govt. Printing Office, Washington, D. C. Revised July 1, 1977.

[2] Ettinger, H. J.; Fairchild, C. I.; Ortiz, L. W. ; Tillery, M. I. Aerosol Research and

Development Related to Healty Hazard Analysis, January 1 through January 30, 1973,

LA-5359-PR, Los Alamos Scientific Laboratory of the University of California.

[3] IMSL Subroutine Library, Vols. 1 and 2, International Mathematical and Statistical

Library, Inc. , Houston, Texas, 1975.

[4] Johnson, W. ; Berner, A.; Smith, G. ; Wesolowski, J. Experimental Determination of the

Number and Size of Asbestos Fibers in Ambient air, Report No. ARB-R-3-688-76-45.

California Air Resources Board, January 1975.

[5] Samudea, A. V. ; Harwood, C. F. ; Stockham, J. D. Electron Microscope Measurement of

Airborne Asbestos Concentrations, A Provisional Methodology Manual. EPA-600/2-77-178,

1978.

156
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**************************************************

STATISTICS FOR FIBFRS ON

NUMBER OF FIBERS - MEAN LEN6TH - 1.59

VARIANCE = 8. 9279

MEAN UIOTH = .16U4

VARIANCE = .0087

COVARIANCE = .03 13

CORRELATION = .184203

ASPECT RATIO < A VF » = 10

VARIANCE (AR) = 392.7113

MASS C6MS) = . 00251193*1

FIBER COUNT ON PLUG =

FILTER

1570815

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161
National Bureau of Standards Special Publication 619. Proceedings of tne NBS/EPA Asbestos

Standards Workshop held at NBS , Gai thersburg , Maryland, October 1-3, 1980. (Issued March

1982)

ANALYTICAL ERRORS IN ASBESTOS ANALYSIS BY ANALYTICAL ELECTRON MICROSCOPY

Eric B. Steel, John A. Small, and Patrick Sheridan

Gas and Particulate Science Division

Center for Analytical Chemistry

National Bureau of Standards

Washington, D. C. 20234

Abstract

Instrument and operator related error can combine to yield large

inaccuracies in analyzing asbestos samples by the analytical electron

microscope. Imaging, diffraction, and mechanical stage problems can yield cumulative errors causing results of the same area of a sample to vary by a factor of two or more. Operator precision and bias vary

considerably. The five operators for which the data was collected have

an average tendency to undercount by approximately 10 percent or greater

in the samples investigated (loadings of 25-45 fibers per 200 mesh grid

square, using NIEHS short fiber chrysotile). Most of this undercounting

occurs in the short, single fibrils that are under 0.4 urn in length. At

or below this length our operators have approximately a 50 percent or less chance of detecting the fibers. Since our operators yield com- parable results with other experienced laboratories, we expect that

other operators are also missing small fibers.

1. Introduction

Measurement of asbestos in the ambient environment is currently accomplished using a filter collection procedure followed by an analytical electron microscope (AEM) analysis.

A 1977 multi laboratory study of ambient air samples using unspecified preparation and

counting methods gave results on split samples that varied by several orders of magnitude

[l] 1. Since that time the Environmental Protection Agency (EPA) has been developing a standard methodology, but even using these techniques and the most experienced laboratories,

i nterl aboratory results generally vary by a factor of two or more [2-4]. These differences

may be due to a number of factors including variations in sampling, sample preparation

(including filter homogeneity), instruments, and operators. Since most i nterl aboratory

studies have used split samples and are currently using the same preparation methods, the

variation should be due to instrument, operator, and homogeneity problems. The subject of

this paper is the data collected on operator and instrument related error during the

preparation of asbestos filter research materials at the National Bureau of Standards

(NBS). The paper by Leigh et al . , in these proceedings discusses filter homogeneity.

2. Sample Preparation

The filter research materials are prepared by filtering suspensions containing small

amounts of National Institutes of Environmental Health Sciences short fiber chrysotile [5]

and St. Louis Urban Air Particulate, NBS Standard Reference Material 1648, in filtered

distilled water onto 0.1 or 0.4 urn filters. Indexed 200 mesh copper transmission electron

microscope (TEM) grid specimens are prepared for AEM analysis using the EPA provisional

method for analysis of asbestos in air [2].

1 Figures in brackets refer to the literature references at the end of this paper.

162
3. Counting Procedures

In order to achieve the greatest accuracy possible, three to five operators were required to analyze each grid opening. Two of these operators had 5-6 years experience in

asbestos analysis and the other three operators had accumulated 1.5 to 2 years experience.

Each grid square was traversed at a magnification of 20,000X. Each operator counted

the square without major stage movement or changing samples between counts and also without

discussion of the analysis with the other operators.

4. Instrument Related Errors

Eight transmission electron microscopes (TEM) were tested to find the instrument best

suited to our analytical needs and to get an idea of the problems associated with instrumentation. These included TEMs from most of the major manufacturers. The instruments ranged

in age from 25 years old to brand new and included both 100 kV and 200 kV models.

A number of the TEMs tested were found to have imaging and mechanical problems which

increased the error in asbestos analysis. None of the TEMs in this study was used solely for

asbestos work and all had multiple users with a wide variety of interests. Many of the

problems encountered could and should have been corrected if the instruments were to be used

largely for asbestos work. But this study was intended only as a survey to see the types

and general magnitude of problems that could be expected in the field.

Imaging related parameters affecting asbestos analysis were found to include operating

voltage, brightness, contrast, resolution, working magnification, and diffraction capabilities. Mechanically related errors appeared to be caused mainly by stage translation

control s.

Under most circumstances the analysis of chrysotile on the TEM is best carried out at voltages of approximately 80-120 kV. At these voltages, chrysotile has good contrast with

the carbon support film and the hollow tubular structure, if present, can be observed.

At higher voltages this contrast can be considerably less and analysts can miss detecting

fibers more easily. For amphiboles, contrast is less of a problem, and often higher

voltages are required to attain diffraction patterns from thicker particles.

The brightness, contrast, and resolution of the TEMs used differed considerably. Some

of the older instruments had considerably poorer images than the newer models, causing

operators to miss up to 50 percent of the fibers as well as prematurely tiring operators

due to the strain of trying to see and distinguish fibers. In addition, it was found that

several older 200 kV TEMs operating at 100 kV often had insufficient brightness and contrast

for easy observation of chrysotile. These problems were not encountered on the new 100

and 200 kV models. Half of the eight TEMs used, increased the error, the time of analysis,

and operator fatigue due to poor image quality.

The diffraction conditions and data also varied greatly from instrument to instrument.

On some of the older instruments less than 10 percent of the chrysotile fibers yielded

good selected area diffraction patterns visible to the operator. On instruments that were about three years old or newer, greater than 90 percent of the chrysotile showed good

selected area diffraction (SAD) patterns. This disparity is probably due to a number of reasons including the longer time needed to set up SAD conditions in the older instruments,

the efficiency of the electron optics, phosphors, etc., and the fact that chrysotile's

crystal 1 inity is electron dose dependent.

Magnification also plays an obvious and important role in both the efficiency and

accuracy of asbestos analysis. At 20,000X a single chrysotile fibril 30 nm wide appears

to be approximately 0.6 mm wide with a hollow tube ~0.1 mm wide. This fiber width is reasonably easy for the human eye to see and the hollow tube can easily be observed by

using a binocular microscope. At magnifications lower than about 10,000X these thin fibers

can be difficult to see and are often missed because they are only a few tenths of a millimeter wide. At these lower magnifications it is also more difficult to distinguish

chrysotile from filter artifacts and other non-asbestos particles, so that more time is

163
taken analyzing fibers that are readily distinguishable at 20,000X. At magnifications of

30.000X and higher more traverses are needed to cover a TEM grid square and the area and

time of analysis increases by about 2 1/2 times. In addition any stage traverse error (see

below) will be likewise amplified. Therefore, for reasons of economy and accuracy we have

found the optimum magnification for scanning grids for chrysotile is approximately 20,000X.

The major mechanical problem associated with the TEMs was found to be the mechanical

stage. Stage movement stability and reproducibility is extremely important. During the

analysis of asbestos a grid square approximately 80 urn on a side is traversed at magnifications of about 20,000X. Any wandering of the stage during a traverse can cause areas and

fibers to be missed or reanalyzed. Some of the stages erratically wandered by as much as 30 urn off a straight line traverse, causing asbestos counts to vary radically (over a factor

of 2 difference on the same square). Most stages have some "backlash" when traverse directions are changed, but these often are reproducible and can be corrected by transversing

past the grid square and then back to it before starting the next traverse. The microscope

used for the data collected on operator error in this paper has a stage which wanders <1 urn during a 100 urn traverse and is very consistent and reproducible in its actions.

5. Operator Error

Operator error was looked at once the machine error was controlled and defined. A variety of methods for reporting and comparing data have evolved during preparation of the

filters. Initially, to determine the extent of operator error, each operator only reported

the number of fibers in the same grid square. These initial values showed sporadic variations of over 50 percent. Some of this variability was found to be due to machine related

problems while some was related to individual operator errors. In one effort to reduce this error and correlate the operators' counts, a counting data sheet was designed. This data

sheet was made to mimic an operator's thought pattern during chrysotile counting. Fibers

that were detected during the traverse of the grid square were catalogued by the following

parameters

- the traverse number in which the fiber was found - whether a fiber was characterized by distinct, poor, or no hollow tube structure

- whether chrysotile 1 s characteristic electron diffraction pattern was observed

- whether the ends of the fibers were visible or unobservable due to obstruction by a grid bar or another particle

- whether the fiber was a single fibril or a bundle of one or more fibrils

- and lastly whether the operator counted a fiber as a chrysotile fiber or as another

type of particle.

The purpose of the chart was to identify which parameters have a significant effect on operator error and to determine operator variability and bias.

It was noticed that one operator had a consistently higher deviation from the mean than

did the other operators. Table 1 shows the average of the absolute percent change from the mean for 30 grid squares by operator. The asbestos counts for operator 3 have averaged

15.6 percent higher or lower than the mean count for the five operators. Normalizing to the low deviation values of 6.9 percent of operators 2 and 4 shows that counter 3 had an average deviation from the mean over 2 times greater than that for operators 2 and 4.

Table 1. Tabulation of the Average Absolute Deviation from

the Mean Grid Square Count in Percent by Operator.

Operator 1 2 3 4 5_

Average percent 10.9 6.9 15.6 6.9 12.0

devi ati on

Normalized 1.6 1.0 2.3 1.0 1.7

164
A "high variance" operator needs to count more grid squares and collect more data to obtain an equally reliable asbestos concentration. In a field laboratory with only one operator, the filter may be erroneously blamed for the large variation. In either case, a statistical analysis should give reasonable error bars. However, if an operator is consistently lower or higher than the mean, the bias will only be detected when compared to several other counters and would not be seen on the statistical analysis carried out on the

data of a single counter (as is usually the case for commercial laboratories). Initially,

the problem of operator bias was looked at qualitatively. For each data set, a ranking

value of one through five was assigned to the lowest through the highest counter, respectively. These values for all the counts were summed and averaged per operator to determine

his average placement or ranking. Table 2 shows such a tabulation. If there were no consistent bias by any of the operators, each operator's rank would be about three. It can

be seen from Table 2 that counter 1 has a tendency to be low while operators 3 and 5 have a slight tendency to be high. The total number of high and low counts is also shown for each

counter. Operator 1 has a large number of the lowest counts, while operator 3 has a large

number of both the lowest and highest counts due to this counter's high variance.

Table 2. Average Ranking of Operators for Thirty Grid Squares and

the Number of Low Counts and High Counts by Operator.

Operator 1_ _2_ 3 4 5

Ranking 2.0 3.0 3.4 3 0 3.4

# lows 11 1 8 3 3

# highs 1 2 14 3 8

An attempt was made to cross reference the counts made by the five operators to discover the cause or causes of the variation and bias. It was found that only the longer

fibers could be cross referenced and it became obvious that all operators were not counting

the same fibers. To prove this and to quantitate what the operators were counting, a system

was devised that accurately located each fiber observed by each operator.

A digital voltmeter was put on the AEM's x and y specimen position controls. This

gives the operator the capability of recording the stage position within approximately a micrometer. Each counter records the position of a fiber as well as the other data discussed above directly into a computer. Later, after all the operators have analyzed the grid

square, a map is made (figure 1) which shows the position of each fiber for each operator.

The fiber count on this grid square is then verified by having an operator reanalyze any

fibers on which there were questions. This includes any fibers that were found by only one operator, or any fibers whose identification is disputed because of ambiguous morphology or diffraction data. In this way, a count approaching as closely as possible to the true

amount of asbestos is derived. Prior to the cross referenced and verified counting method

we used the average value as the number of fibers in the grid square. Table 3 compares the

individual and average operator counts with the verified counts. Thus far, the verified

counts have been greater than or equal to the average counts.

Morphology, diffraction, and sometimes chemistry determined with an energy dispersive

x-ray detector are used to positively identify chrysotile. Because of beam damage to the

crystal 1 inity of the chrysotile only the first operator in our repetitive counting technique uses electron diffraction. This means that the other operators, using only mor- phology, are somewhat more apt to record false negatives and false positives than a field

laboratory that used diffraction. This is especially true of the false positives which

should be almost non-existent, since none of the fibrous minerals with diffraction patterns

similar to chrysotile have been found in our samples. Table 4 shows the breakdown of the errors. Most of the chrysotile fibers that were not counted were simply missed, i.e., only

a small number were misidentif ied as the other. Almost all of the false negatives were

fibers which showed poorly defined or no hollow tube structure and were placed in the

"other" category due to lack of diffraction data.

165
0 50 100 150 200

Figure 1. Geographic distribution of chrysotile fibers on a TEM grid square as analyzed by four operators. The small circles, triangles, squares, and

x's represent individual chrysotile counts for each operator. The large

solid circles are verified chrysotile fibers and the dashed circles are verified "other" particles originally counted as chrysotile.

Table 3. Comparison of Operator Counts and Mean Counts to Verified Count on 75 um x 75

Grid Opening.

Grid / Operator 1 2 3 4 5 Mean Count Veri f i ed

1 20 19 17 18.7 20

2 22 18 21 20. 3 24

3 21 13 21 18. 3 21

4 30 24 21 23 24.5 29

5 24 31 25 25 26. 3 31

6 18 17 19 18.0 18

7 24 23 25 24.0 29

8 24 22 18 27 22.8 24

9 . 22 20 27 20 22.3 24

10 16 19 21 21 20 19.4 22

11 19 16 13 18 16.5 20

12 19 18 20 14 17.8 20

13 31 24 20 25 25.0 26

14 26 19 21 22.0 22

15 27 28 28 30 28.3 30

166
Table 4. Percent Deviation from the Verified Counts by Operator.

Operator D

Percent Fibers

Not Observed

Percent False

Negati ves

Percent False

Positives

•10.62

-2.95

+7.08

•11.20 -29.27 •9.38

-1.30 -0.81 -3.13

+1.82 +14.23 +5.21

One factor correlating with operators not observing fibers is fiber size. Figure 2 shows a graph of the fraction of operators finding fibers of a given length. The scatter

of the points is too large to get a well defined fit, but the trend toward fewer operators

seeing the smaller fibers is apparent. Once the chrysotile fiber length is below approximately 0.3-0.4 urn our operators have at best a 50 percent chance of finding it. This is

probably true for other operators as well.

CO

cro

<

orLU

CL

o

o

o

o

<

en

m-x x)<x Ai x

1.00 -X X0.75

0.50 -

0*25

0*00 —r~*?—i

i

' i—i—i—i—'—i—i—i—i

1—i—i—i—i——i—i—i—i1—i—i—i—

0 1 E 3 4 5 6

AVE. FIBER LENGTH (/*m)

Figure 2. Fraction of operators observing a given length chrysotile fiber

(averaged for each grid square).

The other major error was associated with the way each operator counted fiber

bundles. Our samples contained very few complex bundles of chrysotile fibers, and even though all the operators used the same algorithm for counting bundles there were enough

ambiguous cases to account for up to approximately a 5 percent error. On samples that

contain large percentage of complex asbestos fiber bundles, such as some occupational

samples, one may expect much larger errors due to operator differences in counting the

bundles.

167
6. Conclusions

Machine related error can account for as much as approximately 100 to 200 percent

variation in chrysotile grid counts. Only instruments with excellent imaging and precise

stage controls should be used. Five out of the eight instruments used in this study were considered unusable for routine asbestos analysis because of deficiencies in image quality,

diffraction capability, and/or stage movement precision.

Accuracy and precision vary considerably from operator to operator. This variation

is not necessarily consistent through time, on the first filter operator 1 showed a definite

bias, displayed as low counts, when compared to the other counters. On the second filter

this relative bias was not observed. Among the five operators used in this study, the

average variation for each operator ranged from approximately 7 to 16 percent of the mean number of fibers counted. All of the operators have a definite tendency to undercount

fibers. Most operators failed to observe approximately 10 percent of the chrysotile fibers

in our samples, but one operator missed an average 29 percent. The majority of the fibers

missed are short single or double fibrils below approximately 0.4 pm in length. Below this

length, our operators have approximately a 50 percent chance or less of observing the

fiber. In round robin tests our operators have had comparable results to other experienced

laboratories, thus implying that operators outside of this NBS study can be expected to demonstrate similar biases. Variation in counting the number of fibers in bundles is the

second most common source of error among our counters.

References

[1] "Montgomery County Asbestos Study", EPA Internal Report, U. S. Environmental Protection

Agency, Environmental Monitoring Systems Laboratory, Environmental Monitoring Division,

Research Triangle Park, North Carolina 27711, October 1977.

[2] Samudra, A. V.; Harwood, C. F. ; Stockham, J. D. Electron Microscope Measurement of

Airborne Asbestos Concentrations, EPA Report E PA- 600/2- 77- 1 78, August 1977, U. S. Environmental Protection Agency, Research Triangle Park, North Carolina 27711.

[3] Anderson, C. H. ; Long, J. M. Preliminary Interim Procedure for Fibrous Asbestos,

U. S. Environmental Protection Agency, Analytical Chemistry Branch, Athens, Georgia

30605, July 1976.

[4] Evaluation and Optimizing Electron Microscope Methods for Characterizing Airborne

Asbestos, EPA Report No. 600/2-78-038, U. S. Environmental Protection Agency, Research

Triangle Park, North Carolina 27711, June 1978.

[5] Campbell, W. J.; Huggins, C. W. ; Wylie, A. G. Chemical and Physical Characterization

of Amosite, Chrysotile, Crocidolite, and Nonfibrous Tremolite for Oral Ingestion

Studies by the National Institute of Environmental Health Sciences, U. S. Bureau of

Mines Report of Investigation 8452, Avondale Research Center, Bureau of Mines,

Avondale, Maryland 20782, 1980.

168
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gaithersburg, Maryland, October 1-3, 1980. (Issued March

1982)

STATISTICAL CONSIDERATIONS IN THE PREPARATION OF CHRYSOTILE FILTER REFERENCE MATERIALS:

FILTER HOMOGENEITY

Stefan Leigh, Eric Steel 1

, John Small 1, Patrick Sheridan 1

, and James Filliben

Statistical Engineering Division

Center for Applied Mathematics

National Bureau of Standards

Washington, D. C. 20234

Abstract

The data discussed shows that asbestos fibers can be deposited in a manner compatible with a Poisson statistical process when using careful

liquid filtration onto Nuclepore filters and when using a fiber density

of approximately 10-70 fibers per grid square. However, this hypothesis

should be tested for each filter using methods similar to those

described. The analysis also shows that such filters can be homogeneous

enough to allow individual sections of a filter to be considered as representative of the whole filter sample.

Note: In order to adequately describe materials and experimental

procedures, it was occasionally necessary to identify commercial

products by manufacturer's name or label. In no instance does such

identification imply endorsement by the National Bureau of Standards nor does it imply that the particular product or equipment is necessarily the

best available for that purpose.

1. Introduction

Measurement of asbestos in the ambient environment is currently accomplished using a filter collection procedure followed by an analytical electron microscope (AEM) analysis.

A 1977 multi laboratory study of ambient air samples using unspecified preparation and

counting methods gave results on split samples that varied by several orders of magnitude

[l] 2. Since that time the Environmental Protection Agency (EPA) has been developing a standard methodology, but even using these techniques and the most experienced laboratories,

i nterlaboratory results generally vary by about a factor of two [2-4]. These differences

may be due to a number of factors, including variations in sampling, sample preparation

(including filter homogeneity), instruments, and operators. Since most i nterl aboratory

studies have used split samples and are currently using the same preparation methods, the

variation should be due to instrument, operator, and homogeneity problems. The subject of

this paper is the data collected on filter homogeneity during the preparation of asbestos

filter research materials at the National Bureau of Standards (NBS).

2. Sample Preparation

The filter research materials are prepared by filtering a suspension containing small

amounts of National Institutes of Environmental Health Sciences short fiber chrysotile and

St. Louis Urban Air Particulate, NBS Standard Reference Material 1648, in filtered distilled

x Eric Steel, John Small, and Patrick Sheridan are in the Gas and Particulate Science

Division, Center for Analytical Chemistry.

2 Figures in brackets refer to the literature references at the end of this paper.

169
water onto 0.1 or 0.4 |jm Nuclepore filters [5]. Indexed 200 mesh copper transmission

electron microscope (TEM) grid specimens are prepared for AEM analysis using the EPA provisional method for analysis of asbestos in air [2].

3. Counting Procedures

In order to achieve the greatest accuracy possible, three to five operators were required to analyze each grid opening. Two of these operators had 5-6 years experience in

asbestos analysis and the other three operators had accumulated 1.5 to 2 years experience.

All the data reported in this paper was collected on either a JEOL 100CX or JEOL 200CX TEM

operating at 100 kV and approximately 20,000X magnification. Additional counting information is contained in the paper by Steel et al., in these proceedings.

4. Statistical Analysis of Fiber Distribution Data

The validity of the common Poisson assumption regarding the underlying distribution

for grid counts in the range of loading densities tested, the question of homogeneity of

the fiber counts over the surface of a filter, and the accuracy to be anticipated in

counting filters prepared to uniform loading density specifications are discussed. This

information is based on the results from one trial filter (five operators counting, 0.1 urn Nuclepore) and one uniform filter (three to five operators counting, 0.4 um Nuclepore).

The overall objective of the NBS research filter, and ideally of filter samples collected in the field, is to be able to control the process of fiber deposition and analysis

to such a degree as to be able to measure the average count densities to within some preassigned relative error, where by relative error we mean an estimate of error - e.g.,

standard deviation - divided by the best estimate of count average. We replicate at two

levels during the analysis procedure. We replicate within the grid square by having more than one operator count each square and across grid squares by analyzing many randomly

chosen filter areas (grids) and grid squares.

5. Distribution Assumptions

It is easiest to characterize a process statistically if that process is one characterized by random sampling from a fixed (though not necessarily known) distribution with

fixed mean and fixed variance. The classical assumption in many types of counting situations is that the underlying distribution for the counts is Poisson [6]. In these cases two implicit underlying assumptions are made: randomness and independence. In this

context randomness refers to an "unpredictability" in the number of fibers deposited on any one cell of the counting grid. Independence means that no physical interaction is

occurring among the chrysotile fibers during the deposition process.

Assume a series of grid counts, (x.), has been obtained for a given filter. If the

Poisson assumption is met for the filter, then the Poisson parameter, A, as estimated by

the mean count (x), provides the best single summary of the filter's loading density. For

such a Poisson filter the reported density will be

x ± Vx7n

(1)

where N is the number of grid squares counted.

As a point of reference, typical contemplated NBS filter loading densities would be

approximately 10-100 fibers per grid square. Figure 1 depicts the Poisson coefficient of

variation (yfk/X) for different A (loading densities) within this range. This curve is only

a limiting curve on the relative errors obtainable for filter counts following these

distinct Poisson processes in the sense that given a filter whose count distribution is

known to be Poisson, of known A, with the true counts on all of the grid squares known, then

the relative standard deviation for the filter (the population relative standard deviation)

will be the figure read off the curve. This curve is not, however, to be interpreted as saying that we cannot get as arbitrarily close as we like to the true A in our estimation.

170
35

30 -

25

20 -

15 -

10

0 20 40 60 80

COUNT

REL ERROR « 100 X (SGRT(X)/X)

Figure 1. Poisson coefficient of variation.

100

We can in fact get arbitrarily close to the true K by sampling at a sufficiently high

frequency (N). In fact, if we can assume Poisson, then for prespecif ied maximum allowable

uncertainties p in our estimates of A, a very simple approximate relationship can be derived

which specifies exactly how large N should be to keep the relative error below p. If p equals the maximum allowable uncertainty (specified in decimal terms, e.g., 0.10, 0.15,

etc.) in our estimate of A and is fixed, then by the standard formula for the sampling

variation of the mean (ignoring finite population correction factors)

2(SA/N)

(2)

where the factor 2 is chosen because two standard deviations translates approximately into

95 percent confidence. This can be written

N > (—) \(p/2) x/ v (p/2) x.

Under the assumption of Poisson x = A and s = -JX. Therefore

NA > 4/p

2

(3)

(4)

171
which tells us how large to choose N to get our (x.) based estimate of to within ±p percent

of the true value.

Figure 2 shows the ranges in N corresponding to reasonable choices for p values for

different loading densities. The formula and curves emphasize that ultimately it is the

total number of fibers counted - the product NA. - that determines how close we come to the true value of A for a filter, as we should expect. In field situations, it may be

possible to vary the dilution or time of sampling to attain the most convenient combination

of N and A such that the product is greater than or equal to 4/p 2

. Thus, where applicable,

the Poisson assumption provides a succinct formula determining the degree of precision to be expected in estimating the density parameter as a function of the number of grid squares

counted.

80 I

:

t 60 - \

\\

\ X=25\

40 -

0,035 0*050 0.075 0.100 0.135 0.150 0.175 0.200

P

LOADING DENSITIES (TOP TO B0TT0P1 ) 25,50,100

Figure 2. N for maximum allowable uncertainty in estimate of A.

In fact, experience at some other laboratories suggests that at low and medium loading

densities - the 10 to 70 fibers/75 um2 grid square range - a Poisson distribution of the

counts obtained is to be expected, but at higher ranges the empirical count distributions

may in fact not be explainable by Poisson. At these higher loading densities, fibers may

not be acting in an independent fashion due to clumping or some other process during deposition. Thus one of the Poisson assumptions, independence, would not hold. But alternative

distributions - such as the negative binomial or various compound or generalized distributions used for modeling clustered spatial distributions - may be suitable under these

conditions [7]. For these reasons it is essential that any assumption of Poissonness or any alternative model for the distribution of fibers on the filter be tested.

172
One important limiting factor on all of the checks that were done for Poissonness

(likely to be even more true for commercial laboratories with greater time restraints) is

the small amount of data available. Twenty-five areas on the first trial filter and 38 areas on the second filter were analyzed. This precludes doing certain available checks of

Poissonness, such as a simple graphical test described by Hoaglin [8].

Classical distributional tests that are available and were used include comparison of

moments, chi-square goodness of fit, Kolmogorov-Smi rnov , and probability plotting.

Figure 3 shows the distribution of the counts over the indicated surfaces on the first

filter. The numbers listed for each lettered area are the average counts for five grid

squares counted by five operators each.

A 50,51,53,53,39

B 45,37,41,53,49

C 43,44,40,50,51

D 43,26,46,47,54

E 44,47,42,56,44

Figure 3. Schematic of chrysotile counts collected on the first

uniform 47 mm Nuclepore filter.

173
Figure 4 shows the actual occurrence pattern of the counts over the surface of the

second filter. The picture is a composite, combining counts based on averaging over 3-5

operators counting the same grid square with count values based on the verified method

(starred values) described by Steel et al., in these proceedings. This is done, because,

where available, the verified data do in fact represent the best estimates for the true

grid square counts, while where such values are unavailable the averages over the number of

operators represent the best estimates of "true" counts.

r53~

32

1T(3^

20

30

28

DC

28 35

/28~

/ 16

26

23

22

18

~29\

23\

28*

21

22*

30

'23*

1)7

\20*

\I9 —

24*

21

31*

29

24*

24

22*

20 — /

26*

<20

Figure 4. Schematic of chrysotile counts collected on the second uniform 47 mm Nuclepore filter. Each square represents a filter section from which a TEM specimen grid was prepared. The numbers are the chrysotile counts

from two grid squares in each of the grids analyzed.

Figures 5 and 6 show histograms of the counts obtained on the two filters. Aside from

an apparent biomodality in each, both distributions look reasonably symmetric and normallooking - as they should for loading densities above approximately 10-11 fibers per grid

square.

Comparison of moments, shown in Table 1, show reasonable agreement with theoretically

predicted variance for the first filter and good agreement in both second and third moments

for the second filter. On the second filter the mean does in fact equal the variance out to four significant figures (0.2459) - as it should for Poisson distributed data - and an actual skewness coefficient of 0.245 compares favorably to a Poisson-predicted K x/2 of

0.202. Fourth moments (kurtosis) do not compare as well, possibly because of the valleys

in the middles of the histograms.

174
0

8

j ,

1

,

0 80 40 60 80 10.0

Figure 5. Histogram of chrysotile counts (per grid square) on the first filter.

0

1—j—j—n

1—r~i—n—"-i

i 'i

1 >'1 1

' i' i

' i

1 ,u

~f 1

' »'i

' i i i i i i r—

r

-10 0 10 20 30 40 50 60

Figure 6. Histogram of chrysotile counts (per grid square) on the second, uniform filter.

175
Table 1. Comparison of Moments. ________ Actual --------

Mean Variance Skewness Kurtosis

1st (25-count) Filter

45.86 43.19 -0.976 4. 39

________ Expected --------

Mean Variance Skewness Kurtosis

45.86 45.86 0. 148 3.02

2nd (38-count) Filter

24.58 24.57 0.245 1.98 24. 58 24.58 0.202 3.04

If censor the one outlying count (cf Histogram) of 26, compute a skewness (coefficient) of

-0.08.

Probability plots for assumed Poisson distribution with parameters equaling the

respective average counts are shown in figures 7 and 8. The plots are of the ordered

observed counts vs. "typical" ordered counts that would be predicted if the distributions

were in fact Poisson. The predicted values used in these particular plots are the median

values of the sampling distribution of the ith order statistic from the appropriate Poisson

distribution [9]. If the resulting plot is reasonably linear it provides evidence that the

underlying distribution of the counts is indeed Poisson. The nearness to linearity can be

quantified with a correlation coefficient. The two plots here both look quite linear with

correlation coefficients of 0.95 and 0.98, respectively. Again, censoring the one outlying

count of 26 would improve the first plot.

cO

56.C01

52.25

48.50-

44.75-

41.00-

37.25-

33.50-

29.75

26.00-

35

• • e

• o

e o

e • d

33 4! 44 47 50 53 56 59

Expected counts

Figure 7. Probability plot of first filter.

176
tfi

o

o

o

o

>o

CO O

35.00'

30.25-

25.50-

20.75-1

16.00- o

© 0

O ©

a

o © o •

©

© © o

o

o «

15

1 I 20 -1

1 r

30 35

Expected counts

Figure 8. Probability plot of second filter.

Equiprobable category chi-square goodness-of-f it tests were done for each filter, the

results of which are shown in Table 2 and figure 9. Equiprobable partitioning is used here

because of the relatively small number of counts available. Equiprobable partitioning means choosing intervals over the range of count values such that the theoretically expected

values (of occurrence of count values in that range) are the same. This is done to avoid

inflated contributions to the chi-square statistic from zero count cells and to avoid

weighting the smaller expectation values occurring in the tails of the distribution. In all instances - each of the five equipartitions of each filter - the test is seen to be

compatible with an assumption of Poissonness.

Kol mogrov-Smi rnov (one sample) test of goodness-of-f i t was performed and also confirmed

the compatibility of the frequency distribution of counts with Poissonness for the two

filters as shown in Table 3.

177
Table 2. Equiprobable Chi-square.

Number of Partitions

1st (25-count) Filter

df

Observed

Chi-Square

Cumulative distribution function value of

observed chi-square. The value is from the

null distribution of the x

2 i.e., the distribution of x' when H (Poisonness) is true,

o

' 2 statistic -

' 2 which results

1 . 04

3.32

1.60

2.60

3.84

69%

81%

34%

37%

43%

2nd (38-count) Filter

0.21

4.32

2. 79

3.68

6.21

35%

57%

55%

71%

10

8 -

4 -

0

Figure 9.

10 15 20

Chi-square probability density function for five degrees of freedom.

(1st filter, 7-partition case)

178
Table 3. Kolmogorov-Smi rnov Test.

D sup S(x) - P(x) Percent point value of observed D. The percent

point value is from the null distribution of

the D statistic - i.e., the distribution of D which results when H (Poisonness) is true.

S empirical CDF

P Poisson CDF

For 1st (25-count) Filter

0. 065 20%

For 2nd (38-count) Filter

0. 158 70%

6. Filter Homogeneity

An important characteristic of both reference filter standards and field analysis

filters is homogeneity. Intuitively, the homogeneity of a filter refers to randomness of

scatter plus uniformity, where we can define uniformity as meaning that the projections of

the counts occurring on the grid onto the x and y axes are near uniform.

Two checks of homogeneity were performed. The first was done specifically to see if

there was a discernible radial gradient in the filter density. A quick visual test of the

existence of such a gradient was performed by subtracting off the mean count value (x) for

the entire filter from each of the individual grid square counts, and coding the resulting

residuals with respect to a size comparison with the standard deviation for the counts over

the entire filter: positive residual less than one standard deviation are coded "+"

,

negative residuals less than one standard deviation are coded positive residuals

greater than one standard deviation are coded "++", and negative residuals greater than

one standard deviation are coded Figure 10 depicts these coded residuals over the

surface of the second uniform filter. Aside from a higher density patch in the northwest corner of the figure, no consistent radial density gradient is evident. One possible

explanation of the high density patch might be a slight tilt to the filter during the

preparation procedure.

A second test of homogeneity was done by partitioning the set of counts from the second

filter in various ways to mimic physically slicing the filter as shown in figure 10. Then

the means and variances of the distributions of counts resulting on the various slices were compared using a t-test (not assuming equal variance) for the means and F-test for the

variances. The results are shown in Table 4. The mean counts and variances are comparable

in almost all cases at 5 percent significance. Even for partition number 3, where one might

expect a difference in the mean loadings to be detected from the residual plot shown in

figure 10, no difference was seen. Another method of analysis would have been to do an AN0VA (F-test) on each partition. Generally, the "all possible t-tests" approach leads to more significant differences than the F-test approach.

179
180
Table 4. Results of t- and F-Test.

Means Variances Means

Parti ti on Compar i ng Comparabl

e

Comparabl

e

1 2 3 4 5

1 1-2 Y Y 26 24

1-3 NO Y 26 23

z-j Y Y 24 23

2 1-2 Y Y 25 26

1-3 Y Y 25 23

2-3 NO

• Y 26 23

3 1-2 Y Y 25 25

1-3 Y Y 25 24

2-3 Y Y 25 24

4 1-2 Y Y 28 24

1 - J Y Y 28 24

1-4 Y Y 28 25

1-5 Y Y 28 22

2-3 Y Y 24 24

2-4 Y Y 24 25

2-5 Y Y 24 22

3-4 Y Y 24 25

3-5 Y Y 24 22 4-5 Y Y 25 22

5 1-2 marginal margi nal 25 25

1-3 Y Y 25 27

1-4 Y Y 25 23

1-5 Y Y 25 21 2-3 Y Y 25 27

2-4 Y Y 25 23

2-5 Y Y 25 21 3-4 NO Y 27 23

3-5 NO NO 27 21 4-5 Y Y 23 21

7. Conclusions

There have been several discussions of distributions of fiber counts on filters

[10-13], relating the merits of various assumptions concerning the underlying distribution

of counts.

This paper shows that at least for liquid filtration of chrysotile onto 0.1 and 0.4 urn Nuclepore filters with fiber loading densities in the range of 10-70 fibers per 200 mesh

grid square, one may expect a Poisson distribution. This hypothesis can and should be

tested for every filter. Once the tests for Poissonness have been passed, arbitrarily

precise loading estimates can be established using the above methods and reported with the

filter analysis.

A carefully characterized filter has been shown to be homogeneous on a level that

allows sections of the filter to be considered representative of the whole filter. The next

step of this research is to use such well-characterized filters to carry out split sample

round robins with experienced laboratories to better characterize instrument and operator

effects on asbestos analysis.

181
References

[1] Montgomery County Asbestos Study. EPA Internal Report. U.S. Environmental Protection

Agency. Environmental Monitoring Systems Laboratory. Environmental Monitoring

Division. Research Triangle Park, NC 27711; October 1977.

[2] Samudra, A. V. ; Harwood, C. F. ; Stockham, J. D. Electron Microscope Measurement of

Airborne Asbestos Concentrations. EPA Report EPA-600/2-77- 1 78. U.S. Environmental

Protection Agency. Research Triangle Park, NC 27711; August 1977.

[3] Anderson, C. H. ; Long, J. M. Preliminary Interim Procedure for Fibrous Asbestos.

U.S. Environmental Protection Agency. Analytical Chemistry Branch. Athens, GA

30605; July 1976.

[4] Evaluating and Optimizing Electron Microscope Methods for Characterizing Airborne

Asbestos. EPA Report No. 600/2-78-038. U.S. Environmental Protection Agency.

Research Triangle Park, NC 27711; June 1978.

[5] Campbell, W. J.; Huggins, C. W. ; Wylie, A. G. Chemical and Physical Characterization

of Amosite, Chrysotile, Crocidolite, and Nonfibrous Tremolite for Oral Ingestion

Studies by the National Institute of Environmental Health Sciences. U.S. Bureau of

Mines Report of Investigation 8452. Avondale Research Center. Bureau of Mines.

Avondale, MD 20782; 1980.

[6] Student. On the Error of Counting with a Haemocytometer. Biometrika 5: 351-360;

1907.

[7] Rogers, A. Statistical Analysis of Spatial Dispersion, the Quadrat Method. London:

Pi on Limited; 1974.

[8] Hoaglin, D. C. A Poissonness Plot. Amer. Stat. 34(3): 146-149; 1980.

[9] Filliben, J. Testing Basic Assumptions is the Measurement Process, in ACS Symposium

Series No. 63. Validation of the Measurement Process; 1977. 30-113.

[10] Hallenbeck, W. H. ; Chen, E. H. ; Patel -Mandl i k , K. Precision of Analysis for Waterborne Chrysotile Asbestos by Transmission Electron Microscopy. Bull. Environ.

Contamination and Toxicology 17(5): 551-558; 1977.

[11] Millette, J. R. ; Clark, P. J.; Pansing, M. F. Sizing of Particulates for Environmental Health Studies. Scanning Electron Microsc. 1: 253-358; 1978.

[12] Chatfield, E. J. Preparation and Analysis of Particulate Samples by Electron Microscopy, with Special Reference to Asbestos. Scanning Electron Microsc. 1: 576-577;

1979.

[13] Chatfield, E. J.; Dillon, M. J. Some Aspects of Specimen Preparation and Limitations

of Precision in Particulate Analysis by SEM and TEM. Scanning Electron Microsc. 1: 487-496; 1978.

182
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gaithersburg, Maryland, October 1-3, 1980. (Issued March

1982)

REFINEMENTS IN THE EPA PROVISIONAL METHODOLOGY

George Yamate

I IT Research Institute

10 West 35th Street

Chicago, Illinois 60616

and

Michael E. Beard

U.S. EPA/RTP

QAD/EMSL, MD-77

Research Triangle Park, NC 27711

Abstract

Although the electron microscopical method is the best available

to assess the asbestos minerals in the environment, the credibility of

the analytical technique is weakened due to variations in the methodology with resulting diversity of data. The EPA Provisional Methodology on the electron microscope measurement of airborne asbestos concen- trations was developed from a study under EPA Contract No. 68-02-2251

to evaluate the various electron microscopic methods to arrive at an optimum composite procedure. The lack of properly defined standard

procedures restricted its acceptance as the standard analytical

technique.

The present study under Contract No. 68-02-3266 investigated the

problem areas of: (1) sample collection; (2) sample transport; (3)

sample preparation procedures; (4) identification of particles as fibers; and (5) verification of asbestos materials. The critical

parameters of which filter type (polycarbonate or cellulose ester

type), transfer method, and fiber identification were also examined.

The expertise of the particle technologist and the electron microscopist have been combined to refine the EPA Provisional Methodology to obtain a more consistent analysis. Three levels of analytical sophistication are advocated.

1. Introduction

The EPA Provisional Methodology was first published in August, 1977. A revised edition

dated June 1978 was subsequently distributed. The methodology based on the electron microscope measurement of airborne asbestos concentration was developed under EPA Contract No. 68-02-2251. Various electron microscopic methods were evaluated to arrive at an optimum

composite procedure. The lack of properly defined standard procedures and collaborative

testing of prepared samples restricted its acceptance as a standard analytical technique.

Under Contract No. 68-02-3266, the Provisional Methodology is being evaluated relative

to (1) sample collection, (2) sample transport, (3) sample preparation procedures , (4) identification of particles as fibers, and (5) verification of asbestos materials. Specifically

some of the parameters requiring study were:

1) the recommendation of polycarbonate filters;

2) the acceptance of high volume filtration;

3) the effects of sampler face velocity;

183
4) the loss of fibers from handling and transport;

5) the role of fiber bundles, and

6) the role of energy dispersive x-ray analysis.

The information being presented are preliminary results of the present study.

2. Purpose and Use of the Methodology

We have found that one of the major reasons for the variability in results and con- sequently the acceptance of the methodology was the misunderstanding of the purpose as well

as the misuse of it. The usage of electron microscopy (EM) in asbestos analysis is

tedious, time-consuming, and expensive. The determination of asbestos in air, water, food,

and miscellaneous bulk materials would require a tremendous number of analyses. No one method would provide all the answers such as number concentrations, mass concentration,

length/width size distribution, asbestos or non-asbestos, kind of asbestos, etc.

The Provisional Methodology was designed to provide as many answers as possible, to be

relatively fast, and to be economical - essentially a screening tool to search, recognize,

and pinpoint the presence of asbestos fibers in the air. The method would provide the

number, an unaltered size distribution, a calculated mass derived from it, and information

whether it was chrysotile, amphibole, or not. The method is not for use to determine mass concentration in air or in a bulk sample nor to provide a classification of amphibole

asbestos species (crocidol ite, amosite, tremolite, anthophyl 1 ite , or actinolite), nor to go to court to cite a facility. The method, however, can be used for airborne asbestos

from source emissions, but only within the constraints or limitations of the methodology.

The use of selected area electron diffraction (SAED) from a visual analytical scheme is

recognized as a subjective method for rapid screening. Emphasis here is not on the possibility of misidentification but to err on the side of health and safety. A "false"

positive can lead to a recheck or repeat analysis whereas a "false" negative may lead to complacency.

Knowing its limitations, if the method is used for its intended purpose, a great deal

of uncertainty will be eliminated. However, for classification and to place the methodology

in its proper perspective, we have classified asbestos analysis into three levels of effort;

each level requires a greater degree of expertise, training, time and cost.

A Level I analysis would be the Provisional Methodology requiring an air sample, sample

preparation, and a TEM for obtaining a number count, size distribution, visual SAED analysis, and a derived mass concentration. A Level II analysis will be the Provisional Methodology plus energy dispersive x-ray fluorescence spectra analysis. A Level III analysis

would be a TEM analysis plus energy dispersive x-ray spectra analysis plus quantitative

SAED analysis. A Level III analysis will stress identification of asbestos species with

less stress on the number and size representation in the sample. The Level III analysis

will be used most probably with bulk samples and source samples and these will require

special precautions relative to sampling, contamination, counting, etc. In reality,

polarized light microscopy and x-ray diffraction analysis will probably be used prior to electron microscopy. A combination of levels is possible for special situations.

Placed in its proper perspective, the following discussion is with the Provisional

Methodology and its goal of providing reproducible number, length, width, kind of mineral

(chrysotile, amphibole, or not), and a calculated mass. The emphasis is to eliminate or minimize all parameters that effect reproducibility or the usage of a technique that can be

performed by only a limited few. As a monitoring tool, the methodology must be available

to all laboratories.

3. Sample Col lection

8" x 10", 102 mm, and 47 mm samplers were evaluated. The 8" x 10" samplers were tested out in the field. The 102 mm and 47 mm samplers were tested in an 8' x 8' x 8' aerosol box. Field personnel have difficulty in handling 102 mm and the 8" x 10" filters.

184
Use of a filter cassette for the 8" x 10" minimizes one of the problems for the field

worker, but creates problems for the laboratory personnel. The basic problems are the

fragility of cellulose ester filters and the flexibility of the polycarbonate filters.

Reproducible handling of 8" x 10" and 102 mm filters was impossible in the field, especially

during inclement weather. We do not recommend the use of these larger filter sizes for

field sampling. For sample collection and transport the cellulose ester type filters

collected the aerosol uniformly and the filter could be transported to the lab with a mini- mum of loss. Polycarbonate filters also collected the aerosol uniformly. However,

retention of particulates, especially the larger sizes, were very poor. Based on x-ray

fluorescence data, approximately 40 percent of the mass was lost in transit. Based on these results, cellulose ester type filters would be selected for sampling. However, for

very small particulates, shipping the polycarbonate filters through the mail resulted in

negligible loss based on XRF data.

4. Effects of Sampling Velocities

The 102 mm and 47 mm filters were selected to study the effect of face velocity on the

collection efficiency of polycarbonate vs. cellulose ester type filters. The tests were conducted in the 8' x 8' x 8' aerosol box using chrysotile aerosol. The sampling heads were attached to a probe with the vacuum pumps outside the box. The uniformity of aerosol

and sampling head position were established prior to conducting face velocity tests. Collection efficiency was determined by x-ray fluorescence analysis of the filter surface.

Efforts were made to collect only the fines of the aerosol by delaying sampling for four

hours after generation. The sampling times were carefully selected to obtain a very light

loading of chrysotile on the filter - not to exceed a mono- layer. The emphasis was on the

filter's efficiency and not on the asbestos fiber filtration efficiency. Face velocities

of 3.0, 9.0, 21.2, 28.8, and 34.8 cm per second were tested. At each face velocity, both

types of filters were tested. Lastly, each type of filter substrate was tested at three

different face velocities. Preliminary results show that for these aerosols, capture

efficiency was independent of face velocity. No noticeable difference was found between

the polycarbonate and the cellulose ester. The 47 mm filters appeared to have a higher

collection efficiency than the 102 mm filters. This may be due to the mounting technique

used for the XRF. In the 102 mm filter a circular 47 mm diameter section is cut for

mounting in the XRF cassette, whereas the 47 mm filter required no cutting. For ease in

handling, costs, and filtration advantage, the 47 mm filter is recommended for aerosol

sampl i ng.

The use of a sampling system designed for the analysis of Total Suspended Particulates

(TSP) levels for use in asbestos analysis by the substitution of filter substrates is one

of the causes for the present variability in results. TSP analysis uses high volume

sampling for 24 hours using glass fiber filters. High volume is used to grab enough air

sample to obtain a weighable fraction, 24 hours to average fluctuations in particulate

loadings from day and night activities, and glass fibers to provide minimum pressure drop

with maximum collection of solids, liquids, and reaction products.

For asbestos analysis, a sampling system called "hydra" has been designed and is presently being constructed for testing. The system is a set of individual samplers

mounted in a multi-headed arrangement and utilizing different sampling face velocities for

a constant sampling period. The sampling period is adjusted to accomodate a mi m'mum of

three (3) 47 mm filters for particulate loadings (low, medium, high), a 47 mm filter for

weighing and XRF anslysis, a sampler position for a field blank, plus additional sampling

ports to accomodate a midget impinger as well as a cascade impactor.

Total suspended particulates range from a low of 10 ug/m3 (remote, non-urban) to 60 ug/m3 for near urban to 220 ug/m3 for urban areas. However, for heavily polluted areas, TSP levels up to 2000 ug/m3 may be possible. From the electron microscopist' s viewpoint,

a loading of 5-10 ug/cm2 of filter is adequate for analysis. Values beyond 20-25 ug/cm2 will require a dilution treatment. For a 47 mm filter at a face velocity of 3 cm/sec

(2.48 £/min), 9.0 cm/sec (7.45 £/min), and 21.2 cm/sec (17.62 £/min) in 30 minutes, 74.4,

223.5, and 528.6 liters of air, respectively are sampled. For an urban area of 200 ug/m3

, 14.88 ug, 44.7 ug, and 105.7 ug would be collected on the 47 mm filter (eff. filtration

area of 13.85 cm2 ), or 1.07 ug/cm2

, 3.23 ug/cm2, and 7.63 ug/cm2

. The sampling time could

185
be increased to 60 minutes for a non-urban area or reduced in a heavily polluted area (source emissions).

The system advantages are:

1) It is less costly in time, effort, equipment, and materials.

2) The sample preparation steps are minimized and can be related to TSP levels.

3) The complementary analysis of field blank, TSP, bacteria, size distribution,

etc. , i s possible.

4) It can accomodate ambient air sample and source emission samples.

The system disadvantages are:

1) A short sampling period may catch an episode.

2) The sampling quantity on volume is small and may not indicate the presence

of asbestos fibers.

On completion of the assembly, the sampling system will be tested. The detection limit

for the 47 mm filter sampling 1 m

3 of air is 2 x 10 4 fibers/m3

.

5. Sample Preparation

5.1 Jaffe Wick Washer

Each user of the Jaffe Wick Washer makes slight modifications to the basic methodology.

Some of the modifications require greater dexterity, while others require greater attention. Figure 1 is a schematic of the modified Jaffe Wick Washer. The following changes

were made:

The petri dish cover and bottom were ground to obtain a closer fit and

minimize refilling the petri dish with added solvent. We find that the

substrate should be kept constantly wet for continuous wicking action to occur. Total time for the dissolution of the filter may take up to 40 hours.

The foam substrate or the use of a stack of 5 1/2 cm filter paper had small

problems. A combination of the foam plus a single sheet of 5 1/2 cm filter

paper provided an economical disposable substrate for the Jaffe Wick Washer.

The EM grid-polymer sandwich resting on the stainless steel mesh screen should

be picked up by the mesh screen while wet on completion of the polymer

solution and set on lens paper tacked to the bottom of a separate petri dish.

The grid is then lifted from the screen to dry adjacent to the screen.

The use of the micro-drop of solvent to tack the filter material to the EM grid

resulted in a high percentage of curled and ruined EM grid-filter sandwiches.

We recommend holding the sandwich at the edges with the tweezers and laying it

directly on a damp 100 mesh screen.

5. 2 Filter Selection

In the selection of a filter for transfer, the polycarbonate filters have been accepted

as the substrate providing minimum loss during transfer to an electron microscope grid

using the modified Jaffe Wick Washer. There is a great loss of particulates in transferring from cellulose ester filters using the modified Jaffe Wick Washer. The loss is

reduced significantly if the cellulose ester is first carbon-coated as with the polycarbonate type. However, the analyst has a difficult time seeing fibers in the irregular replica

surface especially of the small fibers. Reproducibility of the results depends greatly on the individual EM operator. The poor transfer technique capability of cellulose ester

filters appears due to the thickness of the filter, which is 150 urn vs. 10 um for Nuclepore.

186
01

tu

187
At present, we are investigating the fused membrane technique utilized by NI05H. Criticism

of the NIOSH technique is the possible loss of small fibers in the fused layer.

Presently, we have no well-defined method of utilizing cellulose ester type filters.

Ashing as practiced successfully in some labs does not give the same consistency when done

by others. The methodology must be refined for others to duplicate the same success reported by Dr. Chatfield at this meeting. If the sampling system called "Hydra" is suc- cessful, we may not need to use cellulose ester type filters.

5. 3 Gold Coating

The prepared grid is gold-coated very thinly in order to establish an internal standard

for selected area diffraction analysis.

5.4 Fiber Identification

During participation in i nterl aboratory testing of fiber counting, both optical and

electron microscopy, we find that counting discrepancies are minimized if more explicit

counting rules are made. The decision to count or not to count can be simplified by

instructions supplemented by sketches. In the Provisional Methodology, the term "fiber" is

used for a particle with an aspect ratio of 3:1 or greater with substantially parallel

sides. Using this definition, edge views of flakes, fragments from cleavage planes,

scrolls, etc. , may be counted as fibers. A consistency in counting is the desired goal.

Unfortunately, all fibers do not stand isolated to be counted. Fibers are with other

particulates in varying arrangements as well as to each other. We now characterize asbestos

structures wherein f i ber is one category. The others are bund! es (B), cl usters (CI), and

matrices (M), by themselves or in combination with each other. Bundles are fibers in a parallel arrangement with each fiber closer than one fiber diameter. Clusters are fibers in

a random arrangement such that all the fibers are intermixed and no single fiber is isolated

from the group. Matrix is a fiber or fibers with one end free and the other end is embedded

or hidden by a particulate. Combinations can be a matrix and cluster (M-Cl), bundle and a matrix (B-M) , etc.

Counting rules for single fibers are:

1) Particulates meeting definition of fiber - isolated by itself.

2) Count as single entities, if separation is equal or greater than 1 diameter.

3) Count as single entities, if 3 ends can be seen.

4) Count as single entities, if 4 ends can be distinguished.

5) Two or more fibers are counted as a bundle if the distances between fibers

are less than one diameter or the ends cannot be resolved.

6) Fibrils attached longitudinally to fibers are counted as part of the fiber

and size estimated (width) based on fiber to fibril relationship.

7) Fiber partially hidden by grid wires is counted if it is on the top or right

edge and not counted if on the left or bottom edge. Here the choice is to be consistent and to use two of the four sides of the grid opening.

At this meeting, counting rules may be adopted that may reflect the concensus of the

group. We are open to suggestions, corrections, and criticisms.

6. Conclusion

This paper is not the complete changes, modifications, or deletions in the EPA Provisional Methodology. We are still working on problem areas in order to obtain more consistent results in asbestos analysis. We appreciate any and all help that we can get.

A sample data sheet for the EM operator is shown in figure 2 for guidance. Asbestos

structures will have a very crude estimate of size.

188
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189
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS, Gai thersburg , Maryland, October 1-3, 1980. (Issued ^arch

1982)

FIBER IDENTIFICATION AND BLANK CONTAMINATION PROBLEMS IN THE EPA PROVISIONAL METHOD

FOR ASBESTOS ANALYSIS

Steven Ring and Robert J. Suchanek

Minnesota Department of Health

717 Delaware Street S.E. Minneapolis, Minnesota 55440

Abstract

Two problems with the EPA provisional method for analysis of

airborne asbestos by electron microscopy are identified. These problems

are 1) an inadequate discussion of blank contamination problems and 2)

fiber identification procedures that are not rigorous enough to prevent

some minerals from being misidentified as asbestos. In order to illustrate the blank problem, the additive nature of chrysotile fiber

contamination in the three stages of a cellulose acetate blank filter

preparation procedure is demonstrated. Increases in fiber lengths and

widths as well as numbers in successive preparation stages are shown to

be statistically significant. Considerable variability was found in

fiber numbers from filter to filter at each preparation stage.

Metropolitan air sample chrysotile concentration and size data are compared with blank concentration and size data. The potential for

confusing amphibole diffraction patterns with other minerals (including

palygorskite, sepiolite, enstatite, hypersthene, magnetite, laumontite

and mi nnesotaite) is discussed. A more rigorous procedure for

identifying asbestos minerals, incorporating both energy dispersive x- ray analysis and tilting the fibers to obtain zone axis diffraction

patterns is recommended.

The EPA's efforts to develop standard procedures for analysis of asbestos by electron

microscopy have vastly improved the quality of those analyses. They correctly emphasized

the development of standard techniques of sample preparation and analysis protocol. Until

this standardization occurred, analysis of the same sample could vary by a factor of 1000

from lab to lab. Under these circumstances, errors due to fiber identification or contamination problems were minor compared to errors resulting from preparation and counting

procedures. At the present time, however, i nterl aboratory agreement on the same sample has

progressed to the point where errors other than those involved with sample preparation are important. Despite the fact that several sample collection and preparation problems remain,

a standard method for analyzing asbestos in air should include a thorough discussion of

other sources of error in the technique.

In our use of the current EPA provisional method, we have found several deficiencies.

In this paper, we will discuss two of them and offer recommendations for improvement.

1. There is an inadequate discussion of asbestos contamination during sample preparation,

particularly with the ashing alterative. The method should require that effective

blank levels be reported with fiber concentrations and some means of determining

whether or not sample concentrations are significantly above blank level could also

be included.

190
2. The fiber classification procedures are not rigorous enough to prevent other materials

from being classified as asbestos. The method should provide for energy dispersive

x-ray spectroscopy (EDS) and indexing of electron diffraction patterns to prove the

presence of amphibole minerals. Then, if there are no significant interfering

minerals present, less rigorous technigues may be used.

1. Blank Contamination

Chrysotile fibers are often found as a contaminant on unused filters and ashing

dishes. The provisional method notes that contamination might be a problem but does not

discuss the possible variability in blank levels nor does it recommend that blank concentrations be reported. In order to illustrate these problems and show how important they are to the interpretation of some air samples, we will give the results of a study of the numbers

and sizes of chrysotile fibers in blanks and air samples.

The components of the sample preparation procedure were examined to determine the blank

levels of chrysotile. This attempt was complicated by the extreme variability in chrysotile

contamination levels found in the blank filters and in the ashing procedure used to eliminate the cellulose acetate filter material. The large standard deviation values for

the concentration of fibers in the Millipore and Nuclepore original filters and the ashed

dish Nuclepore filters listed in Table 1 illustrate this variability. Fiber concentrations

for a given type of filter often varied by a factor of 100 or more.

Table 1. Mean Fibers/mm2 and their Standard Deviations for Millipore Filters,

Nuclepore Filters and Ashed Empty Dishes.

Sample Composite

CHRYSOTILE

0. 1 urn pore size,

47 mm dia. Nuclepore

Ashed empty dish on

0.1 um, 47 mm Nuclepore

0.22 um pore size,

47 mm dia. Mi 1 1 ipore

0.8 um, 102 mm

dia. Millipore

No. Samples No. Fibers

26

31

6

20

219

731

205

641

Original Filter

x Fibers/mm2

97.76

466. 10

340.68

861.02

149.63

870.71

431.85

2224.90

Our blank preparation consisted of three main components. These were:

1. The cellulose acetate (Millipore) filter.

2. The ashing dish in which the Millipore filter was oxidized.

3. The polycarbonate (Nuclepore) filter onto which the resuspended

ashed Millipore was filtered.

Chrysotile contamination from these three components was additive and only the blank

Nuclepore filter could be examined apart from the other components. All of the filters and

the ashing dish were counted on Nuclepore filters.

The median (x) concentration (fibers per sguare millimeter) on the original filter and

standard error of the median (SE~) values for two types of ashed Millipore filter and the

Nuclepore filter were calculated, as well as the median concentration on the grid for the

ashed empty dish Nuclepore and the ashed 0.8 um pore size, 102 mm diameter Millipore.

Table 2 is a list of these values. Median values were used as an estimate of the fiber

levels on the filters since they were not drastically affected by the extreme values

encountered.

191
SE~ Fibers/mm2 x Fibers/mm2 SE~ Fibers/mm2

Table 2. x Fibers/mm2 and SE~ for the Blank Filters and Ashing Dish and x Fibers/Grid

Opening and SE~ for the Ashed Empty Dish the 102 mm Diameter Millipore, and

the 47 mm Nuclepore.

x Fibers/mm2

Sample Composite (On Original Filter) (On Original Filter) (On the Grid) (On the Grid)

47 mm Nuclepore 41.05 37.14 41.05 37.14

Ashed Dish 138.00 195.48 138.00 195.48

102 mm Millipore 164.50 621.88 324.00 234.95

47 mm Millipore 136.00 220.38

Using a z score test [l] 1

, the median concentrations of fibers on the grid for the

various filters and the ashed dish were compared, except for the two Millipores where the

concentrations on the original filter were used. Table 3 is a list of these z score values,

their corresponding p values, and the conclusions drawn from these values. The z score was

used to test the hypothesis (null hypothesis) that there was no difference in the fiber

concentrations on the ashed Millipore and blank Nuclepore filters and on the ashing dish

treated Nuclepore. It was found that the median fiber level values were significantly different for all of the preparations compared except for the two types of Millipore. Each

step in the preparation process apparently added fibers to the total count found on the

ashed Millipore. Both types of Millipore filter contained about the same concentrations

of fibers.

Table 3. Comparison of Median Chrysotile Concentration Values for the

Millipore and Nuclepore Filters and the Ashed Dish.

C omp a r i son

Ashed Dish/Nuclepore

102 mm Millipore/Ashed Dish

102 mm Millipore/47 mm Millipore

z Score

2.70

2.94

0. 17

P (One Tail)

0.00345

0.00170

0.43250

Conclusion (0.01 Level)

Reject H

Q

Reject H

Q

Fail to Reject H

In order to further assess the possible differences in the chrysotile contamination

encountered at various stages in the sample preparation procedure, the size distribution of

the fibers found at each stage were examined. Figure 1 is a set of graphs of the percentage cumulative distributions for fibers found in each of 12 length categories for the

Nuclepore, 0.8 urn, 102 mm diameter Millipore, 0.22 urn, 47 mm diameter Millipore and the

ashed empty dish composite samples. Figure 2 is a set of graphs of the cumulative percentage distributions for the 11 width categories. A comparison of these graphs indicated a trend toward increasing numbers of fibers in the larger size categories for both length and

width as the ashing dish and Millipore components were added to the Nuclepore. Using the

Kolmogorov-Smi rnov two sample test for determining significant differences in cumulative

percentage distributions [2], it was found that the apparent length and width differences

were significant. Table 4 is a summary of the statistical findings and a list of the mean

and median fiber widths and lengths. In comparing the width distributions of the ashed

empty dish and the 0.22 urn, 47 mm diameter Millipore, it was found that significant differences could not be demonstrated. The length distributions, however, were significantly

different. The comparisons of the 47 mm and 102 mm Millipores showed no significant length

or width differences. A one tailed Kolmogorov-Smi rnov two sample test was used in all cases since it was reasonable to hypothesize an increase in fiber lengths and widths from

one blank component to the next, based on the appearance of the cumulative percentage

di stributions.

figures in brackets refer to the literature references at the end of this paper.

192
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Table 4. Kolmogorov-Smi rnov Two Sample Test Results for Significant Differences for

Chrysotile Fiber Widths and Length Cumulative Percentage Distributions and

a List of Fiber Width and Length Central Values.

Compari son Dimensi on , 5 X Conclusion (0.01 Level)

Nuclepore/Ashing Dish Width 42. 60 Reject H

q Nuclepore/Ashing Dish Length 31

71 Reject H 47 mm Mi 1 1 i pore/Ashi ng Dish Width 0. 60 Fail to Reject H

q 47 mm Mi 1 1 i pore/Ashi ng Dish Length 18. 73 Reject H

q 102 mm Mi 1 1 i pore/Ashing Dish Width 13. 23 Reject H

0

If)? i w <_ mmmill Millinore/Ashinn ill i i i j u i \— f n j* i i i i i \j Dish u i j m 1L pV. nn1 IU t«... ht

1 57. 75 Reject H

q 47 mm Mi 1 1 i pore/0. 8 Millipore Width 4. 52 Fail to Reject H

q 47 mm Mi 1 1

i pore/0. 8 Millipore Length 7. 10 Fail to Reject H

q 47 mm Mi 1 1 ipore/C. C. Nuclepore Wi dth 29. 63 Reject H

Q 47 mm Mi 1 1 ipore/C. C. Nuclepore Length 60. 71 Reject H o 102 mm Mi 1 1 ipore/C. C. Nuclepore Wi dth 79. 89 Reject H

q 102 mm Mi 1 1 ipore/C. C. Nuclepore Length 17. 75 Reject H

q

Samples x Width (urn) x Width (pm) X Length (urn) x Length (pm)

Nuclepore 0.0350 0.033 0 335 0.280

Ashed Dish 0.0490 0.042 0 511 0. 370

47 mm Millipore 0.050 0.042 0 728 0.480

102 mm Millipore 0.053 0.042 0. 755 0.550

After determining the median blank chrysotile levels for ashed Millipore filters, an examination of ashed air samples collected on 0.8 pm pore size, 102 mm diameter Millipore

filters was undertaken. Seven ashed air sample chrysotile counts were obtained and mean, standard deviation, median, and standard error values were calculated. Table 5 is a summary

of these findings. The z score test [1] was done to test the hypothesis that there was no difference in the median fiber concentrations between the ashed air samples and the ashed

Millipore blanks. A significant difference was found. More chrysotile fibers were present

on this particular set of air samples than were found on the Millipore blanks. The z score results are listed in Table 6.

Cumulative percentage distributions for the widths and lengths of the total fibers on the air samples and the 102 diameter Millipore are presented in figures 3 and 4. The

apparent tendency toward wider and longer fibers in the air samples was tested using the

Kolmogorov-Smirnov test. The results, summarized in Table 7, show that significant differences in these size distributions do exist.

195
Table 5. Metropolitan Air Sample Data for One Collection Site.

2 Sample Fibers/Cu. Mete r Fibers/mm

1 9.92 x 10

3 844.96

2 1.59 x 104 1369.98

3 8.94 x 10

3 502.50

4 6.04 x 10

3 1850.00

5 1.51 x 10

4 1960.00

6 1.03 x 10

4 1270.00

7 1.09 x 104 4230.00

x Fibers/mm2 (Original Filter) = 1718.20

S- Fibers/mm2 = 1221.17

x Fibers/mm2 (Original Filter) = 1369.98

SE~ Fibers/mm2 = 576.95

Table 6. z Score Results for the Comparison of the Metropolitan Air Sample

and 0.8 pm, 102 mm Millipore Fibers/mm2 Median Values.

Compari son z Score P (One Tai 1 ) Conclusion (0.1 Level)

Air Sample/0.8 pm Millipore 4.66 0 Reject H (Fibers/mm2

)

Median Values

0.8 pm, 102 Millipore x Fibers/mm2 = 861.02

Air Sample x Fibers/mm2 = 1369.98

196
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197
198
Table 7. Kolmogorov-Smirnov Test Results from the Comparison of the 102 mm Diameter

Millipore and Metropolitan Air Sample Cumulative Percentage Distributions,

Along with the x and x Values of the Fiber Widths and Lengths.

Comparison Dimension

102 mm Mil 1 i pore/Air Width

102 mm Mi Hi pore/Air Length

8.78

21.94

Conclusion (0.01 Level)

Reject H

Reject H

Samples

102 mm Millipore

Metropolitan Air

x Width (urn) x Width (urn) x Length (urn) x Length (urn)

0.053

0.064

0.042

0.050

0.755

0.965

0.550

0.790

An estimate of the blank contribution to the chrysotile concentrations on the air

samples was undertaken. Using the median Nuclepore and 102 mm diameter Millipore fibers

per square millimeter values on the original filters (from Table 2) and the sample areas of

the filters, the median total chrysotile fiber number on each filter component of the blanks

and on the ashed air samples was calculated. Table 8 is a list of the total fiber values

and the calculated chrysotile percentage contributions of the Nuclepore (including ashed

dish) and Millipore filters to the air samples. Figure 5 is a graphic presentation of the

data in Table 8. The ashing dish contribution to the Nuclepore was about 70 percent of the

fibers on the Nuclepore.

Table 8. Values for the Percentage Contributions of the Millipore and

Ashed Dish Nuclepore Filters to the Metropolitan Air Sample.

Sample area of Nuclepore = 960 mm

2 Sample area of .8,102 mm diameter Millipore = 6650 mm Median Fibers in whole sample area of Nuclepore (+ Dish) = 132,480

Median Fibers in whole sample are of Millipore (including

a correction factor of 1.23 for the no sample margin) = 1,341,438

Median Fibers in whole sample area of air sample = 9,110,500

Hypothetical Percent of Percent of Fibers Percent of Fibers

Air Filter Ashed From Mi 1 1 ipore From Nuclepore + Dish

100 13. 2 1 4

90 13. 2 1 6 80 13. 2 1 8 70 13. 2 2 1 60 13. 1 2 4

50 13. 1 2 9 40 13. 0 3 5 30 12. 8 4 7 20 12. 5 6 9

10 11 7 12 8

199
SO 60 40 20

% ASHED

Figure 5. Percentage contributions of the Millipore and Nuclepore (with ashing dish)

blanks to the sample chrysotile levels in a metropolitan air sample.

The blank contribution of chrysotile fibers to the sample chrysotile counts can be

substantial, especially when small sections of a sample are ashed. As can be seen in

Table 8 and figure 5, the relative contribution of fibers by the Millipore filter remains

nearly constant. The Nucl epore-ashi ng dish contribution, on the other hand, becomes greater

as the size of the filter ashed decreases. The size of the Nuclepore filter is independent

of sample area ashed. The percentage values presented apply only to these samples. If the

sample load were less, the blank contribution to the chrysotile fiber count would be

proportionately greater.

200
Although additional statistical tests might be used to bolster the case, the fact that

the size distributions as well as the numbers of fibers on these samples differ substantially leads one to conclude that each step in this preparation procedure adds contamination

to the final analysis. In this study, we did not separate those blank samples which were

ashed alone in the ashing chamber from those which were ashed with samples. Thus, there is

the possibility that some cross contamination occurred within the ashing chamber on some of

these samples. Our purpose was to evaluate what was actually occurring during the preparation of air samples. This preparation consisted of ashing two or three air samples along

with an empty dish and Millipore blank in one ashing chamber. If contamination occurred,

it was a part of the samples as well as the blanks.

It is also evident, at least in this study, that large variability in the blank concen- trations makes comparison of a particular sample with the blank that was ashed with it an unsuitable technique for assessing contamination in the sample. The large variability will cause the contamination level to be frequently over- or underestimated. Instead, what is

needed is overall statistical control, where the blank level and its variability are

known. From this, one can determine when a particular preparation is out of control, and

whether a particular sample concentration is significant. We suggest that the provisional

method provide a thorough discussion of the contamination problem as well as sample preparation and counting errors so that when a sample is reported it will include confidence

intervals for the fiber concentrations and an indication of whether that concentration is

significantly above contamination levels.

2. Fiber Classification

In addition to the blank problem, another problem which may seriously affect the

quality of the analyses is the fiber classification procedure. Except for the problems

outlined by Millette [3] and Chatfield [4], the identification of chrysotile asbestos is

easily accomplished using standard diffraction patterns. Chrysotile is scroll-like in

structure and gives the same diffraction pattern regardless of how it is rotated about the

fiber axis. For that reason, there is no problem with orientation when using electron

diffraction to identify the mineral.

The situation for amphibole minerals is much different, and confusion in identification

can result for many reasons. The provisional method provides four criteria to classify a fiber as amphibole. These criteria are that it must have a diffraction pattern with closely

spaced spots arranged in rows, called layer lines, the spacing between the rows should

represent a distance of about 0.52 nm, the pattern should look like those obtained from

standards or published in the literature and its morphology should look like an amphibole.

Because amphiboles are not tubular in structure and are usually single crystals at the

sizes which produce patterns in a 100KV TEM, different diffraction patterns are obtained

with different orientations (rotations) about the fiber axis. Because of this, many

amphibole fibers give patterns which may be atypical of patterns obtained from standards or those published in the literature. Misidentification can be the result. Figure 6 provides

four amphibole patterns which are similar, but differ because of orientation.

201
Figure 6. Four "typical" amphibole patterns (grunerite). Closely spaced spots form

rows approximately 0.52 nm apart. Each pattern is different because

different planes diffract when the fiber orientation to the electron beam

is changed, a) near [301] zone axis, fiber has (100) plane almost perpendicular to electron beam b) near [312] zone axis fiber has (110)

plane almost perpendicular to beam c) arcs of bright spot are Laue zones

d) some streaking evident.

An additional problem to incorrectly classifying fibers which are in fact amphibole, is

the possible identification of other minerals as amphibole. There are several minerals

which give diffraction patterns which closely or superficially resemble those obtained from

amphibole. Palygorskite , sepiolite, enstatite, hypersthene, laumontite, magnetite, and

minnesotaite all give diffraction patterns which could be mistaken for amphiboles. This

problem is further exacerbated by the fact that it is difficult or impossible to estimate

distances in the darkness of an electron diffraction pattern on the TEM screen. Figure 7 shows six electron diffraction patterns which look like amphibole patterns. Plate a is an amphibole pattern. How easily can it be distinguished from the others? Thus, the provisional method is only adequate when the mineralogy of a sample is well known or when

minerals with amphibole-1 i ke patterns are not present.

Unfortunately, uncomplicated or well characterized sources are rare. Increasingly,

the method is being used to measure ambient air concentrations when there is no defined

source of asbestos. Broad mixtures of mineral fibers may be found in rural or urban air or associated with mineral assemblages in mining operations. A laboratory which relies on the

provisional method may find it extremely difficult to classify the fibers accurately. Mistakes are often very expensive and time consuming to correct and can lead to the loss of a laboratory's reputation.

202
Figure 7. Can the amphibole in this group be identified using the provisional

method? a) grunerite (amphibole) b) hypersthene c) minnesotaite

d) sepiolite e) palygorskite f) laumontite.

There are several steps which could be taken which greatly increase the power of the

method to separate the amphiboles from other minerals. As a first step, some means of

accurately measuring the spacing between the layer lines in diffraction patterns should be

implemented. This could be done by evaporating a thin gold film onto the grid to provide

an internal diffraction standard or by requiring that all supposed amphibole diffraction

203
patterns be photographed and the camera constant recorded and used to determine the actual

spacing between layer lines. A gold film of proper thickness will not obscure detail in

the image while providing the ring pattern which can be used as a ruler in the microscope.

Although there is some variability in the spacing between layer lines in amphiboles, some interfering minerals could quickly be eliminated by this technique.

The use of energy dispersive spectroscopy (EDS) in conjunction with the measurement of

the layer line spacing would greatly improve the capability of the analyst to distinguish

amphiboles from other interfering minerals. Several laboratories are already using EDS on transmission or scanning microscopes. Making its use part of the standard method would

greatly enhance asbestos identification capabilities. Unfortunately, there are still

several minerals which might be incorrectly identified as amphibole based on EDS and layer

line spacing. Figure 8 shows four EDS spectra corresponding to plates a, b, c, and d in

figure 7. If the composition variability of amphibole minerals is considered (solid solution series), it is clear that it would be difficult distinguishing these fibers by EDS.

n» 3IS 38SEC 1442? INT n* 36$ 3ISEC 13172 INT M-5J2 H-1MEU 1 10 A««18KEU 18 I 0=512 H-I0KEU 1 I Q A0M8KEU 10

c d

Figure 8. Energy dispersive x-ray spectroscopy (EDS) spectra from

a, b, c, and d of figure 7.

In order to thoroughly differentiate these minerals from amphiboles, it is necessary

to go one step further. If the fibers are tilted in the electron microscope so that the

electron beam is parallel to a set of planes in the crystal (on a zone axis), the resulting

diffraction pattern can be indexed. That is, the planes which produce each spot in the

diffraction pattern can be identified. Patterns can be indexed as illustrated in figure 9 using the computer [5], using common zone axis electron diffraction patterns [6], or by

hand with tables. Used in conjunction with EDS or along with more than one pattern, this

will provide positive identification of the fibers and evidence which can be verified by someone else.

204
3*[l10]

Figure 9. Zone axis patterns. Bright ring is from gold film (111) plane and

represents a d value of 0.2355 nm. Gold ring is sufficient to exclude

a and b from amphibole category (minnesotaite and laumontite). c, d,

e, and f show four common amphibole zone axes. Large arrow gives the

orientation of the fiber to the pattern.

d OJ

205
It may be argued that tilting and indexing patterns is too time comsuming and not suitable for routine work and that it should only be used when needed. Unfortunately, one does not know when there are interfering minerals present unless rigorous techniques are used to identify them. It must first be determined that interfering minerals are not

present then less rigorous and less time consuming methods can be used.

3. Conclusions

The provisional method should provide an adequate method of addressing the contamination levels associated with these analyses. Our experience with the blank problem provides

evidence that contamination is found at each stage of the sample preparation procedure.

Although this analysis pertains mostly to the procedure which uses low temperature ashing,

there clearly is some contamination on the Nuclepore filters themselves. This might be

important in lightly loaded samples. We suggest that the effective blank levels be reported

along with fiber concentrations and confidence intervals for the fiber count. Ultimately,

some expression of the blank variability should be reported. Thus, as suggested in the

provisional method, the minimum detection limits might be the blank level plus three

standard deviations of that level.

Finally, the method should provide for the rigorous classification of the fibers using

zone axis diffraction patterns and EDS. It may not be necessary to index every pattern

obtained from every fiber in a sample, but some part of the standard method should provide

that when a sample with an unknown assemblage of minerals is brought into a lab some percentage of the fibers, and particularly those identified as amphibole, should be examined by

EDS and quantitative electron diffraction. The techniques should become a part of the capabilities of any lab analyzing asbestos by electron microscopy. If a laboratory can show

that no interfering minerals are present, then certainly less rigorous methods may be used.

References

[1] Hodges, J. L. ; Krech, D. ; Crutchfield, R. S. Statlab. New York: McGraw Hill; 1975.

p. 187.

[2] Siegel, S. Nonparametric Statistics for the Behavioral Sciences. New York: McGraw

Hill; 1956. pp. 127-136.

[3] Millette, J. R. ; Twyman, J. D. ; Hansen, E. C. ; Clark, P. J.; Pansing, M. F. Chrysotile,

Palygorskite , and Halloysite in Drinking Water. SEM, 0. Johari , ed. AMF O'Hare, 111. SEM, Inc. 1979 p. 579.

[4] Chatfield, E. C. Analytical Procedures and Standardization for Asbestos Fiber

Counting in Air, Water, and Solid Samples. This publication page 000.

[5] Lee, R. J. Electron Optical Identification of Particulates. Electron Microscopy and

X-Ray Applications. Ann Arbor, Mich.: Ann Arbor Science Publishers. 1978 p. 175-188.

[6] Ring, S. J. Identification of Amphibole Fibers, Including Asbestos Using Common Electron Diffraction Patterns. Ann Arbor Science Publisher: in press.

206
National Bureau of Standards Special Publication 619. Proceedings of the NBS/EPA Asbestos

Standards Workshop held at NBS , Gai thersburg , Maryland, October 1-3, 1980. (Issued M arch

1982)

MSHA STANDARD METHOD FOR FIBER I DENT I FCATION B ELECTRON MICROSCOPY

R. L. Clark

U.S. Department of Labor

Mine Safety and Health Administration

Denver Technical Support Center

P. 0. Box 25367, DFC

Denver, Colorado 80225

ABSTRACT

The EPA Provisional Methodology Manual represents an eclectic

compendium of techniques employed in the analysis of airborne

particulates for asbestos by electron microscopy. Philosophically, the

methods tested and the resulting conclusions have a certain degree of

universal applicability. In practice, however, absolute adherence to the method may not be possible due to differences in mandated standards,

methods, and techniques.

The Mine Safety and Health Administration has developed a standard

method for the analysis of airborne particulates which roughly parallels

the EPA method. Variations in the technique include optical microscopy

for fiber count, transmission electron microscopy (TEM) for

photomicroscopy and selected area electron diffraction (SAED), scanning

transmission electron microscopy (STEM) for image analysis, and energy

dispersive x-ray spectrometry (EDS) for elemental composition.

Applications of the technique as a routine analytical method will be

discussed, with particular emphasis on areas of variation from the EPA

method.

Key Words: Electron microscope; energy dispersive x-ray spectrometry; image analysis; scanning transmission; selected

area electron diffraction; transmission electron

microscope.

1. Introduction

From the outset, it may be advantageous to state basic agreement with what I perceive

to be the pervading philosophy contained in the EPA Provisional Methodology. That is, the

application of minimal sample manipulation which yields a viable analysis reflective of

actual environmental conditions.

However, each phase in the construction of the Provisional Methodology was predicated

upon absolute latitude in the selection process, such that the entire analytical procedure

has an optimized quality. This degree of freedom in the selection process is not always

available, especially at the analytical level, where specific definitions and mandated

techniques demand a different approach to the solution of similar problems.

What will be described in this paper is an independently developed method which generally parallels the Provisional Methodology, but evolved in response to the imposition of

somewhat different constraints.

207
2. Overview of the MSHA Method

The methods employed by the Mine Safety and Health Administration in the analysis of

airborne particulate samples for asbestos is governed by specifications found in the Code

of Federal Regulations [I] 1

. Therein, the term "fiber" is defined as any particulate with

a three to one or greater length to width aspect ratio, and a length of five micrometers

or longer. The term "asbestos" is recognized as generic, applicable to a number of hydrated

silicates, but its use is specifically limited to describe the minerals chrysotile, amosite,

crocidolite, anthophylite asbestos, tremolite asbestos, and actinolite asbestos.

An optical microscopy method, generally known as phase contrast [2], is also specified

for use in obtaining gross fiber counts for each sample filter. It is assumed that the

entire fiberous content of the sample, as preceived by the optical microscopy method, has

the potential of classification as asbestos. Therefore, if the fiber count established by

optical microscopy exceeds the threshold limit value (TLV) for asbestos [3], the sample must

also be examined by electron microscopy for fiber identification.

Electron microscopy is utilized to obtain a physical characterization of the analyte

material, and to make an analytical determination of the nature of specific components of

the analyte. Physical characterization is made by a computer assisted image analysis

system. Analysis of the analyte is achieved by the classification of individual fibers

based upon the crystal lographic data contained in selected area electron diffraction (SAED)

patterns in combination with elemental compositional data derived from energy dispersive

x-ray spectrometry (EDS).

Results from the optical and electron microscopy methods are combined to yield the

number of asbestos fibers per unit of air. These data are used to establish if a health

problem exists in a specific mining operation, and if so, the nature and extent of that

problem.

3. Specimen Preparation

The first variation from the EPA Provisional Methodology, as well as a number of

subsequent variations, was necessitated by the use of the phase contrast optical microscopy

method. This method specified the use of cellulose acetate membrane filters for sample collection instead of the polycarbonate filters recommended in the Provisional Methodology.

In turn, the use of cellulose acetate filters precludes an initial carbon deposition, since

replication of surface features would obscure the analyte. However, this same quality of

surface roughness acts as an effective entrainment media, thereby minimizing analyte losses

that would be incurred during subsequent transportation from remote sampling locations.

Specimens of each sample are prepared on 10 nm thick carbon substrates. The carbon

substrates are produced by floating a thin film of one percent parlodion (high purity cellulose nitrate) dissolved in amyl acetate on the surface of reagent grade water. Electron

microscope grids are laid on the parlodion film, and the film covered grids are removed to the vacuum evaporator for carbon deposition. The parlodion film acts only as a temporary

support for the deposition of carbon, and thereafter is removed by dissolution in acetone

vapor using a modified Jaffe washer [4].

Two sections of the filter are excised adjacent to, and on opposite side from the wedge

removed for optical microscopy. These are laid, filtrate side down, on individual carbon

substrate covered electron microscope grids contained in a Jaffe washer. After about

twenty-four hours exposure to acetone vapor, the cellulose acetate filter is completely

dissolved, and each specimen receives a second carbon coating which effectively sandwiches

the analyte between two layers of carbon.

All substrate and specimen preparation, exclusive of carbon deposition, is performed

in separate class 100 environmental chambers, both for the safety of personnel involved and

to prevent specimen contamination. Sheets of clean room quality polyurethane are used in

preference to fiberous filter papers in the Jaffe washer, and for all cleaning application

Figures in brackets refer to the literature references at the end of this paper.

208
within the environmental chambers. Blank specimens are prepared in conjunction with each

group of specimens to test the integrity of the system.

A number of grid materials and mesh sizes have been explored for applicability to analytical electron microscopy, carbon coated composites, beryllium, and gold in size ranges

from 75 to 400 mesh. The carbon composite grids proved unusable due to an inherent dimensional instability which caused them to curl during substrate preparation. Both beryllium

and gold grids have been successfully applied in the production and analysis of asbestos

mineral specimens. However, the openings of the beryllium grids are poorly defined; they

are difficult and expensive to obtain in large quantities, and their use poses some very

significant health hazards. For these reasons, gold grids appear to be the best selection

for large scale routine asbestos analysis.

With respect to grid mesh size, grids with large openings provide a significantly

higher percentage of open area, for analysis. An example can be found by contrasting a 100

mesh grid with a 400 mesh grid, where the 100 mesh grid will have nearly twice as much open

area. The increase in open area, however, means a loss in specimen support, with a resulting increase in specimen substrate failure. Also, each grid opening contains more than one field of view, and therefore care must be exercised that a small degree of overlap

is maintained in each field of view as a grid opening is examined. These disadvantages

notwithstanding, the increased efficiency realized by the use of large mesh size grids

makes their selection advisible.

4. Analytical Procedures

Specimens of the sample are systematically examined in a scanning transmission electron

microscope (STEM) at an acceleration voltage of 100 keV and a magnification of 3700 diameters. Each fiber encountered during the examination is photographed at 6000 diameters, and

an SAED pattern is obtained and photographically recorded at zero degree tilt angle.

The STEM unit is then switched to the scanning mode at 20 keV, and computer aided

image analysis, is utilized to obtain individual fiber measurements and statistical data

relevant to those measurements. The accumulated statistical data generated by the image

analysis system, and printed at the end of each field of view, is used to determine when

adequate data for the physical characterization of the analyte has been achieved.

A deadtime corrected EDS spectra is accumulated for 200 seconds from each fiber at a rate of 300 counts per second. A computer program compares this spectra to an internally

stored linear regression and quadratic plots of intensity versus concentration for 10 key

elements (Na, Mg, Al , Si, K, Ca, Ti , Cr, Mn, and Fe), and calculates elemental composition

for the unknown spectra. The internal plots utilized in this calculation were generated

by actual analysis of chrysotile and the five specified asbestiform amphibole minerals

obtained from at least two different sources, and represented by at least 10 different

spectra from each mineral species.

At the end of analysis all SAED patterns are intensely examined, and each fiber is classified as either chrysotile, asbestiform amphibole, not asbestos, or ambiguous. The

classification of an SAED pattern produced by chrysotile is relatively straight forward,

since these patterns are composed of uniquely streaked diffraction spots arranged in well

defined arrays known as layer lines, which approximately correspond to a 0.53 nm periodicity [5,6]. Other minerals can also exhibit a 0.53 nm layer line periodicity; however,

none of these display the prominent streaking so evident in the case of chrysotile.

Differentiation between asbestiform amphibole minerals and associated nonasbestos

minerals can also be made by careful and critical inspection of the SAED pattern. The

characteristic patterns produced by the asbestiform amphibole minerals will have prominent

layer lines, numerous reasonably well defined diffraction spots that are closely spaced on each layer line, a 0.53 nm layer line periodicity, and low angle diffraction spots. How- ever, individual members of the mineral group cannot be readily distinguished by visual

analysis of the SAED pattern.

Species identification of the asbestiform amphibole minerals is achieved by confirmation that the SAED pattern is typical of this group, and that the computed elemental

209
composition derived from the EDS spectra is within the published compositional range [7-11]

of one of these minerals for all 10 key elements. If either of these two identification

criterion of positive SAED pattern classification as asbestiform amphibole and a fit of the

EDS spectra to the published compositional data are not achieved, then the fiber cannot be

identified as asbestos.

Fibers which produce SAED patterns that cannot be positively classified as chrysotile

or asbestiform amphibole, are classified as not asbestos. Typically, such an SAED pattern

would fail to have a definite layer line morphology (i.e., quasi-hexagonal array), a layer

line periodicity not equal to 0.53 nm, an inadequate number of diffraction spots, or a combination of the above conditions. Fibers which do not yield SAED patterns, or which

produce patterns with insufficient chrystal 1 ographi c data are classified as ambiguous.

5. Summary

The methods outlined in this paper are the result of both practical experience and

carefully designed experimentation. It is neither the intent nor the essence of this

presentation to advocate a system, method, or technique simply for the purpose of "marching

to the beat of a different drummer".

Instead, what has been described is a viable analytical method evolved in response to

a specific set of circumstances other than those imposed in the development of the EPA Provisional Methodology. Although this violates, so to speak, the letter of the law, the

spirit remains intact.

References

[I] Code of Federal Regulations, Title 30, Mineral Resources, Mandatory Air Quality

Standards, Sections 55.5-1 to 55.5-5; 1979.

[2] Joint AIHA-ACGIH Aerosol Hazards Evaluation Committee, Recommended Procedures for

Sampling and Counting Asbestos Fibers, American Industrial Hygiene Association

Journal: 83-90; 1975.

[3] Threshold Limit Values for Chemical Substances and Physical Agents in the Workroom

Environment, American Conference of Governmental Industrial Hygienist; 1979.

Pages 10 and 32.

[4] Frank, E. R. ; Spurny, K. R. ; Sheesley, D. C. ; Lodge, J. P. The Use of Nucl epore Fi 1 ters

in Light and Electron Microscopy of Aerosols. J. de Micros. , 9(6): 735-740; 1970.

[5] Clark, R. L. ; Ruud, C. 0. Transmission Electron Microscopy Standards for Asbestos.

Micron 5: 83-88; 1974.

[6] Ruud, C. 0.; Barrett, C. L. ; Russell, P. A.; Clark, R. L. Selected Area Electron Diffraction and Energy Dispersed X-ray Analysis for the Identification of Asbestos Fibers,

A Comparison. Micron 7: 115-132; 1976.

[7] Deer, W. A.; Howie, R. A.; Zussman, J. Rock Forming Minerals. New York: Wiley;

1963. Vol. 2, 203-373; Vol. 3, 170-190.

[8] Gaze, R. Physical and Molecular Structure of Asbestos. Annals of the New York Academy

of Science 132; 1965. 20-30.

[9] Hodgson, A. A. A Review of the Structural, Chemical, and Physical Properties of Naturally Occurring Asbestos Fibers. Royal Institute of Chemical Lectures Series 4; 1965.

[10] Huggins, C. W. Electron Micrographs of Some Unusual Inorganic Fibers. U.S. Bureau of

Mines, Pittsburgh, Pennsylvania; 1962.

[II] Speil, S. ; Leineweber, J. P. Asbestos Minerals in Modern Technology. Environmental

Research 2: 166-208; 1969.

210
LIST OF ATTENDEES

Michael E. Beard

U.S. E.P.A.

4004 Brewster Dr. Raleigh, NC 27606

Gene Brantly

Research Triangle Inst.

P.O. Box 12194

Res. Tri. Park, NC 27709

Gerald R. Chase

Johns-Manvil le

Ken-Caryl Road

Denver, CO 80217

E. J. Chatfield

Ontario Res. Foundation

Sheridan Park Res. Community

2395 Speakman Drive

Mississauga, Ontario L5K 1B3

Canada

K. S. Choppa

Union Carbide

4625 Royal

Niagara Falls, NY 14302

Robert L. Clark

U.S. Dept. of Labor

Mine Safety & Health Admin.

P.O. Box 25367

Denver, CO 80225

Philip M. Cook

U.S. Environmental Protection Agency

6201 Congdon Blvd.

Duluth, MN 55803

M. J. Dillon

Ontario Res. Foundation

Sheridan Park Res. Community

2395 Speakman Drive

Mississauga, Ontario L5K 1B3

Canada

Richard S. Feldman

U.S. EPA

116 Howard Dr. Apt. 3 Charlottsville, VA 22903

Mark B. Finn

Univ. of Illinois at the Medical Ctr.

P.O. Box 6998

Chicago, IL 60680

Joan J. Fitzpatrick

Denver Res. Inst. /Met. & Mat. Sci.

Denver University

Denver, CO 80210

Jean Graf

1 1 T Research Inst. 10W 35th St. Chicago, IL 60631

Walter John

California Dept. of Health Services

2151 Berkeley Way

Berkeley, CA 94704

Patricia Johnson

National Bureau of Standards

Washington, DC 20234

R. Keith Kirby

National Bureau of Standards

Bldg. 222, Rm. #B316

Washington, DC 20234

William H. Kirchhoff

National Bureau of Standards

Bldg. 220, Rm. #A261

Washington, DC 20234

Anthony J. Kolk, Jr. EMS Laboratories

12563 Crenshaw Blvd.

Hawthorne, CA 90250

Diann M. Kraft

DOL-MSHA-Metal and Nonmetal Mines

4015 Wilson Blvd.

Arlington, VA 22203

Richard J. Lee

U.S. Steel Corp., Research

125 Jamison Lane

Monroeville, PA 15146

D. E. Lentzen

Research Triangle Inst.

P.O. Box 12194

Res. Tri. Park, NC 27709

J. MacArthur Lonq

USEPA

College Station Rd. Athens, GA 30613

Carl W. Melton

Battel le-Col umbus Laboratories

505 King Ave. Columbus, OH 43201

John L. Miller

Environmental Protection Aaency

Res. Tri. Park, NC 27711

211
James R. Millette

USEPA-Health Effects Res. Lab.

26 West St. Clair

Cincinnati , OH 45268

Ed Peters

Arthur D. Little

15 Acron Park

Cambridge, MA 02140

Deborah G. Piper

Walter C. McCrone Associates, Inc. 2820 South Michigan Ave. Chicago, IL 60616

Harrison B. Rhodes

Union Carbide Corp.

4625 Royal Ave. , P.O. Box 579

Niagara Falls, NY 14302

P. Riis

Ontario Res. Foundation

Sheridan Park Res. Community

2395 Speakman Drive

Mississauga, Ontario L5K 1B3

Canada

Steven Ring

Minnesota Dept. of Health

717 Deleware St. SE

Minneapolis, MN 55440

Kim B. Shedd

U.S. Bureau of Mines

4900 LaSalle Road

Avondale, MD 20782

Patrick Sheridan

National Bureau of Standards

Washington, DC 20234

John Small

National Bureau of Standards

Washington, DC 20234

Eric Steel

National Bureau of Standards

Washington, DC 20234

Jan M. Steward

Walter C. McCrone Assoc., Inc 2820 South Michigan Ave. Chicago, IL 60616

Robert L. Virta

U.S. Bureau of Mines

4900 LaSalle Rd. Avondale, MD 20782

George Yamate

1 1 T Research Inst.

10 W. 35th St. Chicago, IL 60616

212
NBS-1 14A irev. 2-eci U.S. DEPT. OF COMM. 1. PUBLICATION OR 2. Performing Organ. Report No. 3. Publ ication Date BIBLIOGRAPHIC DATA REPORT NO. SHEET (See instructions) NBS SP 619 March 1982

4. TITLE AND SUBTITLE

ASBESTOS STANDARDS: Materials and Analytical Methods

Proceedings of the NBS/EPA Asbestos Standards Workshop held at the National

Bureau of Standards, Gaithersburg , MD, October 1-3, 1980

5.X#KK8®[KDSX Edited by:

John Small and Eric Steel

6. PERFORMING ORGANIZATION (If joint or other thon NBS, see instructions)

NATIONAL BUREAU OF STANDARDS

DEPARTMENT OF COMMERCE

WASHINGTON, D.C. 20234

7. Contract/Grant No.

8. Type of Report & Period Covered Final

9. SPONSORING ORGANIZATION NAME AND COMPLETE ADDRESS (Street. City, State, ZIP)

U.S. Environmental Protection Agency

Research Triangle Park, NC 27711

10. SUPPLEMENTARY NOTES

Library of Congress Catalog Card Number: 82-600506

J] Document describes a computer program; SF-185. FlPS Software Summary, is attached.

11. ABSTRACT (A 200-word or less factual summary of most significant information. If document includes a significant bibliography or literature survey, mention it here)

This publication contains the invited papers which were presented at a workshop on asbestos standards jointly sponsored by the Environmental Protection Agency and the

National Bureau of Standards and held on October 1-3, 1980. The workshop was divided

into five sections: (1) Bulk Materials for Preparation of Asbestos Standards .... This

section includes descriptions of natural and synthetic materials which have a potential use as standards for asbestos analysis. In addition, it also includes a description of the NBS Standard Reference Materials Program. (2) Standards Preparation....

The electron-microscopy preparation procedures for standards mimicking airborne and

waterborne asbestos samples are described. (3) Asbestos Analysis for Standards

Certification. ... This section describes analytical procedures and problems associated

with the intra- and interlaboratory analyses of asbestos standards. (4) Error

Analysis and Statistics .... This section describes the statistical considerations

which are involved in asbestos standards preparation and analysis. (5) EPA Provisional

Method. ... Current developments and selected problems with the EPA Provisional Method

for Electron Microscope Measurement of Airborne Asbestos Concentrations are discussed.

The papers include general reviews on each of the subjects as well as specific

papers detailing current research efforts.

12. KEY WORDS (Six to twelve entries; alphabetical order; capitalize only proper names; and separate key words by semicolons)

Asbestos standards; asbestos statistics; electron microscopy: fibrous minerals.

13. AVAILABILITY

| X| Unlimited

|

| For Official Distribution. Do Not Release to NTIS

FX] Order From Superintendent of Documents, U.S. Government Printing Office, Washington, D.C.

20402. H Order From National Technical Information Service (NTIS), Springfield, VA. 22161

14. NO. OF

PRINTED PAGES

220

15. Price

S7.00

USCOMM-DC 6043-P80

U.S. GOVERMENT PRINTING OFFICE 1982-0- 369-342/713



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