10 PLANNING

IZ REPORT

NBS and Industrial

Biotechnology:

Technical Developments and

Future Measurement Needs

Thomas C. O'Brien, Ph.D.

July 1982

Planning Office

National Bureau of Standards

NBSIR 82-2549

100

.U56

82-2549

1982

C.2

PLANNING

REPORT

tf(Arn>jiAJL auKM.'' Ca BTAKOAtUM

UJBK-oir

AUG

NBS and Industrial

Biotechnology:

Technical Developments and

Future Measurement Needs

Thomas C. O'Brien, Ph.D.

July 1982

Planning Office

National Bureau of Standards

NBSIR 82-2549

2 1982

Acknowledgments

I wish to thank the many people and organizations who have contributed

to this report, both in providing information, and in reviewing the

report and suggesting corrections, modifications and additions. The

author is especially appreciative of contributions to this report by

Peter Heydemann, William Kirchhoff, Elaine Bunten-Mines, Dennis Reeder,

and Kenneth Gordon.

The responsibi 1 i ty of any remaining errors or inaccuracies remains with

the author. The author would appreciate having report omissions or oversights called to his attention so that they can be considered for

future reports.

Executive Summary

Reports and articles have indicated that biotechnology will be one of the

most significant industrial technologies in the next several decades. The major

areas that will be impacted by this technology include chemicals, energy, health

care, food processing, and agriculture.

Beause of the potential long-term industrial importance of biotechnology,

the National Bureau of Standards began in the late 1970s a process to: (a:

examine NBS's role and responsibilities related to industrial development and use of this technology, and (b) identify infrastructure support needed by future

biotechnology based industries. Initial NBS Planning Office reports resulting

from this examination established a policy framework that illustrates the

opportunity for integrating traditional Bureau services and fundamental research

capabilities in chemistry, physics and engineering with long-range industry non- proprietary (generic support) needs in biotechnology and in the applications of

this technology. These reports also identified the commodity organic chemical

industry as an early adopter of this technology, as well as an industry which

would require NBS services.

0

This report describes those initial steps the Bureau could take to develop

fundamental research capabilities and service-related initiatives to meet future

industry needs for infrastructure support in biotechnology. This is accom- plished by: (a) examining commodity organic chemical industry trends, (b)

identifying research opportunities and barriers in biotechnology related to

commercial production of products, and (c) evaluating existing NBS capabilities

in relation to long-term industry needs.

iii
Report Conclusions

0 The commodity organic chemical industry -is important to the U.S. economy and

has maintained a favorable balance of tradOo However, this industry will;

(a) face increased international competition over tne next several decades

affecting its current trade position, (b) undergo a feedstock transition

(coal, biomass) sometime in the future, and (c) undergo structural change as

a result of overcapacity, pressure of economic forces, and technological

innovations. The timing of these events will be dictated primarily by

economic and political developments.

0 The commodity organic chemical industry views its current investments in

biotechnology as a means to keep options open on an important future technology. An industry output from this technology which will appear before the

end of this decade will be higher value added organic chemicals. Application

of biotechnology on an industrial scale for the production of "traditional"

commodity organic chemicals is not expected much before the end of the

century. Further, because of the difficulties that will be encountered in

displacing commodity organic chemicals now being produced from petroleum

feedstocks, the value added component which results from biotechnology may be

a comparatively small $1 billion. However, biotechnology may have a significantly greater market impact in the production of "non-trad i tional

commodity organic chemicals such as biopolymers.

0 Depending upon the economics and politics of producing organic chemicals via

biotechnology, biomass could become an important feedstock for the production

of organic chemicals. However, considerable research effort must take place

to improve the productivity and economics of the biotechnological processes

for biomass conversion that are under consideration. Critical to such an improvement for bioprocesses will be advances and innovations in biotechnology "tools", such as biocatalyst, recombinant DNA, cell culture, and

fermentation technologies. Further, if biotechnology is to be applied on the

industrial scale envisioned by organic chemical producers, innovation must

also occur in bioprocess engineering technology areas, such as process

monitoring and control, product separation and extraction, cell recycling,

and process intensification.

IV
0 This report summarizes discussions with industry representatives on their

perceptions of industry needs for infrastructure support in biotechnology as related to NBS's mission and capabilities. From these discussions, a number

of measurement needs as well as fundamental science and evaluated data needs

were identified. Concurrent with these discussions, an assessment of NBS

capabilities in the physical, chemical and engineering sciences was undertaken to determine how well existing Bureau capabilities could address

these long-term needs. In general, industry needs could be met by expanding

and/or refocusing existing Bureau scientific capabilities. However, new staff capabilities in enzymology, molecular biology/biophysics, and

microbial/cell biochemistry would be required in the next 2-5 years.

0 Should NBS decide, in addition to its current competency development effort,

to address long-term industry infrastructure support needs in biotechnology,

this report provides a framework for possible short-term and long-term

actions the Bureau could take in relation to its major activities which

include Standard Reference Materials, evaluated data bases, measurement and

analytic techniques, and non- proprietary (generic) research. In addition,

this report stresses the need for: (a) a research climate at the Bureau that

is receptive to the interdisciplinary cooperative and collaborative

requirements of this technology; and (b) periodic evaluation of biotechnology

in relation to NBS's mission and capabilities over the life cycle of this

technology.

V
^<1 »,i vr*

WJO-.P y. ,,,-,^1^

..Id

„ uji

' • ,«iiTM»^i "

.! rr’’; mg' :'<^" ! ,*ng»a».--yiSj«— ;y, , 'i. r • .-Mb'-'

c«'t^U£igrp- $d,

^

;v ^^ , , .

. .* 44'^ i -/ V t^eiT^’ •te:tei^

^'v .

. ' ' ":''iSwi®-' '. ' '' i-

' ~/‘ '^y.’j-':^ '.

‘^' - j-i" •'ai£b.

' *

li *.

,^„ .^. ,, ,

,

, ,.„,

;i.' 't(v r 'V 6^

l|^(^;

5

, ‘'.'V «|ili^pli <;: \i

)('.

Kr\.

BliMillW^'l£k> Vitt IMkjnnMI A «<r.» Uli .V >«^^jr'l(i..'4M^ w^T.lltkiiVjC^
Table of Contents

Page

Acknowledgments . i Executive Summary iii

Table of Contents vi

i

List of Tables ix

1. Report Objective 1

2. Introduction 3

.1 NBS and Biotechnology 3

.2 NBS Competency Development in Biotechnology 4

,3 Structure of the Report 5

3. Future Trends in the Commodity Organic Chemical Industry 7

.1 Industry Overview 7 .a Economic Importance 7 .b Definition of Industry 7 .c Growth Projections and Economic Influences 16

.d Capacity and Demand 18

.e Energy Requirements and Pressures 18

.2 Feedstock Alternatives 24

.3 International Competitive Trends 30

.a Trade Liberal ization 32

,b Import Pressures from Neighboring Countries 38

.c Middle East (Saudi Arabia) 39

.4 R&D, Technological Innovation and Industry Prof itabi 1 i ty 41

.5 Industry Strategies 44

4. Biomass Feedstocks for Organic Chemicals 51

.1 Introduction 51

.2 Supply 52

.3 Biomass Chemical Components 52

.4 Technology and Chemicals from Woody Plants 55

.5 Waste Residues 57

.5 Chemicals from Plants 59

.7 Barriers to Research and Commercial Progress 62

5. Conversion of Celiulosic Biomass to Ethanol 65

.1 Overview and General Process and Economic Considerations 65

.2 Pretreatment of Celiulosic Biomass 69

.3 Biological Conversion 71

.a Cel lulose Degradation 73

•b Hemicel lulose Conversion 74

.c Lignin Converson 75

.d Saccharification and Fermentation: Thermostability 75

.e Product Inhibition: Saccharification 77

.f Production Inhibition: Ethanol 77

.g Conversion of Pentose Sugars to Ethanol 78

.h Simultaneous Saccharification - Fermentation 80

.4 Technologies Important for Bio-Industrial Processes 82

.a Biocatalysts 82

.1 Immobilized Enzymes 85

.2 Industrial Applications of Immobilized Enzymes 88

.3 Immobilized Cells 90

.4 Research Opportunities 95

.5 Immobilized Cell Technology and Fermentation 96

V i i
.b Separation and Purification Technologies 98

.1 Membrane Technology 99

.2 Chromatogr aphy 104

.3 Supercritical Gas Extractions 105

.4 Monoclonal Antibodies 106

.5 Biologic Separation/Gerietic Engineering 106

.6 Dry Separations 107

.c Process Monitoring and Control 108

•d Genetic Engineering 112

6. Discussions with Industry and Technical Groups 115

.1 Purpose of Discussions and Listing of Meetings 115

.2 Summary of Industry Views on Needs in Biotechnology 115

.a Standards and Measurement Needs 118

.1 Raw Material Substrates/Feedstocks 118

.2 Biocatalysts/Enzymes 118

.3 Microorganisms and Cell Cultures 119

.4 Proteins and Nucleotides 120

.5 Biopolymers/Polysacchar ides 121

.6 Publ ic-Pr ivate Sector Discussion of and

Consensus on Standards 121

.b Fundamental Science and Information Needs 121

.1 Enzymes 121

.a Enzyiie Characterization 121

.b Enzyme Catalogue 122

.2 Fermentation 123

.3 Separation and Extraction 123

.4 Process Monitoring and Control 124

.5 Chemical Engineering 124

7. NBS Research Capabilities Related to Biotechnology 125

8. Development of NBS Knowledge Base and Capabilities Related

to Long-term Industry Needs in Biotechnology 131

.1 Predictive Models 132

.2 Microanalysis of Complex Mixtures in Bioprocesses 133

.3 Thermodynamics of Bioprocesses 134

.4 Chemical Kinetics of Enzyne Reactions 136

.5 Surface and Interface Characterization 137

.6 Separation and Analytic Techniques 139

.7 Separation Technologies—Membranes 140

.8 Standard Reference Materials (SRMs) 141

.9 Standard Reference Data (SRD) 143

9. NBS Role in Support of Industry Needs in Biotechnology 145

References 151

Glossary 167

viii
List of Tab les

Page

3.1

3.2

3.3

3.4

3.5

3.6

3.7

3.8

3.9

3.10

3.11

3.12

3.13

3.14

3.15

3.16

3.17

3.18

3.19

3.20

3.21

3.22

organic ChOTicals

^ Chemicals to Energy

, petrochemical) Producers

trading Percentage of Total Firm Sales

"®fo’chemirai“ Coma»dity Organic Chemicals

Example of Trade projections oraanic Chemical Firms

Commodity ty^os tions of Some Lead^O ° |

j^^rce of

Feedstock Security .js production Faciiicies

Location of Chemical Firms

.uernative Raw Material

Feedstocks ,„yolved in Selecting Alterna,ive

Some Consideratio

production of Organic

FeedstocKS of oppo'^Tunity foif* P

^°Chem?cris Biotechnology

iSe^rorerh^ -earch Organization 10 Tear

Japanese Investment in and Collaboration

^7?!li?L ' ,oemical Production Projections

SABIC Commodity O^gan

^ ^ Export Targets

-^--g Strategies of the Commodity

“''LrVanic al If^Slcal Industry Strengths, Weaknesses,

^indThfr"c^on?l

industry Competitive ForeExternal Influences

Forms of some^io'^^^^ *^®^^l^°*^a^inmass Comoonents

Use Possibilities

Global Harvest ainal Products . . rpll Cultures

3.23

3.24

4.1

4.2

4.3

4.4

4.5

4.6

4.7

4.8

8

9

11

12

13

14

15

17

19

20

22

23

26

31

33

35

36

39

39

41

44

45

46

47

53

53

54

56

58

60

60

63
Page

5.1 Products of Fermentation and Potential Organic Chemical Feedstocks 66

5.2 Areas for Development Leading to Process Intensification Gains 68

5.3 Production Cost Categories in the Bioconversion of

Molasses to Ethanol 69

5.4 Chemical and Physical Lignocel lulose Pretreatment Approaches 70

5.5 Approaches to Pretreatment of Cellulosic Materials to Enhance

Enzyme Susceptibility 72

5.6 R&D to Improve Thermostability and Reaction Rates 76

5.7 Example of Approaches to Reducing Ethanol/Product Inhibitions ~!^ 5.8 Approaches to Improved Conversion of Cg Sugars/Xylose to Ethanol 80

5.9 Technical Approaches to Simultaneous Saccharification and

Fermentation 81

5.10 Methods for Immobilization of Enzymes 86

5.11 Advantages and Disadvantages of Some Methods for Immobilizing

Enzymes 87

5.12 Application of Immobilized Enzymes in Large-Scale Industrial

Processes and in Pilot Plants 89

5.13 Parameters in Designing Biocatalyst System 91

5.14 Cell Immobilization Procedures 92

5.15 Methods for Immobilizing Whole Cells 94

5.16 Research Activities Related to Immobilized Cells 97

5.17 Experimental Efforts to Stabilize Whole Cells in Polymer Matrices

or on Surfaces 98

5.18 Technically Important Separation Processes 101

5.19 Approaches to Measurement of Biotechnological Processes 110

6.1 NBS- Industry and Technical Group Meetings on the Role of NBS

in Biotechnology 116

7.1 Identification of NBS Generic Research Capabilities Potentially

Related to Industry Biotechnology Needs 126

9.1 Areas of Scientific, Measurement, and Standards Opportunity for

NBS in Biotechnology 146

X
1. REPORT OBJECTIVE

This is the third report* in a series of National Bureau of Standards (NBS)

Planning Office reports on "NBS Contributions to Biotechnology." Its objective

is to provide a description of the climate within '/^ich NBS can evaluate and

begin to develop research initiatives and services to meet future industry

needs in biotechnology. This will be accomplisneo by selecting a candidate

industry--in this case, the organic chemical industry--and for that industry:

0 providing an overview of the industry, industry trends, and the technical

and non-technical forces influencing the industry's strategies;

0 identifying research trends, opportunities and barriers in biotechnology

related to the commercial production of products;

0 projecting industry long-range needs in biotechnology and their

relationship to NBS* mission, as well as tne future direction of NBS

research and services; and

0 summarizing NBS' in-house research and service capabilities, the

relationship of these capabilities to Ipng-range, non-proprietary,

generic industry infr astructure needs in biotechnology, and gaps in NBS

capabi 1 ities

.

This report will not link resource requirements to NBS long-range

initiatives. Rather, this report will serve as a guide for NBS scientists as they conceptualize, justify and develop long-range research and service

initiatives in biotechnology and for NBS managers as they evaluate research and

service priorities and plan for the allocation of limited Bureau resources.

*Bunten-Mines , E., Planning Report 1: NBS Contributions to Biotechnology. A

Prel iminary Report . National Bureau of Standards, Department of Commerce,

Apr VI- n, mo.

Coates, J. F., Planning Report 9: Implications of Biotechnology for the

National Bureau of Standards. A Seminar Presentation . National Bureau of

Standards, Department of Commerce, November 16, 1981.

1

2. INTRODUCTION

.1 NBS and Biotechnology

There are three basic roles of the National Bureau of Standards. (1) The

primary role of the Bureau is to provide a basis for measurements througnout the

United States by means of standards, evaluated data, and test methods. (2) NBS

is also to provide technical assistance to the Federal Government by carrying

out legislative assignments and by providing technical support to other Federal

agencies. This would be accomplished by drawing upon skills and knowledge that

arise from satisfying the first role. (3) The Bureau's final role is to solve

technological problems which are related to national goals as recognized by the

Government's leadership, and which are measurement and standards intensive and

so related to NBS's primary role.

With the dramatic increase in scientific, industry, and public interest in

biotechnology in the mid-late 1970s, NBS began a process to: (a) explore the

long-range technical and nontechnical research and commercialization issues

associated with this emerging and potentially significant industrial technology,

(b) examine the Bureau ' s role and responsibilities related to industrial

development and use of this technology, (c) identify infrastructure requirements

needed for support of future biotechnology based industries and bio- industr i al processes, and (d) identify NBS internal capabilities related to these needs

(1-3). The Bureau's 1981 long range plan summarized the results of this policy

analysis and projected that the commodity organic chemical industry, an industry

facing significant competitive and technological challenges in the future, would

be among the first to adopt biotechnological processes and to require NBS

supporting services (2). As a result of this planning process, a policy framework was established that illustrated the compatibility between traditional NBS

roles. Bureau capabilities in chemistry, physics and engineering, and some biotechnology industry needs for infrastructure support. However, policy

guidance from this process also suggested that there was less of a rationale for

justifying Bureau activities in areas of health care applications of biotechnology. The cummulative effect of this process and the subsequent policy

guidance was to reduce concerns many NBS scientists had about the acceptability

of initiating projects in biotechnology, and to call attention to incipient

industry needs in this area.

3
.2 NBS Competency Development in Biotechnology

The Bureau's Center for Chemical Physics (CCP) proposed in their 1981 longrange plan a competency development initiative on enzyme catalysis, a technology

essential for industrial biotechnological processes (4). In addition, the

Center for Analytical Chemistry (CAC), in their 1981 Annual Report, proposed

competency initiatives in immunoassay procedures, two-dimensional electrophoresis, and solid phase reagent development (5). Resources were subsequently

allocated by the NBS Director to CCP and CAC in Fiscal Year 1982 for support of

these in-house competency development initiatives. The purpose of this

competency development effort was to begin to develop an in-house knowledge base

in an area deemed pivotal to commercial developments in biotechnology. The

development of this knowledge base has already begun in the CCP and CAC. In addition, scientific and general information exchanges have begun at the Bureau

through the newly formed NBS Biotechnology Working Group. Concurrently, the

Bureau began a dialogue with industry, technical and trade associations, and

academia in order to determine what the future needs of biotechnology based

industries will be, and how these needs may relate to NBS measurement, information, and industry infrastructure service roles and responsibilities. This

dialogue with industry formally began in late 1981 and is discussed in more

detail in Section 6 of this report. It is possible that these discussions with

industry will lead to a core biotechnology research and service effort

integrated with complementary expertise already at NBS in disciplines related to

industrial applications of biotechnological processes. This report will provide

(1) a basis for evaluating alternative initiatives for a focus of a core effort

in biotechnology and (2) a framework for understanding some of the presently

disaggregated research elements at the Bureau potentially related to industry

needs in biotechnology.

The Bureau views its investments in biotechnology-related base capabilities

as part of a long-range strategy. This strategy will allow the Bureau to anticipate and to provide in a timely manner traditional infrastructure services that

will be responsive to biotechnology-based industry needs when in the latter part

of this decade industrial processes employing biotechnologies start to move from

developmental and pilot stages to commercial operations.

4
.3 Structure of the Report

The seven remaining sections of this report provide a basis for considering

generic technological problems and issues faced by producers of organic

chemicals in the implementation of biotechnological processes and how these

problems and issues may relate to Bureau research, knowledge development and

service activities.

Section 3 analyzes trends in the organic chemical industry , discusses

industrial technological challenges and competitive pressures, and discusses the

influence of biotechnology on industrial strategies.

Section 4 discusses the problems and opportunities associated with one

potentially important area for biotechnology applications, e.g., use of an alternative feedstock (biomass) in the production of organic chemicals.

Section 5 uses the research and economic challenges of converting

1 ignocel 1 ul ose substrate to ethanol to identify generic barriers and problems

that may be encountered in the scale-up of industrial biotechnological

processes. Biotechnological process technologies, such as fermentation and

enzyme catalysis are discussed. Also discussed are biochemical process

engineering technologies, such as separation and extraction, process monitoring

and control, and substrate pretreatment. Some comments are made on newer technological developments that may have potential applicability as biotechnology production technologies.

Section 6 provides a listing of industry contacts and information sources , and summarizes their perceptions of future biotechnology based industry needs

that may be related to NBS' mission.

Section 7 summarizes current NBS scientific and technological capabilities

that could complement a core biotechnology group at the Bureau. Projections are

also made on areas where Bureau capabilities could be strengthened.

5
Section 8 presents comments by NBS laboratory scientists on possible Bureau

roles and R & D opportunities related to biotechnology and on possible applications of Bureau expertise and services to industrial needs in biotechnology.

Section 9 discusses possible next steps the Bureau could initiate with

respect to biotechnology, industry interactions, and resource allocations.
3. FUTURE TRENDS IN THE COMMODITY ORGANIC CHEMICAL INDUSTRY

The purpose of this report section is several fold:

0 to provide an overview of the commodity organic chemical industry;

0 to discuss hydrocarbon feedstock transitions within the industry and

considerations associated with alternative feedstocks;

0 to illustrate the many internal and external pressures and forces

influencing the competitiveness of the industry; and

0 to provide a framework for considering these forces and for guiding

industry strategic planning.

.1 Industry Overview

.a Economic Importance . The U.S. chemical industry represents a significant and positive economic force domestically . It is the fourth largest

U.S. manufacturing industry. Organic chemicals represent about 44 percent of

the total value of chemical industry shipments (6), and are an important factor

in international trade (7), as shown in Table 3.1. This favorable situation was fueled in part by the significant price increases from 1976-1981 in large

volume, commodity organic chemicals (8). This situation could change dramatically, however, due to domestic and international economic climates, new competitors in world markets ( part icul arly commodity organic chemicals), availability and price of raw materials, world capacity/demand relationships, and

other factors.

.b Definition of Industry . Approximately one-third of the U.S. ctiemical

industry is devoted to producing organic chemicals or petrochemicals . Table 3.8

provides a listing of the primary (commodity) chemicals and a partial listing of

intermediate organic chemicals (SIC Codes 2865 and 2869). There are, in fact,

about 100 intermediate organic chemicals of major industrial importance, several

hundred more of lesser importance, and about 7000 different organic compounds in

7
Table 3.1

United States Trade Balance in Chemicals

U.S. Chemical Trade

($ billions)

Bal ance

1981 1980 1979 1978

exports 21.20 20.74 17.31 12.62

imports 9.60 8.58 7.49 6.43

bal ance 11.60 12.16 9.82 6.19

E/I* 2.21 2.42 2.31 1.96

B. U.S. Organic Chemical

(S bill ions)

Trade Balance

1981 1980 1979 1978

exports 5.87 5.70 5.06 3.37

imports 3.10 2.54 2.16 1.73

bal ance 2.77 3.16 2.90 1.64

E/I* 1.89 2.24 2.34 1.95

Source; Reference 7.

*E/I = Export/ Import ratio

8
Commodity

Organic

Chemicals

(Petrochemical

s)

and

Uses

®u

tn 3

<D +-

©

MW L_ c (0

<n -Q

u o 0 3 C U_ >• § 0 to u

•o t; oC

u

© c

LU < t3 in ‘S c

V.. 3 m -C

L. c U c s o 0 0 4- ©

in o s

(Ti c in c S CO o •— ©

— t o\ cu _

O)

© c — o 3 © — 1- c. c Li. ® c O) 4-

in

_ o (0 10 c m © a

in jc

©

to 5 L.

U -O

i. 0 § Q.

© 10 c

tX

X < X 0.

—

-1-coc

a)CL in

G) L. in »

(/) CO L_© U Iw

CO 3 © 3 *—

u TD +* ce o <0 s s u O

a) D. WM 6 o ©

4-© o

L. in 4- 4-

f- (0

C

C

0) >-

a. C/) CO ©

T3

V.o

Ul (D o —

©

(0 a) x>u -C *o© o H MM. o *— —

MW <0

— •—

©

F o o Q 0 L. c O T3 _ •I- (0

(D

®

©

® •—

X 0 e: c c c o C © Ou

L.

0)

®

®® o

+- •H >. >* >- >. >-

—

a)

C r; J= c 1. o

0. +- MW o >.

UJ LU LU > o

vlc

'4«®

in o

(0

c- y)

©©

©

®

® c ® c c > F -t- c © c © © a (0 © — © — —

(0 L.

— o E 3 > & > (0 _ 0 4-

©

L. L. 10 L. 3 u

a.

m LU 0. CD CD < aa 5

(/) (0

(J 153

L. *3

4“

O

U© in t

LU 4- L. T3

inQ.

C

(0 _s

C

10 8 ©

-9 CO © cc

in

©

© '^ CO o U © 3 L +- \in T3© c 4- <D c © 4- 0 — 12. © > © 3 (/) CO in MW U

4“

U 4- in

—

U

— U)

©C© L.

© L- :>•

w jC -Q O) <D © *o ©

LU < 1—Q < U.

in©u

inu in

©

© L. o N © © N E — *—

i- MW ©

0. t© t

Li.

©

•dS in Um

4w in 3 o m C — 0 MW © © © C 4- *3

t3 c

8,

©

-C o

lO *u o CL Vw •— L. ^ ' 4-

l. •3 4- 0 in 3 © MW -C ©

CO

©

X

X

*3 *3

l_ LO © *3 < s >.

© 0 u c c C WW © 0 < —

N X © C ©c o

WM

4-

© 0 © O. >. 1. c JZ © £ >• © 4- L. 4* 4= -C ©

LU CO

X 1— (m1

xed ) ©

©

©

©c©

m©

© in c c c © o C c © © © ©

—

MW

©

© c -W

— 4- o

4-N 3 © >. >* >• -C c C w. X X X © © 0 >• 1 1 1 ©

1. <

XJ hX 0 E Q.

X 4-©

ON o

SOURCE:

Reference
chemical production. This table (Table 3.2) also illustrates the products and

end uses of these organic chemicals. The dominant market for SIC Code 2865 and

2869 organic chemicals is polymers such as plastics, elastomers and fibers,

representing about 60-65 percent of total U.S. commodity organic chemical

consumption ( 9)

.

Table 3.3 provides general information on the hydrocarbon feedstocks

(petroleum, natural gas, and coal) for primary (commodity or bulk) organic

chemicals. These chemicals, such as ethylene, propylene, toluene, benzene, and

the xylenes, serve as the key building blocks of many industrial organic

chemicals. They are the foundation of the chemical industry, and rarely find

direct consumer use. Ethylene also serves as the base chemical for about 50

percent of the largest volume industrial chemicals.

Table 3.4 provides production data of commodity (primary) as well as major

intermediate organic chemicals, and general information on hydrocarbon feedstock

requirements, number of manufacturers, and major product outlets. Comparing

production information in this table with price information in Table 3.5

illustrates the sensitivity of primary (commodity) organic chemicals to the

energy price increases of 1978-1980 . These commodity chemicals are more sensitive to such energy price increases than intermediate organic chemicals or organic chemical -dependent products, since ethylene, toluene, propylene, etc.

are made from hydrocarbon resources or raw materials that require hydrocarbon

fuels ( 8 ). However, the problems associated with obtaining fuels and with

producing chemicals are not closely related (12). For the period 1976-1981,

primary organic chemicals had the greatest price increases relative to other

chemical s ( 8 )

.

Tables 3.6 and 3.7 list the major United States producers of organic

(commodity) chemicals. These producers listed in Table 3.6 account for 80

percent of U.S. total capacity; 74 percent of this production capacity belongs

to the petroleum companies, in part because of their process capabilities to

produce a stream of to 63 chemicals ( 6 , 9). Petroleum companies are also

consumers of about 40-45 percent of their organic chemical building materials

(9). Tables 3.6 and 3.7 also illustrate the variety of recent research and

10
Table 3.3

Hydrocarbon Feedstocks for Production of Commodity Organic Chemicals

Hydrocarbon Feedstock (1978 data)

Pr imary

•ganic (Commodity)

Chemical Source

Approximate % of

Chemical Product

methanol synthesis gas 100 %

olefins natural gas

liquids (ethane,

propane, butane)

43 %

petroleum liquids

(naphtha and gas

oil

)

57%

aromat ics petroleum liquids 97 %

coal based

(coal tars and

1 iqht oils)

3 %

Source: Reference 9.

11
(j 0 5 — -*• S u 'J s ^

3.

1 r

o

^

^11

i b 5

— O *» ^ is ^

> (j ^

U (O — 5.3>• 9 ^ M

c a (p s ® i

> s

i- s r ~

« 2 5 g,

i- 9 9 —

% i

« 5

e ~ zll

i

8

c

w

8 ^

>- e

•5 ?

6 S

01 5 p —

9 8» Z 0 0 c — 9

<<* -C —

« y >• c X

J ^ i

>. s

t; > 5 >> >• 9 ii ^

t s

O —

i. h z

tfl W99

•A • :^

05 9

9 * —

3 (A . 05

f 8

® “ - “ 2

"3

2. 2

9 U $

-c ^

c

9 0 ^

irt. n>* *- e u o — O (jB lA pa 9 £

O i ^

= - ? o >. —

o f 3

i

" O —

. . a

£ £

U i 5

* So lA y ^

9

>. • » . y P ^

. 9 9 . P — « Z ^

0 W ^ 9

y 9 3 e

>. ^ . u

8

0 9

9 ^

9 C 9 4- —

C 2 5

c

9 £ 0 9 • — . 05 S - a u. 9 e . 9 jT O — U —

9 9 >® y *9 e ® S' b

i £

^ <o o <S <N ^ iT> <13 — O

5 S

9 05

S 3

05 \j

Z 5

C .

5 o

35

(A .

22 s a y

e .

- O O — — 5 o

>» 05 ^ .

r *5 s i= u O

^ c 9 5

05 — C 9

y fl 9 — -

u O ^ 2 y c

1A.9.99A. PV.COCCPV. -C. 9 C 9 9 ^ —

p . o —

— o « 5

9 ^ 4. o 9 0 ^ - 3 3 "O »

OT O T3 >. • — 0 4= • A . • U 9 9 p Nw A a ”o c

9 I — P

r >- o 4-

. 9. 9 O

*9 A A

• 99 i

9 C C 9

9 e 99 *9 c 9 — — — ^09^

£ = a- “ ^ A 9 ^ — 2 9

8- £

£ £

® ® -5 9 ^ i y

<0 ^ ^

9 'O'C r»^«r

r> lA <r>

r» o — O

A ^ P-4 »A <0 ^

(M -O A CX A <M

<M r* A A ^ «r O ® A r*.

. A A O ^

^ O A >0 — O f"* CM A

PM —

CM P» ^ r** A A — ^ r* O S R

PM . <0 a r • r

— CO g

o o ^ r* 0* PM

P«* PM • P^ . 95 05

PM — . —

05 • 2 p

9

9 j p

c 9 9

9 — C

i- I

® - w O

9 a «•

*0 <0 'O A A ^ V

O 9

T5 .

i £ — — P > >. ^

^

9 y<43 e 9

9 — c

y >• 9

>. c —

f £

11

c c —

9 9 >» — — X

_ > >

U ^ £ t S « -

y —

. X P o % * p^ o

y O o

4* p a

9 JZ —

y “9

5 -seo

y —

o1 i

9 ^ ^ e s “

^ (/>

— o

3

y —

i 2

. £

. ;A A .

a L. 9 A

9 « y ^ . c 9 w p 9 9 y

U X c

A . w 9

9

y 9 c y ^ 9 e A

>0 U 9 A A 9 U A

9

9 9 A

y X 9 O

e w e 9 w 9 W 9 u

9 c e 9 . 90 -^ 9 U •• 9 X > ^ X ^ g W z a

o e ^ A

i 'S o i

J

. A a ^ .

P ^ C 0 u y p — A . 0 £ i3 A

9 -3 9 O ^ A ^ O

y 05 0 y

0 c . e a

u . 4- o 0 iz3 c 3 — r ^ s 3 f

1 ^ t

L. — S' 9 O - I

*“ o C

J2 H “

A .

0

o 9

9 (X y -

5 o

•f- ^ 9 3 ^

i

»

« . A

P 0

>» ^

8- 2

I. .

a I z

3 ^

9 3 — a

3 3

y

£

A —

3 9 9 y

e- s

^ 3 P o

1. y

y u 9 9 . ^ 05 . ^ j3’ 3

>• U

9 — 1? I5

» “9 i s

§• a

= 3 . "

lO I I - i-* . A

OCPUA 9

^ ^ i

>• Q

a ^ ii i

a z

3 z

£ ^

E i i

£ -O ® « U

i a 5

9 9

O O X

t £

>*-CggAa533 9 *- 3

* 0 — OO*/ P^5tOZ</5 X«/5 CJO« y*9 9^ 05£.

0 C L

03 S O
Table 3.5

Sensitivity of Commodity Organic Chemicals to Energy Price Increases

Average Price Range of Organic Chemicals

(cents)

Product ion

Energy

Commodity

Organic

Chemical Year: 1970 1974 1978 1979 1980

Requi remen

( Therm

1981 ton)-

Tol uene [if gal .) 22 58 61-70 99-135 115-140 123-145 556

Benzene (4/gal .) 24 90-105 70-85 85-165 145-190 160-200 608

Xylenes (4/ lb.) 3.3 6.7 6. 5-8.

4

8.4-18 18-23 19-24 714

Ethylene (4/lb.) 3.3 7.9 13.3-14 13.3-20 20-24.5 24-28 603

Propylene (4/lb.) 2.3 -- 9-9.3 9.8-16 17-20 18-22 584

Butadiene (4/lb.) 9.5 9.5-13 20-22 20.5-26 32-34 33-34 666

Source: Reference 10.

^Chemistry and Industry, May 1, 1982, p. 285.

13
Table

3.6

Leading

U.S.

Canmodi

ty

Organic

Chemical

(Petrochemical)

Producers

+- cn c i.

(D 0

E c

LU

to0

> 0

c L.3

«<- >.

CO *3 0 O

0 u

c Q1 <

u

<v

-1- to 0 —

•— <0 03 o

E0

o

to TD ^ i — Q.

(J

(0CL O

(0 O ^

c — O

IT3 ^

4- —

o —

O 'O c (0

(T3 U

CO —

J? E o o ^ 8 ~ i. -a +- Q m

E a. O m

(0

1- o

(0 —

e £ — ®

0. o

CTc

"uiui®

1.

0.

JZ

+- ® — i_

ro (0 0 o

ly) 0

c

0

>J X

X X X X

XX X XxXXX

X X X X X X X X X X X

X X X X X X X

O

VO O . • 0\ 03 c^l’-^00000^^0'— 0(NK3(N — CNCM — inCNCSIlf' O 1^ .s- lo Osl 0'L-i (N 01 O' l^l^l^r-iriin'3-K>rOK'CM(NCN(N*— —

X X

X X

X X

X X

X X

X X

XXXXX X X

XXXXXXXXX

XXXXXXXXXX

XXX

X X

X X X X

XXX X X X X

XXXXXXXXX

XXXXXXXXX

XXXXX

XXX

0

0 o £ 0 o u

o -o .C to •— 4- _ 4- to > 0 o •T3 ce JO 0 c. 0. C L. <0 X 0

"O (0 u 0 to o 0 0 CO C

•— o Q. 4- to (t3 4- “3 0 — <0 4- o C c 0 *3 C (0 C u

c T3 C C C — U 0 o c 0 o 0 3 — o c — ro O *4- (0 to t- 0 1. 0- *— 1. 0 X (0 C X c c JO 4- .C 0

.c X -H 4- c 3 .C o 3 © o o 0 E 3 O to £

CO UJ CO < 3 o 0- Q CO h- s o o Q s o < <

Source:

Reference
Table

3.7

Leading

U.

S.

Chemical

Producers

r: • o 1 Q. u

u

>- 0 ,2 L. 0 0 o in s 0 0

l_ ex 0 0

*o 0 • jji JO X o 5 y 0 0c 0X </) 2 0 —

•H .c <9 0 o ik to 0 c </> X TO o *o L. I \ » C ' O c 1. 0 to 0, E 0 U 0 0 J= > 05 0

4* c 0 • to JO o > •—

—

—

{fi j: C75 o u 0 uC .c 0 4-

O o t o C

— 0 4- <9

X o o 0 > o 0 UJ 4- X c X 0 c c >- h“ 0 0 c 0

4- » C CO c «k 0 to

O • 0 0 c 0 c 4- _ • >. 3 •4- 2 O O 05 0 0 JO i_

O'. 1. 05

C o o 0 0 O UJ C • CL % 0

“O JO OO c — .c O) 3

"o c 0 0 to to

O CO 0 0 C • 0 — o O) 0 0 0 3 ^ ©

.C

—

— o a

— 4- 0 4-

U

— o 0 0 Q 0 0 — 0-0 0 0 o o o CD >- o X o < s O

to to to to to (0 to to c E 0 0 0 0 0 0 0 0 0 0 ..p. 0 0 0 0 0 JT 0 0 0

>- 4- 0 u 0 E E E p £ E ff E

4- 0 0 0 E 0 0 0 E "> E E 0 o e 0

to MB .c pC 3 .c -C .c 3 3 3 .c 3 3 3 0 0 0 0 0 0 0 — fj © 0 0 0 0 0

*o 0 “C to 0 0 0 0 0 0 0 0 0 o O

O 0

to0 to to t. 4- to to to

u

4- *05

J— to l.

to to 0 0 0 0 0 0 0 5. 0> 0 0 0 0 0 0 a X X CL X X X X CO a. X X X X X X

fj) 0 to o o vO o o ON ON

X o m in in 0 0 o •— ir\ K*N iTn tn o X o m in ON — X — CN 04 O O CN O Ln CM o C7N E ON •k 0 to — O to m in rO N-> CV4 CN CM CM •—

vO

OO o _ •o > 0 *..» C 0 o 0 — u _

CL£ E0 <5 o

4- s o o c .c c 0 o o O C c 0 c — X O 0 to 0 3 X c 0 3 0 X c o 0 JC

ClQ UJ

X 2 o X 0 « -o — TD -o 0 C 0 — L — E k*» 0 0 JT c X « 0 0 0 CL - — >. 0 U o X O 0 0 o u — 4- o -D 0 0 c O C U 0 *— 0 • O 0

-o 8 0•o 3

4- 0

ce H-

— c 0 0 t.

— 0 — 0 U 0 • 3 3

CO 8 <

4-X

4-< J o

00

ON

CVJ ro

m vo 00 ON

O

m <0-0 0

Rank

based

on

1980

sales.

Source:

Reference

17.

Sources:

References

13,

16,

and

18
other investments by these companies in biotechnology. Table 3.8 illustrates the

diversity of major firms in the chemical industry and is a relative indicator of

their commitment to this industry.

The following characteri st ics can be generally applied at present to the U.S.

commodity organic chemical industry;

0 capital, energy, feedstock, and R&D/technology intensive;

0 established distributing and servicing networks;

0 global operations;

0 high plant and equipment investment in contrast to low labor content in cost

of manufacture;

0 highly skilled work force;

0 high growth rate in past dependent on commodity chemicals (future growth

potential may depend on specialty chemicals);

0 integrated plant complexes;

0 world scale capacity plants for commodity chemical production (current

capacity, however, exceeds global demands);

0 chemical plant sites near sources of raw material (raw materials represent

over 50 percent of manufacturing costs and industry outputs are highly

dependent on their supply, availability, and price);

0 world's largest producer of organic commodity chemicals;

0 competitive position based on low price, ready availability, and high purity

of commodity chemicals;

0 second only to West Germany in exports; and

0 signs of industry maturation and structural change on horizon.

.c Growth Projections and Economic Influences . The growth rate of primary

(commodity) organic chemicals has been declining for the past three decades:

1950S--17 percent, 1960s--13 percent, and 1970s--4.6 percent (20). The growth

projection for the 1980s is 4 percent, only slightly above the projected growth

of the GNP (2.8 percent). But probably more revealing is the declining ability

of the industry over the same time periods to rapidly penetrate end markets or create new end markets (20). This is an indicator of a maturing industry.

16
Table 3.8

Leading Chemical Industry Finns' Percentage of Total Firm Sales to Chemicals

Ccmpany

Chemica

1

% of

Total Sales Company

Chemica

1

% of Total Sales

DuPont 82 Esmark 6 Union Carbide 57 Gul f Oi

1

3 Monsanto 80 3M 9 Dow Chemica

1

69 Pennwa 1

t

45 Exxon 6 Airco 39

Celanese 76 Pf i zer 17

Al lied Ch©n i ca

1

66 Tenneco 5 W.R. Grace 36 Dow Bad i sche 100

Hercules 83 Kerr -McGee 25

Occidental Petroleum 28 Na 1 CO Chemica 1 89

Easi'man Kodak 20 American Hoechst 65

American Cyan am id 50 El Pa so Natural Gas 15 Shel ! Oi 1 15 Baychem 63

FMC Corp. 39 Un ion Oi 1 (Cal .) 7 Phil lips Petrol eum 19 Lh iroyal 8 PPG Industries 33 Cabot 56

Stauffer Chemical 85 GAF Corp. 20

Rohm and Haas 70 Atlant ic Richfi el d 4 NL Industries 41 Borg-Warner 1 1 Standard Oi 1 ( Ind.) 7 Chemetr on 47 Akzona 71 General Electric 1 Texaco 4 National Starch 79

Diamond Shamrock 60 Wi tco Chemical 47

Ethy 1 55 Freeport Minerals 88

Cities Service 18 IFF 97

01 i n 31 CF Industries 71 Standard Oil (Cal .

)

5 Morton-Norwi ch 30

Goodyear 7 National Di sti 1 lers 10

B. F. Goodrich 18 Northern Natiral Gas 19 Mobi 1 Oi 1 3 Willi ams Ccmpan i es 21 Air Products 74 Arco Polymers 92

F i restone 9 Farmland Industries 13 BASF Wyandotte 71 Eli Lilly 14 Merck 23 Procter & Gamb le 2 Lhited States Steel 4 Dart Industries 12 Asniand Oi 1 12 Dow Corn i ng 78

ICI America 86 Kewanee Oi 1 66

Lubr i zol 99 Texasgul f 34

Continental Oil 6 Houston Natural Gas 33

Reich ho Id Chemicals 96 Ccmmercial Solvents 90

CIBA-GEIGY 41 Emery Industries 90

IMC 37 Sun Oi 1 5 Borden 9 Ka i ser Al umi num 1 4

Source: Reference 19.

17
Part of the problem in the poor growth prognosis for the U.S. commodity

chemicals industry lies with the economic viability, vitality, and confidence of

its major customers--automobi les , housing, textiles, and appliances. High

interest rates, availability of capital, domestic and foreign economic

downturns, increased production capacity, value of the U.S. dollar, active

foreign government involvement in commodity organic chemicals, etc. will affect

the rehabilitation of the chemical industry customers and, in turn, future

growth prospects for the industry (8, 21, 22).

.d Capacity and Demand . Current capacity and future demand projections for

U.S. organic commodity chemicals (based upon economic forecasts, projected

chemical customer needs, new plants coming on-stream, new competitors, etc.)

indicate that capacity will exceed demand by about 20-25 percent (10, 23, 2^).

In spite of this, U.S. market growth for organic chemicals is orojected to be

1-2 percent over that for other industrialized countries (25). This will create

additional competitive pressures on the U.S. commodity chemical industry in

domestic markets, as well as on our chemical trade balance overall, and the

trade balance for organic chemicals in particular (Tables 3.1, 3.9, 3.10).

.e Energy Requirements and Pressures . In spite of extensive chemical

industry energy efficiency and conservation efforts in the 1970s, the chemical

industry, and in particular the producers of commodity organic chemicals, ^s one

of the most energy intensive and sensitive manufacturing industries . About 5.7

percent of U.S. petroleum and natural gas liquids are used by the chemical

industry as hydrocarbon feedstocks for the production of primary organic

commodity chemicals (ethylene, propylene, benzene, etc.), and an additional 1 percent is used as fuel in the manufacturing processes (23). In fact, chemical

industry natural gas consumption represents about 10 percent of total U.S.

consumption, e.g., 3 percent for feedstock and 7 percent for fuel (27).

A guiding principle of the commodity organic chemical industry has been

conversion of low cost raw materi al s , such as petroleum and natural gas, into

higher value added products. Geographic proximity of industry plants to such raw materials have been considered and will continue to be important in the

strategic planning of firms in this industry (24).

18
Example

of

Trade

Balance

Changes

In

Conmodity

Organic

Chemicals 03

(0 > 0

O)

g o KN 'O

—

CO

C o 00 S

K*^ IN 5 4= o

ON + IN+ 1

*- in

m o o o CO • • • 0 • ON 'h vD VO O (O' vO m 0

hOCN

GO CM o

G.E c

(03

cH

bO. b«l. o o o (N in. CM ON c 0 GO T

CO

(0 c O' + 4- 04 1 £ +- o +

CO CO vO o ON

GO 0 • • • • • • ON 4- (N GO CO CN 0 m CM 'O 1—

in -HUO

ClX

LU 030>C03oi 0 bQ. O) g o C7N c 0 00 • b^ b^ bQ. ON 00 •— O O + + !

CO ON in CO

GO 0 • • • • 2

to ON 0 in o> VO 1- to 0 b^ b^ bO. b^

U) E o CM to r*. in c 0 GO o 1 0 t. ON CN+ o

to

m s 0 • • • • O' 4- in rj* 00 0 rO to lO

+-

to \0

IT'

inm ^

fO« m

to

CSi

in

in

cn

<ncO 0 E u•— C • £ •— • 0 0 o 3 W o 0 • 0 0 > • • — o T3 C — 0 • >*

fl. 0 0 U)

U

O) O) O)

— cn o u >• w o +- 0 0 0 0 c 0 c u

4- c c 0 c 0 C 0 $ — 0 0 N 3 >. u >. >• 3 c X >• c

O* 0 0 1 4- 4" 4- a. (/> 0 >

vO<N0

(JC0u0H-0

cr0o

L-3o

CO
Ccmmodity

Or

ic

Chemical

Projections o

0^ ^ VO vO I m Hi

vb vo CO I PS HI CN4

HI vO

CM

CO o CSI I ON ^ CM — CSI — —

CSI vo I o ir\ I HI — CM CM ^ in o I ov \o t CM CM CM

to

© to

©

©©

©

u MW

u

O

O

_ •— u L

•— MW L. 4- 1. '4- 4- i/) MW C © LU © MW C © u 3 P u 3 UJ

(0 0 c © 0

CL o u CL o © c 1 c © c c — © © • MW 4- © • © *3 • a. MW p 4- © 4- — p d. *o CO

(0 U) o (0

©

© i/) o © • < o wJ 3 < o 2 Z)

O

O CO 00 ^ ^ CM

{fi l_©

in

oo

a^

o>

<7V ^ O CO H^ I ^ CM CM CM o o^ — I I ON I — CM n*

P^ CM

vO

O ^ CO

Csl I I p*^ CSJ oo

<0

<J1 3 ©

Q.oc

1.© • 4- to to • ©

=> 3 oO 4-

to

lO UJ

© c ©

•0 3 —

(t3 S -O s S 2 o o s I

Soiree:

Reference

27
In spite of current perceptions of an "oil glut" (28-30), oil is a wasting

resource (31-34). The era of low price and ready availability of these hydrocarbon feedstocks is growing to a close. Although deregulation of oil has

increased new drilling as well as oil prices, production of oil from existing

facilities will decline from current levels and total oil production will

probably peak around the year 2000 (35, 36). Also, natural gas reserve

additions, usually associated with oil discoveries, have been less than

production for over a decade (35, 37). With projected decontrol by 1985,

natural gas price increases have encouraged greater exploration with some encouraging results; but documented reserves of natural gas have not been

sufficient to reverse the decline in the reserve/product ion ratio (37). In addition near term prospects for decontrol of natural gas by the 1985 timetable

are not promising, and eventual decontrol greater than 50 percent is unlikely

(27, 38).

To a considerable extent, the pace at which the curtain is drawn on the

petroleum era will depend upon the oil production and policies of and the

economic and political stability of Saudi Arabia. Other key uncertainties

include; (a) U.S. and global economic growth, (b) energy conservat ion , (c) oil

production and stability of other OPEC nation, and (d) energy contributions from

nuclear, and coal, and from other sources--tar sands, shale oil, solar, biomass,

hydroelectric, wave power, etc. (35).

In spite of these projections, estimates have been made by some that 75-80

percent of U.S. organic chemical production will still be based on petroleum and

natural gas feedstocks in the year 2000 (8). Table 3.11 illustrates the variety

of strategies (e.g., securing hydrocarbon feedstock positions, forward

integration into higher value added products) that chemical companies are pursuing in response to hydrocarbon feedstock price and availability, as well as future competitiveness pressures. Table 3.12 also reflects the increasing

presence of petroleum companies in the production of organic commodity chemicals

and the location of their production facilities at the sources of lowest price

and of available hydrocarbon raw materials (41). Petroleum companies control 20

percent of U.S. coal production and 25 percent of U.S. coal reserves (35, 42).

21
Table 3.11

Feedstock Security Positions of Some Leading Organic Chemical Firms

Chemical Company Hydrocarbon Base

DuPont merger with Conoco, expanded oil/gas exploration

effort through recent acquisition (Terrapet)

Dow owns oil/gas properties*

Union Carbide purchases feedstock supplies

Monsanto oil/gas exploration revenues indirectly support some

of liquid feedstock needs

W. R. Grace oil/gas exploration revenues indirectly support

feedstock needs; has coal resources

Allied Corporation oil/gas exploration revenues indirectly support some feedstock needs, plans expanded energy exploration

effort through subsidiary (Union Texas Petroleum)

American Cyan an id purchases feedstock supplies

Cel anese purchases feedstock supplies

PPG Industries purchases most feedstock supplies

Source: Reference 39.

*Recently sold domestic oil and gas reserves to Apache. Still owns Freeport,

Texas crude oil processing refinery (Chemical Week, May 26, 1982, pp. 34-38).

\

22
Table 3.12

Location of Chemical Firms' Production Facilities at Source of Feedstocks

A. Saudi a Arabia ( SABIC) ^

COMPANY^ PRODUCT

Cel anese methanol

Dow

Exxon

ethylene, polyethylene

polyethylene

Shel 1 ethylene, ethanol, ethylene dichloride,

styrene, benzene

SOURCE: Reference 22.

^ SABIC = Saudi Basic Industries Corporation

° Production facilities at A1 Jubail.

B. Can ada

COMPANY^ PRODUCT

Cel anese methanol

Union Carbide

DuPont

ethylene qlycol

polyethylene

Exxon benzene, styrene

Shel 1 benzene, styrene, polyethylene

Dow polyethylene

SOURCE: Reference 40.

^ Production facilities in Western Provinces.

23
In spite of these company activities, a chemical industry hydrocarbon

feedstock transition will take place in the future. This transition will

involve a shift from oil and natural gas feedstocks to coal and to renewable

hydrocarbon feedstocks. Economics wi’l dictate the timing of this transition.

Technology and technological innovation will be essential components in shifts

made. The transition to alternative hydrocarbon feedstocks will greatly

influence strategic decisions being made now, and stimulate evolution and

structural change in the chemical industry.

.2 Feedstock Alternatives

The raw material hydrocarbon feedstock bases for organic chemicals will be

undergoing changes over the next few decades. Although projections indicate

that the transition to alternative feedstock bases will not be dramatic or complete, the organic chemical industry in the future will depend upon a different mix of feedstocks and on new feedstock conversion processes (43),

Petroleum is currently the primary feedstock of the organic chemical

industry. In spite of the current availability of this hydrocarbon feedstock,

there are long-term chemical industry concerns regarding its price and

availability. Since any organic chemical can be made by either chemical or biological synthesis , two alternatives to petroleum and shale oil include coal

and biomass.

The basic technologies supporting the conversion of either coal (44, 45) or biomass (46) have been known for several years. However, to improve the

economics of these conversion technologies, significant investments in R&D and new and innovative processes are required over the next several years.

In addition to capital investments in existing plant and equipment, basic

factors, such as; (a) technical feasibility, (b) process economics (cost of the

raw materi al s , efficiency of the process, cost of product recovery, cost of

recovery and value of by-products, cost of waste disposal and/or of conversion

of waste to useful products), (c) cost of process development, (d) market size

and future growth potential, and (e) present and future competition will govern

24
the choice of hydrocarbon feedstock, of target organic chemicals, and of

synthesis routes (47). Table 3.13 provides an overview of some of the economic

considerations involved in determining what alternative feedstock to use and

when to convert to an alternative hydrocarbon feedstock.

The projected time table for the commercial production of organic chemicals

via coal conversion is about 1990 (20, 50). However, recent developments in

coal conversion technology in the United States by Eastman Kodak (acetic

anhydride), by Gulf-SRC-II (commodity olefins, BTX), and by Mobil-zeolite

catalysis (methanol, ethylene) (44, 52, 60), and in West Germany (45) suggest

that the timetable may be advanced to the mid-to-late 1980s.

Although use of corn starch as a chemical feedstock in the production of

ethanol is becoming increasingly competitive at commodity levels with ethanol

produced via chemical synthetic routes (61, 62), commercial production of

commodity organic chemicals from biomass via biotechnological processes is not

expected until the mid-to-late 1990s (51). In spite of this, the chemical

industry's emphasis on conserving energy, on improving process efficiency, on reducing hazardous wastes and less caustic reaction processes, and, where

applicable, on moving to production of higher value added products favors use of

fermentation and other biotechnology tools (e.g., enzymes, plant and animal cell

cultures) in the production of organic chemicals in the long-term. Although

biochemical engineering process scale-up barriers (such as heat removal, mixing,

steril ization, instrumentation and controls, and environmental/nutri tional

influences on the stability of genetically engineered microorganisms) are present (63, 64), supplementation of traditional mutation and selection

techniques with gene amplification, fusion and recombinent ONA techniques (46,

51, 65-67) and improvements and innovations in bioprocesses (63, 68, 69) will

eventually shift the economic momentum currently favoring conventional chemical

synthetic processing approaches to bioprocesses for production of higher value

added chemicals and some commodity chemicals.

Biotechnology, via chemoautotroph ic microorganisms, can also provide a range

of organic compounds from hydrogen and carbon dioxide which are available from

coal following reaction with steam (70).

25
IIII 2

.*0 Qi

3)

— <1

fO CDC—

T3

15>>C I

tncO

m

m 1 1 c '4> 6 © 6 c 0 m — 0 C

m

©

—

>- -3

m

© 4-

—

—

© 0 m

©

m >- Q.

© >-

—

m E ©

© 3 % c 0 4- © © © © -H- >. © © © © © © >*- —

V- E m c 0 m Cl 0 —

C

—

C 3 © 0 0 © 8- 0 © © — —c —> — — © _ 2 c © 4- -Q 0 c © _ m 0

4-

© 0 3 4- © V. m © © — 3 © © 5 © © — m © 0 2 © 4- 0 c 3 c s ^ © +- 4»N © 2 C m m c © a. 0 1 m m m m ©

8- 1^ “ ©

— j)

u

©

m

m

©©

m 3 E ©

©

© c 4- c ©

©

©

L.

—

©

y ^ © 0 0 m e s •3

© o c © c m 0 J2 © © © 0 © C © •— 0 i. = 3 © W 2 — © © © 3. G. ul 4- © w i3 4- m *3 3 m 0 — ITS >- m

m

+-

m 3 4-

© “• 4» 3 © 0 0 >* CT cn

©

© 3

U)

— 8-

©

©

© ^ m© ON

m

© c © c c 3 “C C c 2 r\ in © 0 ©

m 3 — © U 3

8,

© c —

—

— c —

m c — © c © 0 0 Cl 1 © © © 0 > © 2 © c 1 >» © c © — — -C © *o c >- 0 0 © >. © — >* 0 c *—

— © c 0 4* — • >• © A © © © © 2

-2

m

© c m

©

—

— *c

m

©

m

©

© CL 0 w © 0 4- 4- — © • — © c 2 — — E 0 c C cn © — 0 — E 0 t © © c — , m m 2 c t S — © "D © 0 © J2 © 3 t 8 — — •w © © © *C 4“ O. m 3 © c S E 2 © cx m +- c m © © © © 3. 3 0 0. 3 © 3 m 0 © — Cl © © © C C m I 1 1 © s • m "c) 8) V 3 © Cl 'O 0 © C © © © m 3 0 m a. > n 0 © 4- © © c m > 2 — 0 0 0 0 0 0 0 0 1©>cO

(J

(/) in

13c5mO3©UcQ

cc©>

<J©o i! «O ! o 3 1 <n O Oo

s.o

1T3 o m — © ^ 8 6 o ° . 8 5 ^ c 8 .2

<n o i

13

O

— o • 'T

©

(/) I — O i2 4-

lr^

V

—

u c © O . 8 = mo — ^ I- O I 05O

Ou c ^ —0 ^ 4-u % 13 c i- o ^ — X

4-

V

V©

*©

3^3 — C m 2 ^ 2 O ^

i.>*

m

Q.

©

2^ c S’ - ° o U 4- C — 13 m u 'j S z 2 I m

U5

13cO

©§

s/>

mc0 m 4- 2 m © © © ©

q:

©

© 5 -3 >- 4-

©©

— 4-

— > CL m — — c

4-— 0 —

©© CJ -3

© c ... — 2 © © © X © © © — © C

4- 4-

p

—

i. < 1 cu.

O'© • • UJ CN

>*.

L.©>C

cS

rO

cons i

tier

dbIe

Investment

by

petroleun

conpanles

slon

In

coal

R&D

technology

(syntiBl

energy

Industry

developments

could

have

significant o

improvement

s/innovations

in

process

spill-over

effects

to

organic

chemicals)

technology (

cel I

ul

ose

convers

ion,

ci
(/>

nn

A I m * 'U\ 2

l/t < 1 o

-r

a iIp

!/) i J

s. cUO ;3 1 •

IS.

m

Q , ® r. I n ® t '/>

n • —

0)? ‘ Of I « . - ; n j -< i — n 0 c ^ 1 - 2 1 ri — a>

'oS 35

f*. $ • ® 3 < r. Q — r ®

C"

— !T

— O'

X

® ^ n — c = — Qi S. 0 - $

o» < = X 3 O 2 2 c ~ tn — O X

S' 0 X r- o a><

cn

X 0 !£. ^ tS X — S O

01X

o.oo

vO

Q»o

cro o *o ^ 1 a — B 3 — o> o0 —

V) ”< 1I •4* X o

3* -O ^ I ^ I s i — X 03 O i - — Q. I X too 0 O COX — a 2L

X. 2 ^ I

-f

X

X ? X

cr

X

a.

(CX2 C — 1 O

t:

n o X c. 3- 2-1 X 2 ' — O , n j ^ 2 i o 1 X 21 1 o2 O i c. , 1 ^

h.

i!

(/) 0 0 0 0 0 0 i c ; X s ly) 0 tft X X 3 s X o 3 o < < o 2 o o VI X «. 0 2 X 01 I '-" 1 c. — 5o 0 •n u> X —

0 X i1 0 0 2* 0 X 3 c — 2 X 0 O 1 s.X • 3 X3 1 X X

U1

Vft s nX 2 1

CL 2. 2. § oX 1 s 3 X3 acr

V) c a

o’ o a

a

X

® i o 2 O’ Q X X »“ X :; X

9, “ X a. o X \ 0 3 CL -4 -2 a o 2 o a n 2 5 o 0 2 —

t X U) Ui —

—

0 < X *2 X C •< X 0 —

X X — — 0

'fi

a

X « o O o — -4 o o — 3 VI — * s. o 5 2 a « o 0 — o gi *2 X -4 S'

(/> VI cr -«

Q 0 2 rs V) o -<

X

X

X

a o X o 0 n 0 X < o < o X X 0 —

— 0 < O' — 2 < o o o> 3 0 < X — X «»• Q 3 < *2 n O 0 X VI — — — X u> X X cr X *• X X a. 3 a. X — v> 0 2 2

>< 5 X

— o X • 2 3 o C. \ X 0 •• 2 c 0 X «i» •2 3* X X o cr 0 VI X C 0 x* O o “1 s •«» 2 0 3“ 3 o Q

•<

— -< o “ 0 X “1 o a V)

n »> * V*. M.

C VI VJ 3 X X X2 X s n0 2<

c. <cX o oX 1

VI O’ 3 X 3 XX

a oX

n

X

"S

X

Q

X3 5 o 2 2 s X 3 2 0 o c X tn • — 0 -4 2 — X X a

Q s 2 X -+• cn X 0 -1 X i 0 W o 0 VI 3 VI X • *1 n V) O 3 a a « 0 V) o 0 T < 2 — n 2 n 0 a X X ? 2 o VI — 0 n o n X i — < -f o X VI

(/) y) cr cn 0 1/1

X 0 o o o 0 0 O’

Q 2 X c —

Q cr o 3* 0

ss 0 tA • — y> c

iS 2 O'

X c_

X 3 3 A

X

X

a 2

2“ S — • 2

*o 0 2 3 X

_

X

X

X

X < v» i a

X

n 0 • _ 2 U) 2.

*2 a. 0 CL 0 X 3 VI 3 a.

X 0 3 O’

a i o

i/I

n

X 3

2.

0^ 2 VI cT

—

0^ 0 —

X o -4 o X 7T o X o *o V) • X X*

A 3 o X

O o X o X 3 2* X 0 X X 3 • «• •M C o i 0 “ a o — • X 2 X 0 o 3 X X O’ 2 — -4 o. 0 X 0 0 o s — o s X O M —

X 2 3 < •2 — VI X < « 3 X M. 3 c

.o • ««k VI

X 3 3 3 • o VI 0 0 V)X 2 X V) o 0 o U) o X X 3 X cx a. O’ o 0 VI X X o o o O X 2* to X O’ VI o •2 X

•1

X •< 3

'S

VI ‘a 0 0 o X — ^< Q • X < X X 0 o V? X — o X

X 0 X a 0 o

VI cr

a

O’X o o0 0 0

VI V) >< 2 X

v> 2 crX s 0

VIa 2a < 2 X 2 •< VI 3 X X VI 2. 0 1 n O* n X tn *< X o 2* Q. X 0 < ? a cr X X X “1 O C C X 1 T 3 — VI v> (A 0

•<

yrT • ' V)

'•*
o 0 0 0 0 c 0 0 0

*o 3 © 3 n

a 0 n 01 3 O *o *o 0 ©

_

n 0 © 3 ©

© 0

"1

©

© -<© 0

-1 3 ©

Q o 3 'O 3. 01 3 © 3 —• 3 « 01 0 © © c © © •o © © 01 0 -f g. 2.

©

© < ©n

—

*1 vl 5 1 01 c s O 01

01 3 ©©

© 0 ©

3" o -r

01

— O'

—

© 3 o “1

—

o"

— o «K-<» o —

01 cn — 5. 8- 3 © © CO r> 01 Q o a— 3 o 3 n

-f 3 © CO CL 0 'yi -f 3 T

© 0 CO c ©

©

© >— c © T3

Q 3 01 3 ©©

—

— 3 o -< n o 01 © © n © 1 o 3 v. -f

©

©

3*

©

© 3 3 o o 01 0 3. o © • 0 o 01 0) 3. < -f — -f 01 — 0 n — 3 O 0 ;r 0 c © 0

“1

—

3 01 o © 0 01

•aa • 3 n 71

©

© 01 c — 3 3 \

.2

— S </)

O 0 9 ©©

n

Q

— 3. o o • © 3 01 3 < © •o 3 — © Q -< © © 3 a —

© © — “1 — -< 3

3.

Q

©

— 5 O 3 ? 3 n

O

— 3 © 3 3 3 3. C 0 — © o 01 0 X ©

01 o —

i s v> 0 -t'

© 3 mmm "1

C

©

©

— ?

"3 v-n 3 O 1 0 5. 0 X 0 1 3 —

© < -1 3. -f

n 3 © CO 3 ©

fs:) CO

O T3 3* -J S'

®

-H

3*

—

—

O

— 3 3 (jy *3 ;r o —

3“

(O

O

<D

O ^ 3-o o -

to

(9

—

T u» S O o — © --* o —I U1;r oo

ca ^ B O 1

-I o oo

•n ®

O

-0 3 ® 1/1 1 -I- o — — <

ffl

—

® 3 '< O -r O 0 o ® o — 3

u) "0 — T

CL

O8

CU , ^

a, I- ^,o ©

©

©

©

© 01

a 01

©

—

©

n

© 0 €

CO 3 — -h 0 1 0 3 — *3 — © ©

© 3 3"

—

—

*o

©

—

©

— Cl

© -f

© 3 ©

© 3 3 0 CO _ 3 © © O © -1 n © © —

01 0

•<© 01

O 3. 01

(/» -• < o © o

?r *3

<• 3 - 3 O ©

-f 3

S' 3 © © I O

-t* 3- o — o

9- ^

S’ I

01 3_O « — I p ©

-+ 3 s ©

©

©

® < © © S

3.

©

© -!

— -<

© *3 3 © © © — (ft 3 -f a CO O. © n © © © © — 01 3 3. — O© ©3 "1 1© © <

*1 0 3 © © 3 Q.

01 01

01s 01 o © ©

on ‘i

©

0) 3 *3 -< ©

3.

“1Q 0_ © © n c 3 3 1

Qt3

C.

O'

—

n

Q. o © — 3 3 —

0)© 3

Q» ^ © -N

3*©

O" *s — c — 0 o o» — o ? O

O 3 — 3 O

— o ©

01 01

01

— < o ©

-f» 3© © o

-»• uc

3* •<

3-

O

—

"O5-

n o

0)— *o S 5 o — © o

01 01

—

—

©

S'©

0)n 3. 0 S

-f

©

© c 0 © n n < 3 n 3 3 — CO © c © 3.

01 < “1©© 3.

X

©

© •< 3 3.

© ^ 3

*ooo

0)

-f 01 ^ o

T* S ©

3.

—. — 01

©o

3*3O o 3 © Ctf

—

O

©o

9L ^ — ©

01

01

©

©

01 01

S' 1 o —

— 01 3< 9“ 0 ©

01

=r

'< *3

©

n

© 3 3 X -f o © -f 3 “1 3 © © 0 © 3 3 n 3

©*

©

© 0 c n

— 3. c 3 ©

01

©

*3

© 01 -T© 5. 3

O. • 01 © © 3 —

*1

© 3 © "1 © CO

'<

*o5.

01 2 o 0 c ©

01

©

01 cn©

©©

CL

—

— 01

© 01 o 3

CD (II
I I ... 0 -3

in 0 c ... in

Q 0 0 L. 0 0 in c o u .c 0 c o o 0

h.

4- o 0

JZ

b in in 0c i 4- 4-0 0

in o

in t3 0 0 0 m 0 0

in .o 4- 0 O

u CL c

L. 4- 4- c E O

8. o 0 in 0 o c4- 4- 0 U

—— 4- 0

in in 0 4- * 0 — 8 0

Q. >0 0

*3

4-C

4- w. 4- 0 0 O 0 —

>-

U 0 3 in c L. c c ..

in mmm 0 a. 0 CD 3 0 U 0

in > a. c t. 0 im Lm s2

o. 0 (T3 C ... u T -3 0 £ O 0 o in 0 c Lm mm 0 0 •— o •mm 8 u 0 0 u c c •— 4- 0 Lm CD a. 0 3 0 • m, Q 0 *3 3 0 .Q £ in '+- in C •3 Q b

... o 1 0 0 in

C

m 0 1 •3 0 o 0 m 80

(j c in

C 0 4-

8, in oti 0 *3C 0 c3 -3

C

+- 3 »+- 0 4- 0 c 0 C

*0

—

— *o JZ 0 £ 0 0 a.

1. c ...

O o o > 0 M.

m 3 0 u 4- L. "3 4- E 0 0 3

L_ Q.

u 0 0 g cn

C

U) 0 — in 3 4- 0 5 L. 3

(0 c C *o 0 c c C

C Lm .. 0 0 s •mm 0 c .c 8) o 0 0 0 .C

(7! in O) 0 2 o E in > o b 0 O 0 O o 0 c *3 0 •—

•—

U

4- 0 0 s o o _ 0 0 E © 0 — "3 0 3 4- y

•mm

—C

C 0 X

U 0 L. o

4- c in C s TD in C 4- 0o in 0

l.

C0

T5C 8)

C0

4- *— 0 0 E > E

J= 0 • C c in 0 3 0 JZ 3 3 M. 0 m C

O) 4- 0 -3

U 0 —0 c C C 0 U •mm 0 0 4- 0 0

inm

— 0 "3

u 0 in 0 0 4- 0 Lm _ X s o Q. 4- 8, 0 % 0 o C .C •— 0 o fe. -o 0 0 4- 3 *3 c C 0 in 0 X S <J n3 0

L. 0 3 0 • N in *3 .. -3 ON

*—

X >. 3 in

— 0 in 3 4- in

C 1 (0 4- _ 0 0 oo o U JZ 0 >* 0 0 in Lm

mm.

— in *3 in c 0 — S 3

fe) 0 u * 0 0 4- L. 0 Lm — .. c 0 in O) 0 C •mm in c

L.C 0 cn c im ..

—

m 0

8,

Lm 0 0 E 0

Q. E £ 0 L. o O O 0 O o

CM

CM

•—

m

in 0 00 s 4- c0 in

in 4-

—

a 0 3 in (0 •— o

<0 0 u

O .. c

ct o 0 0

in 0 0 © O Li. 4- in Lm u •» +* C c 0 > CL in O) u 0 c c 3 E 0 4- — 0 ... 8 c 4- QC

(T3 (0 o u

O 0 C *— 3 U Lm U U u u Q. 0 • • 0 0 o 1 > *3 0

+- 4-

g L. >•

C 0 < 5 c

CL CD LU in

V

Lm 0 • • • 3

(J CM K) 0

Ui *—

— </)
In spite of deficiencies in data on the economics (a) of biotechnological

processes and (b) of starting materials in the production of organic chemicals,

some examples of possible targets of opportunity (specialty and commodity

organic chemicals) are listed in Table 3.14. Although several of these

chemicals represent higher priced/value added chemicals, possible future

increases in their volume of sales could shift these currently high priced, low

volume chemicals to low cost/high volume commodity chemicals. Critical to

improving investment decisions on (a) industrial scale production and (b) the

economics of a bioprocess to produce a particular product is a fundamental

understanding of cell metabolic pathways. Information on pathway throughput

capacity; on pathway intermediates; on pathway enzymes, their stability, the

amount of enzymes produced and enzymatic reaction rates; on substrate/ product

relationships; and on the significance of the metabolic pathway will be

essential for industrial investment decisions on bioprocesses (72).

.3 International Competitive Trends

Chemicals, particularly primary and intermediate organic chemicals, are important contributors to the U.S. export performance (Table 3.1) and chemical

industry profits, and an increasing proportion of domestic chemical production

has been exported (7, 23, 73). However, a number of factors will tend to

moderate the rate of growth of exports of U.S. commodity organic chemicals and

increase the rate of growth of organic chemical imports.

Some U.S. export moderating factors include:

0 continued near-term reliance on petroleum and natural gas as hydrocarbon

feedstocks and declining feedstock price advantage;

0 lower demand for organic chemical -dependent products by chemical industry

customers, such as automobile manufacturer, building construction, fibers,

plastics industries;

0 domestic and foreign economic pressures and currency val uations; and

0 protectionist barriers.
Table 3.14

Sorne Possible Targets of Opportunity for Production of Or^nic

Chemicals via Biotechnological Processes

Product ion Sales Quant it/ Pr i ce Category Chem i ca

1

(xIOOO lb.) (X 1000 lb.) ($/lb.

Am i n o Ac i d s g! ufamic acid 650,000^

129,000^

NA 1.80

lysine NA 2.10

met h ion i ne 210,000 NA 1.20

Vi tamins^ (total

)

42,577 26,535 9.04

vitamin E 7,254 5,287 16.23

0. A 1 i phat ics^ acetic acid (synthetic) 2,976,772 514,995 0.18

aery lie acid 61 1,172 68,966 0.40

adipic acid*^ 1,500,000 NA 0.57

ethanol (s^'nthetic) 1 ,450,769 1 ,159,446 0.27

ethylene glycol 4,385,731 3,008,147 0.27

ethylene oxide 5,220,400 530,986 0.34

g lyceroi 140,578 137,784 0.57

prepy lene glycci 487,526 426,762 0.37

propionic acid 102,975 69,753 0.23

Arcmat ics^ aniline 659,421 1 16,223 0.35

benzoic acid 73,414 32,220 0.40

phthal ic anhydride 818,247 418,519 0.34

A 1 gi nates^ (e.g. , sodium alginate) NA NA 2.00

Enzymes ( for 20.00

industrial bioprocesses) — 22,000^ NA to 700.00

Pept ides'" (e.g., aspartame)^ 40 NA 1 10.00

Polymers (for fibers)" — 6,583,357 454,338 0.58

Polymers (water soluble)^ — 316,016 269,720 1.38

Plastic and resin materials^ — 1 1,753,214 9,606,419 0.66

Rubber^ — 2,798,951 2,249,397 0.27

NA = not ava liable

^ U.S. production only. (Source: Synthetic Organic Chemicals, U.S. Pr oduction and Sales, 1980. ) 1982 prices are about 30 % higher than 1980 prices. 1982 production is about 12 % lower than 1980 production,

b Source: Reference 72.

c Source: References 51, 71. Peptides could be used as a nutrient medium component (synthetic) for cel I culture

on a laboratory and industrial scale, and as biological regulatory substances,

d source for U.S. production information

-Chemical Products Synopsis , Mannsvi I le Chemical Products, August 1981.

^ Aspartame, a synthetic sweetener, consists of the amino acids aspartate and phenylalanine.

Represents global production. Japan, primarily through Anjincmoto Co., Ltd., produces about 70^ of the world

volume of amino acids. Glutamate, methionine and lysine currently represent over 99| of the world production

of amino acids. (Source: References 51, 71, 72).

^ Represents global production. Microbial amylases (94$) and proteases (5$) represent 99$ of current global

commercial production of bulk enzymes. Major companies in bulk enzyme production are foreign owned (for example.

Novo, Gist-Brocades, Miles-Bayer AG). A U.S. commercial bulk enzyme producer, Rohm and Haas (enzyme production

facilities), has been acquired recently by Corning Glass Works. Prodjction figures are under est i rated since firms

such as CPC, Clinton, Staley produce starch hydrolyzing enzymes for captive use. (Source: References 51, 71).

31
Some U.S. import enhancing factors include:

0 emergence of Canada, Saudi Arabia, and Mexico as major commodity chemical

manufacturers;

0 chemical industry industrialization plans and export strategies of developing countries such as Brazil and Saudi Arabia; and

0 industry nationalization and state/government ownership and direction of

industry and industry economic incentives in some countries.

These and other factors, individually or collectively, will over the next

decade and one-half present significant challenges to the international

industrial competitiveness of U.S. organic chemical producers, and may cause

U.S. producers to lose their position in world commodity chemical markets.*

The following will highlight some examples of the pressures from industrialized and developing countries that will influence the U.S. commodity

organic chemical industry competitiveness over the next several years.

a. Trade Liberal ization . The European Economic Community (EEC) is the world

leader in both chemical exports and imports (7). Nearer term competitive

pressures from the United States and Canada as well as intra-EEC exports may

force structural changes in the European commodity organic chemical industry

because of overcapacity, weakened economies, lower profit margins, and projected

pressures from producers in Saudi Arabia and other countries in the Middle East.

In spite of the Tokyo Round of Multilateral Trade Negotiations (MTN), nearerterm responses to these mounting pressures may include actions counter to trade

liberalization. These may include delays in implementing multilateral trade

agreements, shifts to bilateral trade initiatives, import relief through protectionist policies, joint industry ventures, countertrade agreements, mergers,

plant closures, government ownership of chemical industries, and government

directed competitive practices and industrial policies (22-25). The outcome

*0ne projection indicated that U.S. net chemical trade will decline at an average annual rate of 15 percent between 1980-1985 (Chemical and Engineering

News, May 24, 1982, pp. 21-22)

.

32
from these pressures affecting the European organic chemical industry and from

their subsequent responses to these pressures will probably be considerably

fewer producers in the commodity chemical marketplace and an industrial movement

to higher value added organic chemical products. European chemical industry

interest in biotechnology as one alternative route to production of higher value

added products is illustrated (a) in Table 3.15, (b) by West German industry and

government substantial investments in this area since the early 1970s*, (c) by

Elf Acquitaine's investment in Engenics (16), (d) by the French Government's

increased investments in biotechnology and efforts to orchestrate a national

public-private sector initiative in this area (74)**, (e) by the United

Kingdom's Agriculture Research Council hybridoma research center (75), and (f)

by British Technology Group investments in Speywood Laboratories ana in CeUtech

(76, 77).***

Japan '

s

commodity organic chemical industry, without local access to raw material hydrocarbon feedstocks, may be facing pressures even stronger than

those being experienced by European chemical manufacturers, including substantial competitive pressures from the United States and Canada. Industry and

government responses to these pressures have focused on protectionist measures

and industry cartelization (21, 22). However, like the European industry,

chemical industry structural changes will be required in the long-term, probably

resulting in fewer companies and with a movement toward production of higher

value added products. Japan, however, plans to utilize its industrial

experience in fermentation technology (for example, Anjinomoto Company Ltd. and

MITI's Fermentation Research Institute) to leverage its industrial movement to

high technologies, such as biotechnology, and to higher value added products.

*It is estimated that the West German government invested about S60-65 million

in biotechnology R&D between 1972-1978, including funding responsibility for

the Gesellschaft fur Biotechnologische fur Forschung (Nature 283 : 124, 1980).

**The French government's funding in biotechnology is expected to be about $35

million in 1982 (Chemical Week, March 10, 1982, p. 36).

***By the year 2000, the European biotechnology market is estimated to be $16-17

billion (Information Research Ltd., November 1981). Also, of the 125 firms

currently in fermentation production, nearly one-half are European (L. Hepner

& Assoc., London, February 24-25, 1982).

33
Table 3.15

Leading European-• based Chemical Com pan i es

Biotechnology Investments

1979 Sales Home Equity ownership;

Company ($ million) Country In-house other

BASF 16,014 W.G. X Heidelberg Un iv

.

Hoechst 15,474 W.G. X Mass-Gen. Hosp.,

Hoechst-Kawagoe

Bayer 14,953 W.G. X Miles

ICI 11,757 U.K. X Cell Tech

Rhone Poulenc 7,719 Fr. X Transgene,

Genetica,

Institute Merieux

Montedison 7,592 Italy X Adri a Lab

.

DSM 6,711 Neth

.

Akzo 6,324 Neth

.

Ciba-Geigy 5,557 Switz. X DNAX Res.; A1 za Henkel 3,886 W.G.

Solvay 3,873 Belg. Biochem

Sources; References 13, 16, 18, 73.

34
This long-range government-industry strategy was emphasized in the report.

Mill's Vision of the 1980' s (78, 79).* Examples of Japanese implementation of

this strategy with respect to biotechnology include the following:

0 formation of a chemical industry consortium in biotechnology (Mitsubishi,

Sumimoto, Mitsui, Asahi, and Kyowa);

0 formation of a 14 company biotechnology research organization in

technology areas key to the successful implementation of industrial

strategies in biotechnology and in areas that will complement existing

industrial experience in fermentation technology (Table 3.16); and

0 Japanese firms' active interest in investment in and establishment of

cooperative technology transfer and other agreements with IJ.S. biotechnology firms (Table 3.17; 80-86).

This approach certainly reinforces trends to reduce their dependence on high priced hydrocarbon feedstocks and to embody greater value and higher

technology in organic chemical products.

Developing countries such as Mexico and Brazil were not included in the 1979

MTN agreements and codes, and, under the Generalized System of Preferences

(GSP), qualify for tariff-free access to U.S. chemical markets (22, 23). The

rationale behind GSPs is to allow developing countries to establish an industrial base. Although this approach is reasonable, it is extremely difficult to

determine when a developing country has developed a base for an industry and

when GSPs are no longer required. Two of the objectives of Brazi 1

'

s

Proal cool

program for production of ethanol from sugar cane are to reduce dependence on petroleum imports and to improve foreign currency exchange. Of the program's

1985 goal of 10.7 billion liters, less than 30 percent (3.1 billion liters)

would be used for blending with gasoline (87, 88). With only a small percentage

*Japan's industry is currently spending about $200-220 mil lion per year on biotechnology R&D. MITI is predicting that the value of Japan's biotechnology

industry will be $21-34 billion by the year 2000. It is estimated that the

biotechnology contribution of Japan's chemical/fermentation industry will be

about 75 percent of these year 2000 projections.

35
Table 3.16

Japanese Biotechnology Research Organization

10 Year Biotechnology Research Program

Government

Area of Research Funding

Emphas i s (millions) Compan ies

rONA applications $ 43 Sumitomo Chemical

'Mitsui Toatsu Chemicals

Mitsubishi - Kasei

Institute of Life Sciences

Bioreactor development $ 43 Kao Soap

Daicel Chemical Industries

Oenki Kagaku Kogyo

Mitsui Petrochemical

Industri es Mitsubishi Gas Chemical

Mitsubishi Chemical

Mass cell culture technology $17-22 Asahi Chemical Industry

Aj i nemo to

Kyawa Hakko Kogyo

Takeda Chemical Industries

Toyo Jozo

Sources: References 81, 82.

36
Table 3.17

Examples of Japanese Investment in and

Collaboration with U.S. Firms

Japanese Firm

Mitsui Toatsu Chemicals

Takeda Ahiza

Green Cross

Mochida

Nissho Iwai Corp.

Kinto K.K.

(trading company)

Japanese Financial

Institutions (unnamed)

Mitsubishi Chemical

Industries

Yamanouchi

Toya Menka Kaisha, Ltd.

Mitsui

Fujizoka Pharm.

U . S . Firm

Gen ex

Gen ex

Bristol-Myers

Collaborative Research

Genex

Interferon Sciences

G. D. Searle

Cetus

Enzo Biochem

Genentech

Genentech

Hybri tech

Schering-Plough

Biosearch Lab.

Gentronix/EMV Assoc.

Biotech Res. Lab.

Investment or Biotechno logy

Area o f Interest

urokinase; genetic

engineering technology

enzymes

interferon

equity; urokinase

human serum albumin

interferon

interferon

equi ty

develop R & D projects

with Japanese pharmaceutical and

chemical investors;

establish subsidiary

f i rms

equity

human serum albumin

monoclonal antibodies

interf eron

polynucleotide

synthesizer

biol og ical -molecul ar electronic circuits

monoclonal antibodies

37
of the balance being used by the domestic chemical industry, a significant

volume of competitively priced ethanol could be released for export to industrialized nations. In addition, Brazil's low technology-based turnkey plant

operations for ethanol production could be exported to other developing

countries (87, 89). Brazil may also develop other low technology bioprocess

approaches for world scale production of organic chemicals such as acetic acid.

'//hen Mexico'

s

government-owned Pemex petrochemical complexes produce a sufficient volume of commodity organic chemicals to satisfy its growing domestic

market and when a sufficient industrial infr astructure is developed, Mexico

will, because of its raw material feedstock base, become a significant commodity

chemical exporter (21, 22).

b. Import Pressures from Neighboring Countries . The United States is the

world's largest (and still growing) market for industrial chemicals.

As indicated above, products from Mexico's commodity organic chemical

industry may be competitive in the future in United States markets, although

U.S. organic chemical imports from Mexico presently represent only 2-2.5 percent

of U.S. imports (73). Mexican government intervention in the market place, for

example, currency devaluation, and the development of an industrial infrastructure will in large measure determine Mexico's future organic chemical

industry export successes and the timing of these successes.

Despite the economic climate and current falling demand, Canada , and the

world scale production facilities coming on line in the Western provinces, will

provide substantial competitive pressures on the U.S. commodity organic chemical

industry in the near term (24, 40, 90). Already Canada, the major source of

U.S. chemical imports, exports about 24 percent of its petrochemicals to the

United States (73). A number of factors will enable Canada to double its share

of world production, particularly of commodity organic chemicals such as ethylene, benzene, polyethylene, styrene, etc., by the mid-to late 1980s and

make significant inroads into U.S. domestic industrial markets. Some of these

factors include: Canadian Government intervention to fix natural gas prices

below comparative oil prices, the current magnitude of and plans to expand

capacity, the value of Canadian currency, and the increasing petroleum costs of

its major trading partners (United States, Japan; 21, 22, 40, 73, 90).

38
c. Middle East (Saudi Arabia) . As Saudi Arabia is a critical determining

factor in the future price and availability of petroleum, so it may also become

a significant competitive factor in the global production and trade of commodity

organic chemicals. Table 3.18 provides some primary organic chemical oost-1985

pi^oduction plan estimates for SABIC operations. The overall goals and targets

for Saudi Arabia exports are listed in Table 3.19.

In addition to the availability of a hydrocarbon feedstock, Saudi Arabia

projects that the construction of modern, flexible chemical manufacturing plants

will help it to gain a competitive edge in international markets over industrialized countries with out-of-date production facilities. Because of

construction problems and other industry infrastructure considerations and

barriers, whether the timetable for Saudi Arabia plant commerci al i zat ion is

achievable and the projected scale of production is realizable is the subject of

much discussion. However, in spite of these difficulties, Saudi Arabia's global

market share of commodity organic chemicals will increase, but it may take

longer to reach its target share than it originally projected (22, 74).

These represent some of the major pressures facing U.S. producers of

commodity organic chemicals over the next several years. The magnitude and

integration of the domestic chemical market and U.S. chemical industry

management and technological know-how will help U.S. manufacturers to weather

some of these international competitive pressures from new commodity chemical

producers. One alternative, but not exclusive approach, to coping with these

competitive forces and feedstock disadvantages would be for the U. S. organic

chemical industry to use its technological edge to produce higher value added,

high technology products.

39
Table 3.18

SABIC* Commodity Organic Chemical Production Projections

Projected (mid-late 1980s)

Annual

Product ion

Chemical Complexes Commodity Organic Chemical Capacity (mil 1 ion lbs)

SABIC +

National Methanol methanol 1400

(Celanese, Texas

Eastern)

Saudi Methanol

(Japanese Consortium)

methanol 1300

Petrokemya (Dow) ethylene 1100

Eastern Petrochemical ethylene glycol 660

( Mi tsubi shi

)

polyethylene 286

Kemya

( Exxon

)

polyethylene 572

Sadaf ethylene 1440

(Shell) ethanol 618

ethylene dichloride 1000

styrene 649

Yanbu ethylene 990

(Mobil

)

Table 3.19

SABIC Commodity Organic Chemical Export Targets

Percentage of

Saudi Arabia

Exports Absorbed

Target Country (goal

)

United States 20%

Japan 20%

Europe 22%

Middle East and African Countries 38%

SABIC = Saudi Basic Industries Corporation

Source: Reference 22.

40
.4 R&D, Technological Innovative and Industry Prof i tabi 1 ity

The chemical industry is an R&D intensive industry. A high percentage (97

percent) of its R&D is supported directly by private sector chemical firms (91).

Table 3.20 illustrates trends in chemical industry R&D spending. It is

estimated that petrochemical (organic chemical) R&D currently represents about

two-thirds of all non-drug R&D expenditures (6). Firms witn 25,000 or more

employees and with annual R&D expenditures greater than $100 million represent

about 50 percent of the total industry R&D spending (91).

The data presented in Table 3.20 show that the annual growth rate in

industry R&D spending for the past decade was about 10 percent. However,

estimates of total industry R&D spending from 1980-82 suggest a renewed

commitment to investments in R&D. For example, the annual growth rate from

1979-1980 was 14 percent, from 1980-1981 was 16 percent, and from 1981-1982 is

expected to be 17 percent. When this rate of increase is adjusted to projected

rates of inflation, there is a real growth (or a real expansion of R&D activity)

of about 7-10 percent (92), as compared with a real growth rate of 4.5-5 percent

in the previous decade. Also R&D as a percentage of sales, at a low of 2.5

percent in 1979, is projected to be at 3 percent in 1982, similar to the 1973

level (92)

.

There are a number of factors that will maintain and enhance trends in

industry R&D investment levels. Examples of these factors include:

0 new entrants to and increased international competition;

0 world overcapacity pressures in commodity organic chemicals;

0 regulatory cost requirements;

0 price and availability of hydrocarbon feedstocks;

0 Economic Recovery Act of 1981;

0 need for innovations in process technology to increase efficiency of

existing and of new operations;

0 use of alternative feedstocks;

0 need for automation to optimize plant operations and management and for

flexibility in new plant design to meet market demands;

41
Chemical

Industry

R&D o ^n

— in

r*

m CO in

ON o CM

•—

CM ON ON

tn o

ON CO o in CM

CMr*

Ov

CNro

ON

O o Oinos

inmooovOo

CMoCM

CM

ON

ONCM

ON

CO

ON

vO

GO

ON

ONam

CM ON o

in o nr

in

CO in

—

NO nr

in o ON

p^ p^

GO CM

NO GO CM M- NO O in

CMO

O o o m

pn NO NO CM *—

CM

ON

L. • CL m 10 CM • 5 0

nr •I- O)

ON 0 c E •—u 0 •H 0

10 0

to 0 c B— c O) 0 0 *o

to MM c u 0 s •

to 0 nr CO < o ON c *—

_ (0 E a, l_ 1. 0 m 0 0 ON 0 1. o c 3 to + c 0

CO o 0 (J 423c

to od

to

(0 q:

to MM E - 0 c 10 0 0 0 0 0 -H 0 M. S o •M u u 0 — 0 3 u O s [A — u MM +- E 0 *i E 0 — *— — 10 0 to Im «M 0 0 O MM 1. 3 O) 0 0 o to u s 0 0 a. *3 3 J= o o CJ LiJ 3 C 0 o — Q o 0 a U </)

CM

i;
0 possibilities of new processes and products via biotechnology (Tables 3.6

and 3.7) ; and

0 promise of discovering or of acquiring higher value added speciality

oroducts such as pharmaceuticals (23, 25, 92, 95-97).

Investment in and ultimate successes in research and technology development

are but one element in determining the economic success of an innovative product

or process and the level of industry profitability. Recent experiences in the

chemical industry bring home the fact that possession of state-of-the-art

technology is little consolation in the face of global overcapacity (95). The

probability of the success of a product or process resulting from a firm's R&O

depends on three separate but interdependent factors: 1. the probability of

technical success; 2. the probability of commercialization; and 3. the

probability of economic success (98).

The research and technology development efforts of major organic chemical

industry producers can be categorized as dynamic rather than static. These

firms have but one goal--to avoid industrial stagnation and decline by rollingover investments (99). Proponents of the "long wave" theory suggest that

current industrial upheavals and the technological ferment, also character! st ic

of the current status of the commodity organic chemical industry, may indicate

the end of a "long wave" (100-103). Here there is a struggle between

establ-ished industries and the industrial identification of those technologies

that will become the bases for the establishment of new, future industries. In addition to technological innovation, a major force influencing the initiation

of each "long wave" in the past appeared to be energy (fuel wood, coal,

petroleum and natural gas). In the future, technological innovations linked to

energy efficiency may be the driving force in the next "long wave". Two key

technologies with important long-term industrial and economic implication, and

that also meet this energy efficiency criterion are microelectronics and

biotechnology. The identification of important future technologies and the

timing of an industry's move to these technologies as an integral part of its

plans for future growth are critical factors in the future viability and

profitability of a firm (104). Present investment in R&D on a variety of

fronts, such as appears to be the pattern of major organic chemical firms, will

create the options necessary to facilitate future industry decisions to move to

43
new technologies. Decisions on company selection of specific new technologies

and on the timing of company investments in technologies selected will require a closer integration and responsiveness of current industrial R&D efforts to

future industry business prof itabi 1 i ty and success requirements (95).

.5 Industry Strategies

In spite of the inherent technical structure that defines functions within

the organic chemical industry, there is considerable heterogeneity in individual

company motivations and stakes in this industry (19). Thus, no single industry

strategy would be applicable in addressing the many forces and challenges

influencing the survival of producers of organic chemicals over the next two

decades. Table 3.21 outlines some of the forces that are guiding industry

strategic planning. Tables 3.22 - 3.24 list some of the factors that will

influence industry strategic decisions.

In addition to the dynamic influences of economic and political forces,

major challenges that will affect the strategic plans and directions of

commodity organic chemical industry include the following:

Feedstock and Energy

0 feedstock security

0 feedstock availability at competitive prices

0 feedstock flexibility

0 alternative hydrocarbon feedstocks and synthesis processes

0 process efficiency

Growth and Differentiation

0 R&D investments in process innovations vs. development of new and

innovative products

0 high technology, higher value added specialty and fine chemicals vs. commodity chemicals

44
Table 3.21

Overview of Some Forces Influencing Strategies of the

Commodity Organic Chemical Industry

Industry Strengths,

Weaknesses, and + Other Considerations

Industry

Competitive

Forces

+ External

Inf 1 uences

0 technology 0 competitors 0 social

0 manpower 0 suppliers 0 po 1 i t i c a 1

0 raw materi al s 0 buyers 0 economic

0 products 0 markets 0 technical

0 manufacturing/ 0 new entrants 0 Governmental/

distribution requl atory

0 foreign trade

0 margins/profits

0 internal competition

0 structural adaptab i 1 i ty

Sources: References 105- 107 ,

45
Table 3.22

U. S. Commodity Organic Chemical Industry

Strengths, Weaknesses, and Other Considerations

Technology

0 increased real dollar expenditures for R&O; technology leadership position

0 emphasis on process >^esearch (e.g., process energy efficiency; position

alternative feedstocks)

0 broad range of RS<D as window on future technologies and on new markets

0 proprietary knowledge base

Manpower/ Management

0 organizational ability

0 process engineering and other high skill position requirements high

Raw Materials

0 increased emphasis on geographic positions relative to feedstock sources

0 priority placed on feedstock security as a fuel, chemical feedstock

and/or revenue source

0 need for development of alternative supplies of hydrocarbons for chemical

feedstocks (synfuels, biomass)

0 increasing presence of petrochemical companies

Products

0 need for product differenti ation--f ind specialty product niches; new and

higher value added products (by acquisition, R&D)

0 greater need for successfully exploiting patent protection

0 need for continued reliance on maintaining complete product lines

0 maintaining quality of products and reliability of supplies to markets

Manufacturing/ Distribution

0 over capacity, lower demand, lower productivity

0 world scale and highly integrated plant complexes

0 good supporting industry service infr astructure

0 proximity of manufacturing operations to energy/chemical feedstocks

0 capital requirements for plant/equipment expansion and modernization

Foreign Trade

0 increased global trade competition

0 decreasing rate of growth of exports and increasing rate of growth of

imports

0 Eurooean recession plus strong dollar equals curtailed demands for U.S.

exports

Margins/Profits

0 slower growth, profitability projections

0 maturing industry, declining rate of penetration of USGNP

0 increase is costs of oil and natural gas

0 declining share of world productive capacity

0 recent rapid increase in value of petrochemicals more from price than

quantity increases.

Internal Competition

0 serious price competitions forcing industry structural change--shakeouts

Structural Adaptability

0 flexibility to use alternative feedstocks for fuel and chemical feedstock

needs; capital requirement demands for flexibility

0 manufacturing plant obsolescence and capital invested in existing plants

and processes

0 pressures between low growth projects and need to develop innovative and/or

retrofit existing facilities in plant design

0 need to build management of plant into new designs

46
Table 3.23

Industry Competitive Forces

Competitors

0 increased price competition from industrialized countries

0 growing competitiveness of developing countries; exemptions from GSPs

0 middle east petrochemical capabilities for share of global capacity

0 dynamic technologies for production of commodity chemicals and impacts on global economics and competitiveness

0 foreign competitor investment in plants in United States and impact on U.S.

markets

0 increasing presence of Canada as major exporter to United States of

commodity chemicals causing reduced U.S. competitive advantages in domestic

and some loss of position in world markets

0 excess capacity, declining demand for certain U.S. commodity chemicals and

increasing capacity of Canada, Mexico and Saudi Arabia

0 trading partner willingness to live up to MTN agreements

0 plant obsolescence in Europe

0 prospects for reduced number of chemical companies

Suppliers

0 decontrol of U.S. crude oil

0 feedstock availability and competitive pricing of feedstocks

0 increasing probability of petroleum feedstock shortages

0 rising feedstock and fuel prices

0 pace and extent of natural gas deregulation

0 feedstock shifts

0 expanded oil and natural gas recovery efforts

0 declining domestic supplies and increased costs of oil and natural gas

0 coal as an increasingly important feedstock (fuel and chemicals)

0 nation/state control of operations and of exports

Buyers

0 economic health of major chemical customers--housing, autos, plastics,

fibers, paint, rubber

Markets

0 United States has large and integrated domestic market

0 United States has growing market and is target for foreign competition

New Entrants

0 new sources of higher value added chemicals expected and impacts on smaller

chemical firms

0 price competitiveness of substitute materials vs. organic chemical end

products

0 food processors (corn to ethanol), forestry companies (wood to organic

chemicals) and other new competitive forces in domestic markets

47
Table 3.24

External Influences

Soci al

0 aging workforces (industrialized countries)

0 worker entitlement demands

0 unemployment, workforce disruptions, economic conversion needs

0 workforce education/training/safety

0 public perceptions of chemical industry

Pol itical

0 delays in deregulation of natural gas and real extent of deregulation

0 foreign country investment policies

0 protectionism/reciprocity/import 1 0 responses to increased trade press

United States, Asia) of manufactur

Economic

0 emergence of internationl trade in

0 debt positions of countries and tr

0 disurptions in balance of payments

world credit markets

0 high interest rates, availability

0 national economic downturns and si

chemical industry outputs (autos,

0 high underlying inflation in some

0 currency manipulations to improve

0 government deficits

0 linkages between European and U.S.

0 value of dollar vs. other currenci

0 global interdependence considerati

Techn ical

0 increased pace of technology trans

0 patent protection and longevity of

icense policies/cartel formation

ures on Europe because of imports (from

ed good with high chemical content.

U.S. economic plans

ade efforts/strategies to service debts

and global money flows; defaults in

of capital

owed consumer demands for some of

housing, clothing)

countri es export positions

economies and slow growth projections

esons

fer and of technological change

patents in foreign countries

Government/Regul atory

0 foreign government industry nationalization, national industrial policies,

nation/state support of uncompetitive industries

0 chemical industry priority for available feedstocks and government

allocation priorities

0 regulations and their costs (air quality standards. Clean Air and Water

Acts, National Contingency Plan, environment/health/cancer policies, TSCA

and pre-manufacturing notices. Resource Conservation and Recovery Act, FDA

and/or USDA requirements)

0 shifts from multilateral to bilateral trade negotiations

0 growing foreign government/state owned and directed competition (new

chemical producers with access to low cost feedstocks)

0 foreign national/state policies on price and supply of energy feedstocks,

subsidized worker training, preferential access to capital and to

restrictions on quantities of imports

0 pace and costs of country/state development of industry service

infrastructure

0 government subsidies

48
Marketing

0 capacity vs. proprietary base

0 market share increase vs. competitive advantage increase for market share

niche

Biotechnology has and will play an important role in the investment

decisions of firms and in the long-term future of the commodity organic chemical

industry (97). However, substantial impacts of this technology on the

commerci al products and processes of this industry will probably not begin to

take place until

1995-2000.*

* In the interim, industry investments in

biotechnology will provide organic chemical firms with a window on a potentially

significant technology, as well as expand their range of strategic options.**

*The value added component of commodity organic chemicals (SIC Codes 2865,

2869) in 1980, based on a total shioment value of $54.2 billion, was $22.6

billion (1981 U.S. Industrial Outlook, Bureau of Industrial Economics, U.S.

Department of Commerce, Washington, D.C., January 1981). It is estimated that

by the year 2000 biotechnology could contribute $1 billion (William Chang,

Corning Glass Works, personal communication. May 25, 1982; Daniel Wang, MIT,

seminar, January 22, 1982) to $7-10 billion (Chemical Week, September 30,

1981, pp. 36-41; Industry Week, September 7, 1981, pp. 67-70; T. A. Sheets

Co., Cleveland, February 24-25, 1982) in constant 1980 dollars to the value

added component of commodity organic chemicals. The conservative estimate ($1

billion) may be more realistic, since biotechnological feedstocks and

processes for the production of commodity organic chemicals will not easily

displace existing petrochemical feedstocks and production processes. Both

estimates, however, are focused primarily on projections for traditional

commodity organic chemicals, and do not necessarily take into consideration

expansion of uses for existing organic chemicals or development of new,

"non-tradi tional" commodity organic chemicals such as biopolymers (Inside R&D,

May 12, 1982, p. 4; Biotechnology Newswatch, April 5, 1982, pp. 1-2). Market

impacts for these new, "non-tradi tional " commodity organic chemicals are unknown but could adjust upward significantly the above estimates.

**Conservativel y, about $500 million has been privately and publically invested

in new biotechnology companies to date. Most of these investments have been

made in the last two years. Of the amount invested in new firms, about

one-half has come from venture capital sources (Chemical and Engineering News,

March 29, 1982, pp. 10-15; Genetic Engineering and the Engineer , National

Academy of Engineering, National Academy Press, Washington, D.C., 1982).

Figures are not available for established chemical, phaimiaceut ical , agricultural, etc. firms in-house investments in biotechnology. A projection on the value of the U.S.'s biotechnology industry is $64 billion by the year 2000

(Genetic Eng. News, May/ June 1982, p. 5).

49
Projected initial impacts of biotechnology on this industry will be most

likely in the higher value added specialty chemicals. This may fit closely with

the strategies of those firms lacking a secure feedstock base. Also, some of

these specialty chemicals may, in turn, become the commodity chemicals of the

next century.

In addition to products, biotechnology will also influence process

developments and selection (63), and industry structural changes via new

entrants and use of alternative (biomass) hydrocarbon substrates (53, 57, 61).

Although it may not be evident in the near term, changes now in motion both

internal and external to the industry will significantly affect both the form

and substance of the industry by the close of the century.

50
4. BIOMASS FEEDSTOCKS FOR ORGANIC CHEMICALS

.1 Introduction

The decade of the 1970s demonstrated clearly our national industrial

vulnerability to foreign sources of petroleum. Increasing economic, industrial

and political concerns about the continued availability of petroleum and natural

gas at competitive prices has stimulated efforts to seek alternative sources of

hydrocarbon raw material s, such as coal and biomass, for both energy and

chemical uses.

Biomass can potentially be used by the organic chemical industry in certain

circumstances as a renewable energy source. Non-renewable petroleum reserves

can then be extended for their more exclusive use as primary organic chemical

feedstock and the economic viability of capital intensive organic chemical

process facilities tied to petroleum feedstocks can be prolonged. The environmental and energy economics and concerns associated with this approach have been

discussed elsewhere and will not be discussed in this report (49, 55, 108).

In this report, biomass is considered in the context of an alternative

hydrocarbon feedstock for the production of commodity organic chemicals. In the

previous section of this report. Table 3.13 illustrated the many technical,

environmental and other considerations involved in determining the economic

viability of using biomass as a feedstock or substrate in the production of

organic chemicals. It should be emphasized that, even if successful approaches

are developed for converting biomass to organic chemicals (for example,

cellulosic materials to ethanol), industry must continuously weigh and compare

the return on investment (ROI) expected from such an approach versus alternative

approaches." These comparisons will include evaluation of ROIs from existing

processes, capital investment in existing processes, processes using alternative

feedstocks (for example, coal), and other projects or programs competing for a firm's limited resources.

51
.2 Supply

Biomass is composed primarily of lignocel lulose materials, and includes

forestry residues, agriculture residues, and municipal/agricultural/industrial

waste residues. Table 4.1 illustrates the forms of biomass in the U.S. and the

relative amounts produced per annum. As one proceeds from primary to secondary

to tertiary biomass, both the quantity produced per annum and the biomass

product value decrease. Currently there is large scale production in the U.S.

of ethanol from cereal crops (primary biomass) such as corn and wheat. However,

in the future, woody plants and underutilized secondary and tertiary biomass may

be the substrates of choice for fermentation processes for three major reasons.

First, neither the technology required for their full conversion nor their

economic potential has been fully realized (58). Second, use of crop plants for

chemicals may conflict with global food needs. Third, nature has, particularly

in woody plants, provided a mechanism for the abundant production of liqnocel lulose and for concentrating it. In addition and in contrast to other

products of photosyntheses (such as grass, algae, and crop plants), woody plants

can be collected on a year-round basis and can be stored for long periods of

time ( 109)

.

. 3 Biomass Chemical Components

Biomass, for example, 1 ignocel 1 ulose from woody plants, is complex and

consists of three major organic chemical components: cellulose, hemicel lulose,

and lignin (Table 4.2). Cel lulose (approximately 40-45 percent of the dry

weight of wood) is a high molecular weight, crystalline, relatively insoluble

polymer consisting of B-1-4, linked D-glucose (hexose; Cg) units. The

composition of hemicel lulose (about 20-35 percent of the dry weight of wood)

varies considerably with the type of biomass (for example, hardwoods as compared

to softwoods are relatively xylan rich), and consists of hexose (Cg) and

pentose (Cg) non-crystalline, relatively insoluble polymers, such as D-xylose,

L-arabinose, D-mannose, D-glucose, D-galactose, and D-glucuronic acid (110).

Table 4.3 provides information on the percentage composition of hexosans and

pentosans in primary and secondary biomass sources.

52
Table 4.1

Forms of Biomass in the United States

Annual Amount

Biomass substrate (million metric tons)

Primary biomass

Crops

720

Corn 195

Wheat 85

Hay 150

Soybeans 105

Wood 286

Secondary biomass

Crop residues 430

Cereal straw 141

Stover-cobs 145

Forest residues 260

Tertiary biomass

Feed lot manures 237

Municiple waste 150

Forest product waste 100

Industrial wastes 45

Sewage solids 15

Reference 58.

Table

Chemical Composition of

4.2

Some Biomass Feedstocks

Chemical Components {%)

Biomass Substrate/Feedstock Ce 1 1 u lose Hemicel lu lose Lignin

hardwoods 45 30 20

softwoods 42 27 28

wheat straw 30 27 11 corn stover 38 26 19

hemlock bark 26 16 15

feedlot manures 17 19 7 municiple solid waste 61 - NA

domestic sewage sludge (dry)

References 53, 58, 111, 112.

NA = Not Available

35 6 NA

53
Table 4.3

Composition of Hexosans, Pentosans and Other Biomass Components

Percentage Composition

Biomass Substrate/Feedstock Tfexosans Pentosans Lignin Pectin/^tarch

Primary

Woods

Softwoods

White Fir 58 9 28 - Douglas Fir 57 4 27 - Redwoods 44 6 34 - Hardwoods

Oak 45 20 25 - Sycamore 46 14 22 - Sweet gum 45 19 19 - Crops

Corn/Wheat 4 - 2 85

Secondary

Agriculture Residues

Cereal Straw

Wheat 39 19 14 - Rice 39 17 10 - Barley 40 19 14 - Corn Stover 36 16 15 - Corn Cobs 36 28 10 - Bagasse

Seed Hulls

41 20 20 —

Rice 36 15 14 - Oat 34 30 14 -

Sources: References 110, 113.

54
The third major chemical component, lignin (about 15-30 percent of the dry

weight of normal wood), is a high molecular weight, relatively insoluble, non- polysaccharide cross-lined polymer of sinapyl, coniferyl, and p-coumaryl

alcohols. Softwood lignins can be distinguished from hardwood lignins by the

predominence of guaiacyl units. These chemical units contain more reactive

sites and contribute to a greater degree of cross-linking (53). Thus, softwood

1 ignocel 1 ulose complexes are more difficult to degrade into individual chemical

components

.

.4 Technology and Chemicals from Woody Plants

The basic technology for the conversion of wood biomass to commodity organic

industrial chemicals is known (12, 57, 114). However, the economics for using

this technology and substrate as a feedstock have not, until recently, been

favorable because of the price and availability of competing petroleum feedstocks. In spite of this, some chemicals from wood or wood pulp are made in

large quantities (Table 4.4).

The forestry industry and pulp and paper manufacturers have been quite

cognizant of the importance of efficient use of forestry by-products on their

process economics, and have made substantial R&D investments in this area in the

past. This is illustrated byKoppers', Mead's, and Canada's MacLaren Power and

Paper's equity investments in Engenics and by Weyerhauser' s contractual

relationship with Cetus. These firms view biotechnology not only as an attractive alternative for improving the quality of forest stock more efficiently and

economically but also as a potential vehicle for improving forestry product and

by-product processing and as a vehicle for their becoming a greater market force

in the production of organic chemicals. Also Canada, in addition to its strong

petrochemical and natural resource base, has a vast lignocel lulose natural and

renewable resource base. Canada's biotechnology development plan has given high

priority for the application of biotechnology to cellulose and to forestry

residue waste utilization (115-117).*

*An example of Canadian application of biotechnology to forestry waste use is

the University of Waterloo/Envirocon process for converting pulp mill sludge

waste to protein or gasohol (Canadian Research, April 1982, p. 8).

55
Table 4.4

A. Organic Chemical By-products Formed in U.S. Pulping Processes

Kraft pulping, tons X 10^ Sulfite pulping, tons X 10^

Thiol ign in 8,800 Lignin sulfonic acid 1,920

Fatty and resin

acids

405 Hexoses 550

Formic acid 600 Pentoses 87

Acetic acid 840 Acetic acid 140

Acids derived

from

carbohydrates

4,500 Formaldehyde and

small amounts of

ethyl alcohol,

furfural, and

acetone

13

All other organics

(phenols, etc.)

650

Source: Reference 53.

B. Si Iv ichemical

s

of Commerce

Cel lulose

Cel lulose esters

Cellulose ethers

Rayons

Wood resin

Turpentine

Pine oil

Tall oil

Tall oil resin

Tall oil fatty acids

Dimethyl sulfide

Dimethyl sulfoxide

Kraft lignin

Lignin sulfonates

Ethyl alcohol

Vanillin

Yeast

Charcoal

Bark products

Hemicel lulose extracts

Source: Reference 57.

56
In addition to an improved economic climate for use of wood biomass as a feedstock in the production of organic chemicals, improvements are also needed in

basic process conversion technologies and conditions. Selective adaptation of

these technologies and conditions to a variety of wood biomass substrates are needed to achieve optimum separation and subsequent conversion of major chemical

components. Table 4,5 illustrates the variety of industrially useful chemicals

and products that potentially can be derived from wood biomass. Using current R

& D activity as an indication of technological and commercial developments, a near-term commodity organic chemical target of opportunity will be ethanol via

(a) hydrolysis and saccharification of 1 ignocel 1 ul ose and (b) fermentation of

C

5 and Cg Sugars. In the longer-term, a better understanding of the complex

structure of lignin and improvements in process technologies associated with its

modification offer the potential for providing an important and renewable source

of monoaromatic chemicals and higher quality and value added products such as surface coatings, adhesives and foams.

.5 Waste Residues

Wastes from industry, agriculture and mun ici pa.l i ties have been defined

broadly as resources misplaced in time, location or need (118). Characteristically, these waste residues have a high cellulose content, are generally of

low commercial value, and, in many instances, are major sources of pollution.

There appears to be renewed interest in the possibilities of applying biotechnologies for upgrading these residues to more valuable organic chemicals. For

example, the economics are becoming favorable for the anaerobic digestion of

agriculture or sewage waste residues or organic solid waste residues from

landfills to methane (119-124), and for the industrial use of an immobilized

enzyme (lactose) to convert dairy industry waste (whey) to commercially useful

syrups and protein concentrates (125).

Based upon experiences in the pharmaceutical industry, fermentation

by-products are a major wastewater source (118). The dairy food processing

industry is also a major contributor to biodegradable waste (126). With

projections for expanded industrial use of fermentation process technologies for

57
-

Gas

(carbon

monoxide,

carbon

dioxide,

hydrogen,

hydrocarbons)

—

methanol

SCP

PYROLYSIS

-

Liquids

(methanol,

acetic

acid,

acetone,

phenol

derivatives)

•

formaldehyde

-

Charcoal

1/1 QJ>

(O>

s-O) -a

<u

<13 X

<D

r— CD o CD o i4-

>1

CTi

O) _E

c

1 OJ <13 >1 >1

Cl cl ! 1/1 O O O CD

s- s_

•

1- >

CL Q. s- -I- O

1/1 >i+J

a;

»> O) <—>1^

^ <D

3 -O

-a

<d

O) >1 <u — o

cn <o

4-J <Do

<13

</1 O

o

o

1/1 oi o =

>1 o

o

a_

i/1 +j "O 1/1 •r- <13 u o) c >-

J

</i o

C I— O O ^ >)

s- C_) <au -a o c

s- <o -o

>1 1/1

<T3

<oo

</) OJ>

o<13 </1 o>

c

3>

O)

<o

s_3

MS_3

>1 JZ o

c

<d >1X

u •<- O!

•1- O)

4-1 4-! ^ ID CD u o X 1— 1/1

<13 S_ O o o

r— (2. S_ >1 o

-a 1— 3

O)

<13 S- ^ o

x: Q_ C3

U1

<13 D

§1

O)

5-<D

C

o

c •<- O• f— <o o i/1 4-> N1 I— •>— <o •>—

-t-j i/) c >—

<13 a: I

i_ I— CD <13 T3 O O -t-> >1 S- S_ iD JC T3 T3 >1

<U >1 >> S- O X X o

o

Q_

>1 X

1/1 “O c fO o<~1 1/1 i_ <— i/1

<13 O cr U X c

o o •I— S_ <D 4-> T3 +-> <13 iD

<13 O S- X O O <D

s=>1

i/1 i/1 . • r—

CD CD iD s- o

> > I— o Q.

•I— -r- O C4- +J 4-> C CD

<13 <0 CD i/1 1—

•1—> > X CD X •!— Q.4-> CD 3

s- i- <13 CD 1—

<D CD •> > O

"O TD c •o •1— (D

•r^ CD i/I ID 1 1— r~” r~” E I— s_ ID O S- o o .— S- <D <D E 1— CD c c •>- CD 3 X 1— CL4-J

CD CD C -t-J <4- •r— •!— X <13 X X <13 C <4- M- CD 3

Q. Q_ > 1—

1

<13

J—L_t 1

i-<D U_

L.

O -•-> C •!— <13 O M

•f- <0 -r- c —

<13 O) .

S_ CD <13 "O O -U >) i- ID X -O >1

O) >1 S- 0^0 J L

c

o

c

o

<13 O

' O

c

</) O)

<13 X

i/1 a)

i/1 o

X

OJ

i/1 O)

i/1 o

-I

i/1 <0 C

C iD o

O) >>•!-

oil 4-> 0 o <13 S- S- T3

-O "O •<- >1 >> X X X o

1

1 L

O) o

I

OI

OJ

OJ

i/1 o

O)o

<13 u

i-O)Q.

<o QOJ Xo

Sources:

References

53,

108,

109.
feedstock conversions and waste residue utilizations, development of new, less

empirical, and more sophisticated bioprocess approaches to the conversion or disposal of waste residues will be essential from both an environmental and

economic perspective. From an economic perspective, bioprocess economics will

become more favorable with (a) improved approaches to recycling chemical

by-products from bioprocesses, (b) production of commercially useful chemical

products from biomass waste residues, and (c) an expanded market for feed

supplements and nutritional components derived from biomass waste residues,

.6 Chemicals from Plants

One possible alternative to synthesizing chemicals from biomass is to obtain

industrially useful chemicals from plants directly. Currently the^f'e is global

interest in finding chemical derivatives from both aquatic and terrestrial

pi ants

.

One approach being investigated is the development of aquaculture management

technologies for the production of algal biomass (127). In the past, considerable emphasis was placed on the high nutritional value of algal biomass, with

protein up to 70% of the dry weight in Spirul ina species, a blue-green algae

(128). Commercial application of algal cultures for food production have

generally not been successful because of high capital costs, poor yields,

product quality control problems, and lack of market outlets. Use of large

scale algal ponds for liquid waste treatment have been more successful (129).

There is much current interest in the use of ponds and coastal waters for the

cultivation of algal biomass as an energy source, for example, methane biogas

(127, 129, 130).

The global harvest of marine algae is illustrated in Table 4.6. Major

commercial products of marine algae processing are shown in Table 4.7: polysaccharide polymers agar, algin, and carrageenan (108). With increasing

industrial interest in biotechnology and in immobilized biocatalytic processing

systems, both alginate and carrageenan are receiving consideration as polymer

support and entrapment matrices for biocatalysts (131-133).

59
Table 4.6

Global Harvest of Marine Alge

(metric tons)

Year

Brown

Algae

Red

Algae

Green

Algae

1971 483,600 375,500 1,100

1972 506,500 322,800 700

1973 586,900 450,300 900

1974 696,567 525,654 2,237

1975 633,182 422,424 2,487

Source: Reference 108.

Table 4.7

Sources and Uses of Algal Products

Algal Algal

Product Source Uses/Properties Industry

Agar red suspending, thickening, stabilizing

solutions

bakery, confectionary

Algin brown gelling, thickening, suspending,

emulsifying, and water-holding

properties

paper and pulp,

pharmaceutical

,

food processing

Carrageenan red thickening, stabilizing, gelling

agent; texturing binder; gel base

to control release of substances

food, health

care

Source: Reference 108.

60
In addition to possible use of algal biomass as a source of vitamins and as

an organic fertilizer, algal biomass with high concentrations of polysacchar ide can be easily converted via anaerobic fermentation to produce organic acids

which can subseguently be converted to olefins, acetic acid, and liguid hydrocarbon fuels (134), Cither research advances in this area include reports of the

economically competitive production of glycerol from algae; of future prospects

for production of plant growth hoi^mones, pharmaceuticals and specialty chemicals; of the use of immobilized algal cells for specific bioconversions; of the

use of carrageenan or other saccharides in vaccine detoxification; and of the

possibilities for transforming algae via DNA from other species (135-139).

Another area of active research interest involves hydrocarbon-producing

crops as sources of biologically synthesized organic chemicals. Species from

Euphorbi aceae and Asclepiaeceae plant families have received much attention

because of their possible cultivation on "petroleum plantations" as energy

sources and as sources of latex-i soprenoid hydrocarbons (140, 141). Oilseed

plants (sunflower, linseed, rapeseed, buffalo gourd, Chinese tallow tree) can potentially provide a renewable, domestic resource of natural glycerin and of

fatty acids of varying chain length containing epoxy, hydroxy, and keto, as well

as othe*^ functional groups, in addition to being a possible source of the

nutritionally important amino acids lysine, cystine, and methionine (49,

142-145). The chemical constituency of peat also indicates some prospects for

its alternate use as a natural resource chemical feedstock. Peat biomass

extraction processes can yield a variety of potentially useful chemicals such as peat wax, asphalts, peat coke, alkaloids, tannins, resins, fulvic acid, humic

acid, carbohydrates , amino acids, and B-vitamins (146). To improve the

commercial prospects of these and other domestic agriculture-based, biomass

alternative chemical feedstocks, however, a number of technological, economic,

and environmental barriers need to be overcome, for example:

0 selected plants need to provide high yields per acre on marginal lands in a particular climate or to serve as an acceptable alternative cash crops in

agricultural crop rotation management schemes;

61
0 chemicals produced from such plants must have a higher value than other

plant product uses such as fuel; and

0 efficient and cost sensitive extraction technologies need to be developed

for separating valuable chemicals from other plant constituents.

In addition, natural fats and oils from domestic renewable resources will

have to compete in the future with similar imports of these natural products

from Southeast Asia and the Philippines coconut and palm oil plantings, as well

as tallow from domestic sources (145).

These considerations also apply to the production of ethanol from high

carbohydrate content root crops, such as the Jerusalem artichoke, cassava, yams

and fodder beets (48, 147-149).

On a smaller scale, plant cell culture biotechnology including the use of

immobilized plant cells has the potential to produce a number of specialty, low

volume, high value added chemicals that can impact on a number of industries

(Table 4.8; 150-153). Technologies such as plant and protoplast cell culture,

protoplast fusion/somatic hybridization, protoplast-liposome interactions, and

genetic engineering, used individually or in combination with conventional plant

breeding technologies, offer the promise in the long-term of revolutionizing the

production of biomass crops with desired characteristics, such as:

0 genetically uniform and predictable character! sties;

0 tolerance to adverse environments; and

0 ability to produce high concentrations of biologically synthesized chemicals or to produce plant constituents tailored to chemical feedstock needs.

In the interim, considerable R&D is required to realize this promise (151,

154, 155).

62
Table 4.8

Potential Range of Chemical Products from Plant Cell Cultures

Alkaloids

Allergens

Amino acids

Anthraquinones

Antileukemic agents

Antimicrobial agents

Antitumour agents

Antiviral agents

Aromas

Benzoquinones and related compounds

Carbohydrates

Cardiac glycosides

Chalcones

Dianthrones

Enzymes

Enzyme inhibitors

Flavanoids, flavones

Flavours (including sweeteners)

Furanocoumarins

Hormones

Insecticides

Latex

Lipids

Napthoquinones

Nucleic acids

Nucleotides

Oils

Opiates

Organic acids

Protei ns Peptides

Perfumes

Phenols

Pigments

Plant growth regulators

Polysaccharides and derivatives

Steroids and derivatives

Sugars

Tannins

Terpenes

Vi tarn i ns and Terpenoids

Source; Reference 151.

63
.7 Barriers to Research and Commercial Progress

Impediments to biomass research progress and to commercialization successes

using biomass ( 1 ignocel 1 ulose) as a substrate that are relevant to NBS's mission

include: (a) lack of standard biomass samples for comparison of experimental

(domestic and international) findings, (b) lack of standardized and reproducible

biomass ( 1 ignocel 1 ulose) assay methods, and (c) in general, lack of information

for characterizing 1 ignocel lulose and its major chemical components (156).

These barriers and possible approaches for overcoming them will be explored in

more detail in subsequent sections of this report.

64
5. CONVERSION OF CELLULOSIC BIOMASS TO ETHANOL

.1 Overview and General Process and Economic Considerations

The process of converting lignocel lulose biomass to ethanol via hydrolysis

and fermentation technologies was selected for more in-depth discussion in this

report because it serves as an example of an area: (a) where R&D interest is

intense and industrial and economic potential is high, (b) where solutions to

scale-up and commercialization problems and barriers may have generic applicability to production of other organic chemicals using different substrates and or bioprocesses, and (c) where NBS can possibly provide a range of measurement,

information and other infrastructure services to facilitate commercial

applications in biotechnology.

The desire to replace petrochemicals with biomass (such as lignocel lulose)

as a feedstock or substrate for the industrial production of organic chemicals

such as ethanol has been stimulated by fluctuations in price and availability of

petroleum feedstocks over the past decade, and by projections on the amounts of

under-utilized lignocel lulose that may be available as a substrate for

bioconversions

.

The primary constituents of lignocel lulose biomass are cellulose, hemicellulose, and lignin (refer to Section 4). The major physicochemical

characteristic of 1 ignocel lulose substrate that limits its direct fementation to

ethanol is its complex and polymeric nature (157). Hexose and pentose polymers

of cellulose and hemicel 1 ulose by themselves can be enzymatically converted to

organic chemicals, but their embedment in the lignin matrix inhibits the

activity of microbial cellulases by steric hindrance. In addition, the presence

of lignin in certain concentrations may be toxic to microrgani sms . Thus,

chemical, mechanical, and/or enzymatic hydrolysis pretreatment of 1 ignocel lulose

is a prerequisite for the conversion to simpler sugars.

Depending on the nutritional environment, the atmospheric conditions of the

reaction vessel, and the microogranism( s) selected, these sugars can, following

pretreatment, be converted by fermentation to a number of high volume, low

margin acids, alcohols, and gases that could serve as organic chemical feed65
stocks (58, 113, 158; Table 5.1). For example, lactic acid produced in this

manner has applications in both organic chemical commodity and speciality

markets as a component of new plastic materials and of controlled release

matrices for pharmaceutical agents and pesticides (159). In addition, the large

scale of fermentations required for commodity organic chemical production will

produce process reaction by-products that will necessitate the development of

efficient recovery processes and of market outlets to improve process economics

(e.g., enzymes; vitamins; substrates for methane production; residues for

fertilizers or energy sources; and protein concentrates). Improvements in

process economics will also depend upon developments in continuous industrial

processes including technologies for process water, nutrient medium, and cell

recycle ( 160)

.

Tab le 5.

I

Products of Fermentation and Potential Organic Chemical Feedstocks

Organic acids Alcohol s-ketones Esters

Propi onic Ethanol Ethy 1 acetate

Acet ic Acetone Ethy 1 butyrate

Caproi c Butanol Pol y-3-hy dr cKbutyrate

Lactic 1 sopropanol

S ucci n ic Gl ycerol

Butyr ic Methanol

Formic 2, 3-butanediol

Aery 1 i c

Sources: References 58 and 158.

Gases

2CHCO

4

2

Thus, the basic steps in the conversion of 1

i gnocel lulose biomass to

ethanol can be represented as follows:

0 feedstock collection, handling and pretreatment for biomass size reduction

and for dissociation of lignin and cellulose;

0 saccharification of more complex sugars to simple sugars using the

cellulase system;

0 fermentation of simple sugars (Cg and Cg) to ethanol; and

0 ethanol separation and recovery.

66
A simplistic model of the bioconversion of cellulose to ethanol is

illustrated below:

Cel 1 ulose

endoglucanases

exogl ucanases Oligosaccharides /3-gluccsidase

^ /pv

I

1 ' ' ' * _

(

I L*— Cellobiose !*

yeast

Glucose Ethanol

—

1

*Indicates product inhibition

There is much in the historical literature on the technologies associated

with bioconversion of cellulosic materials to ethanol. However, improvements in

the industrial economics of this and other fermentation processes in comparison

with chemical synthetic processes will occur only with significant advances or improvements in many areas such as:

0 handling large amounts of fermentable materials:

0 direct enzymatic conversion of cellulosic materials;

0 production of cellulase and other process biocatalysts;

0 product tolerance and higher substrate and product concentrations;

0 product separation and recovery from reaction by-products;

0 productivity of fermentation reactions;

0 understanding interactions between various unit processes and an overall

process; and

0 closer integration of the biological system with process steps (161, 162).

Several of these areas will depend upon improvements in bioprocess

engineering technology. Strategies for improvements in this technology,

referred to as process intensification, are concerned with:

0 increasing the volumetric rates of reaction;

0 improving yield coefficient ratios;

0 increasing the number of reactions in a single reaction vessel;

0 reducing the number of reaction and recovery steps;

0 reducing the number of separate, dedicated reaction vessels; and

0 improving recycle engineering technology and reactor design (160, 163).

Table 5.2 identifies several areas where technological innovations could

lead to significant process intensification and improved process economics.

67
s na ta

(C

<D

ClO

•A

(T3 k &

T5OUCc _ UJ -O—

X

(/) <]) <D — o — 0) —

(D *— (0

I/) Q. ^ 3 O

T33 U

(/) tn©oou

Q. ^ 5 S 15 S

-O E -5

0) 3 _ ir c >

T3

u3 C T3 O m O

f- S h, ^ 2 S 3

O'

a 8 ©

u

Q.E O iU

—

— Q.

w

-o©

"O©

©o © © N4 q: u

© o — > u c © 4- 0 0 Q © ©

•—

O

g U) 4“

L. 3 ©

U 0 u © —c O © © > u U

>-©>o 1

cnc c si/) E©

4-©

+- 0oU) c©

T3c

Q. E1.©

©

O.©

u. > © 4-

C (/)

© O) 4- 0 © © © © Q. 3 © U) 3

S’

u

CLE

1.

(Jc

*o©

L. oco 1 ©

“Oc 0 "O©

L. *oc u CL o CD — 4- CL © 8u

CL£ ©

L.Q.E

(/>

® §

U)c +-

® s © u - I — c ©

JZ

a. —

© 0)

u *o S- o c 0 c o o — © ©

•o 0 4- 4- u 4- C © © © >. © E

>•

u 3 c o 0 C

*o

©

L 0©

(X

00

vO

dilute,

suspension

free

aqueous

properties,

particularly

viscosity
It should be noted, however, that the major component influencing any shift

to biomass based processes for the production of organic chemicals will be the

price of the biomass raw material relative to the price of petroleum or natural

gas hydrocarbon feedstocks. This is illustrated in Table 5.3 by categorizing feedstock.* the costs of ethanol production using molasses as the hydrocarbon *

Product!

Table 5.3

on Cost Categories in the Bioconversion of Molasses to Ethanol

Cost Category

raw material (molasses)

fixed costs including labor

electricity + steam

materi als

water

% of total production cost

68%

14%

10%

5%

3%

Source: Reference 158.

Although the costs of both petrochemical and renewable biomass will

increase in the future, the rate of increase for biomass will probably be less

in the long term. _ When the costs of these two feedstocks begin to intersect,

greater emphasis will be placed on seeking technological innovations to improve

the efficiency of the process technology. Process efficiences will be a major

factor in determining what bioengineering processess will be used and when it

will be economical to make them operational in industrial biotechnological

processes

.

.2 Pretreatment of Cellulosic Biomass

In order to enhance the conversion of lignocel lulose raw material to simple

sugars and chemicals, active research and technology development on a variety of

chemical, physical and mechanical processes are in progress. Table 5.4

illustrates a number of pretreatment process approaches (used individually or in

combination) that are attempting to expose the molecular bonds of major

1 ignocel lulose constitutents (cellulose, hemicel 1 ulose, and lignin) to enzymatic

*Although molasses is a feedstock for ethanol production, the economics of its

conversion are marginal. Hydrolyzed starch and cellulosic materials are more likely biomass feedstock candidates for the production of organic chemicals.

69
Table 5.4

Chemical and Physical Li giocel I ul ose Pretreatment Approaches

Pretreatment

L i gnocel I ul os ic Feedstock (substrate) Chemical Meehan ica I /Phy sica

I

Process Reference

saw dust grinding and heating Gul f -

U. of Arkansas

111

crop residues o

o

Sul fur ic acid

(95-120*0

2-A% (90*0

( hem i cel 1 ulose

hydrol ysi s)

cadoxin solvent

( 1 i gn i n & cel 1 ul ose

separation)

T sao 157

wood ch i p s steam explosion:

24 0-300 *C 500-1000 psi

lotech 164

saw dust sulfuric acid

1-1. 5$

o extruder screw

o 230*C, 500 psi

NYU 164

wood chips,

pulp mill wastes

wood chips

ethanol/water (200“C)

- lignin:

ethanol phase

- hem icel 1 ul ose: water phase

- cel 1 ul ose:

und i sso 1 ved o so 1 vent

hammer mill General Electric 165, 166

del i gn i fi cat ion o d i 1 ute acid, low

temperature

prehydrol ysi s o di 1 ute acid, high

temperatire hydrol ysi

steam explosion

s

Georgia Tech - Stake Technology

165

wood logs o hot water

o 0,6% sul f ur ic acid (140-180*0

wood debarking Inventa 167

agriculture

cel 1 ul osic waste material

sodium hydroxide

de 1 i gn i f i cat i on

Trivedi and Rao 168

wood ch i p s hydrogen fluoride wood chip drying Michigan State

Uh i versi ty

165, 166,

169, 170 wood pulp for papermaki ng

sul fur ic acid

0.1-1$ (190*0

gamma irradiation David et al. 171

aspen wood chips 2$- 10$ ch 1 or i te steam explosion

250*C, 560 psi

Sadd ler et al

.

*

a gr i c u 1 1 ur e/ forest residues

phenol (100*0 Battel le-Geneva »»

Biofechnol. & Bioeng. 1389-1402, 1982.

**Chemlcal & Eng. News, May 31, 1982, p. 38.

70
have progressed on pretreatment alternatives, a number of problems have become

evident, for example, loss of carbohydrate material, corrosion of equipment,

undersirable side reactions, and high capital costs.

Some advantages and disadvantages of alternate p-^etreatment approaches are summarized in Table 5.5. If pretreatment of cellulosic material will be

required in the future, and if pretreatment methods are to be cost-effective,

many research and development hurdles must be overcome. For example, there is a critical need to identify and to select optimum conditions and pretreatments for

cellulosic materials from specific sources and to relate these choices to

process economic, efficiency, energy use, and scale-up considerations. Ideally, one would like to circumvent these problems by developing biological systems

(microbial and/or enzymatic) that would allow the direct conversion of cellulosic materials to hexoses and pentoses and then to ethanol. This approach

would avoid or at least reduce the need for the currently used and proposed

capital intensive, destructive, and relatively inefficient pretreatment

processes. The physical approaches are particularly costly and time consuming.

.3 Biological Conversion

For the efficient and economical biological conversion of renewable

1 ignocel lulosic material, extracel 1 ul ar enzymes from microorgan i sms capable of

degrading cellulosic biopolymers to mono- and di saccharides will need to assume

an increasing importance in biotechnological processes. Before economically

feasible process technologies are developed, several complicated and interrelated problems need to be addressed, for example:

0 thermo- and chemical stability of enzymes;

0 enzyme(s) production variables;

0 multiplicity of enzymes and substrates complicating enzyne assays;

0 optimization of activity and rates of reaction of individual enzymes in a multi-enzyme system;

0 multiplicity in terms of origin of 1 ignocel lulose substrates and varying

degrees of reactivity of such substrates to enzymatic hydrolysis;

0 low rates of hydrolysis of lignocel lulosic materials;

71
Table 5.5

Approaches to Pretreatment

to Enhance Enzyme

of Cel I ul osi Susceptibi I i

T~ype Advantages

Phy sica

I

Hammermi 1 1 Co 1 1 o i d mill

Bal 1 mi 1 1 Two-rol 1 mill

Less Costly

Improves hand 1 i ng

E f feet i ve High bulk density

Steam explosion

220-240“C,

600 Ib/in.^

E f feet i ve Less costly

Alkali swel 1 i ng NaOH Moderately effective

Less costly

Pe 1 i gn i fi cat ion

Chemical pulping

Biological

Moderately effective

Conserve enzyme and

reactor space

Pissolve, reprecipitate

Cadoxen (toxic)

Hydrazine (toxic)

Ferric tartrate E f feet i ve MaOH

Cuprammon i urn V i scose Concentrated acid

Source; Reference 172

c Mater i a I

s

ty

Pi sadvantages

I nef feet i ve

Energy- i ntens i ve Expensive

Low buik density

Ineffective for softwoods

Low bulk density

Must wash or neutralize

Ex pens i ve Low bulk density

Pol I ut ion

Low bulk density

Expensi ve

Pi f ficult hand I i ng

Solvent recovery

S i de react ions

Pollution, lignin degradation

72
0 changing patterns of substrate susceptibility to enzymatic hydrolysis with

time;

0 nature of substrate handling and pretreatment;

0 product inhibition of enzyme activity;

0 enzyme inactivation by heat, pH, mineral ions, proteases, etc.;

0 identification of microorganisms that can enzymatically attack specific

1 ignocel lulose substrates such as wood, bagasse, straw, etc.;

0 molecular level understanding of the metabolic pathways and the efficiency

and stability of these pathways used in the decomposition of cellulosic

materials; and

0 identification of important process enzymes and approaches to regulating

their activity at the gene and transcription levels and to increasing their

productivity in biotechnological processes.

The following sections provide a description of some areas of active R&D in

the bioconversion of 1 ignocel lulose residues to ethanol. These sections also

provide a sampling of the many problems that still need to be addressed before

economically feasible commercial products can be developed.

.a Cellulose Degradation . Considerable research work on the enzymatic

hydrolysis of cellulose has been done with two fungi, namely, the white-rot

fungi _S. pulverulentum and Trichoderma reesei (173). The three primary

hydrolytic enzymes of the cellulase complex system of these microbes are:

(a) endo-^-1, 4-glucanases (act randomly on cellulose);

(b) exo-1, 4-<3-gl ucanase (splits off cellobiose or glucose units from the

nonreducing end of the cellulose); and

(c) /3-glucosidase or cellobiase (hydrolyzes cellobiose and cel lodextr ins to

glucose and cellobionic acid to glucose and gl ucono 1 actone; 173).

A recent review by Ryu and Mandels summarizes research progress with the

cellulase system and the need for research progress in the enzymatic hydrolysis

of cellulose in priority areas such as;

73
(a) substrate characteristics, properties and multiplicity;

(b) multi-enzyme system kinetics, characterization and interactions;

(c) enzyme purification and characterization; and

(d) enzyme inactivation and process design improvements (172).

The sequential and/or simultaneous mode of action of the cellulase system on cellulose is illustrated below;

Endo-/3-l, 4-glucanase

.b Hemicel 1 ulose Conversion . In addition to dilute acid treatment, a variety of endo-mannanases , -mannosidases, and -xylanases exist that can release

Cg and Cg monomeric sugars from the water soluble dextrin, hemicel lulose

.

These enzymes are also produced by microorganisms ( Trichoderma , Asperqi 1 1 us , Pen i c i 1 1 urn ) that produce cellulases. A major problem in comparing and in

extrapolating from experimental findings where hemicel lulose is used as a substrate is that the variety of hemicel lulose conversion approaches being used

also produce a variety of reaction products.

74
.c Lignin Conversion . In several of the R&D, pilot plant phase and

commercial processes used in or proposed for in the conversion of lignocellulose to chemical products, lignin has been separated from the cellulose and

hemicel 1 ulose components and later used as an energy source to improve process

economics. Less is known about the opportunities for bioconversion of lignin.

lAihite-rot fungi appear to be one of few microbial species capable of enzymatically degrading the lignin component of lignocel lulose through phenol

oxidases. Metabolic products of this catalytic reaction include vanillic acid

and quinones (173). Recently, research findings on the enzymatic activity of

the fungus Cyathus stercoreus in the breakdown of lignin also appear promising

(174). Additional research on physicochemical aspects and kinetics of

del ignif ication on a molecular level also appear warranted (175).

It would appear that a systematic review and evaluation of the enzymatic

activities, properties and pathways of other wood degrading fungi may reveal

microorganisms and microbial enzyme systems that can more efficiently hyd*^olyze

1 ignocel 1 ulose chemical polymers to simpler monomers under temperatures required

for higher reaction rates necessary for industrial processes.

.d Saccharification and Fermentation: Thermostability . Two major

problems encountered in the conversion of cellulase polymers to simple monomers

are the thermostability of cellulase enzymes and low hydrolysis rates of cellulosic materials. Thus, a significant process parameter in saccharifications is

temperature. With increasing operating process temperatures, the rate of

enzymatic activity may increase 2-5 fold until the enzymes are thermally

inactivated. However, at these higher temperatures, the potential for growth of

culture contaminants also decreases.

Microbial enzymes show considerable variability regarding their thermostability. The activity of the enzymes of thermophilic (45°C+) microorgan i sms

is more stable then those from mesophilic (37°C-45°C) microorganisms (176).

Table 5.6 gives some examples of the kinds of R&O activity in progress to

improve the thermostability and the reaction rates of the cellulase process

system.

75
Table 5.6

R&O to Improve Thermostability and Reaction Rates

Optimum Temperature Reaction

Microbial Strain(s) Range (experimental) Reference

Trichoderma reesei 45-50°C 111, 157

Sporotrichum thermophile 55-60°C 177

Thermomonospora sp. 55-60°C 176, 178

Thermomonospora sp. + Clostridium thermocellum

60-65“C 165, 166

Cellulomonas sp. +

T. reesei

37-45°C 179

Fusarium sp. 50°C 180

Thermobacteroides

saccharalyticum 55°C 181

An alternative to the above and to using the popular cellulolytic fungus,

T. reesei , is to isolate cellulase genes from other themostable, cellulolytic

fungi and to clone these genes into yeasts. Such an approach is being pursued

at Stanford Research International with Thiel avia terrestris , a more heat

resistant fungus with a higher temperature optimum (70°C) than T. reesei (182,

183)

.

A related approach is being pursued at USDA where the cellulolytic fungus

Penicillium funiculosum is viewed as an alternative, less complex source of

genes than T. reesei for a cellulase system (184). Attempts will be made to

isolate the three cellulase genes from P. funiculosum and to transfer them to a microorganism that could participate in combined saccharif ication-fermentation

.

In addition to genetic manipulation approaches for improving process

thermostability and reaction rates, recent experience with the microorgan ism

Caldariella acidophilia , a thermophile with a growth temperature optimium of

87°C, suggests the possibility of using polyurethane foams to immobilize whole,

thermophilic microbial cells for industrial biocatalytic reactions (185).

76
.e Product Inhibition: Saccharification . Saccharification end products

include not only glucose and cellobiose but also other sugars such as xylose.

Cellulases are inhibited competitively by both glucose and cellobiose and the

extent of inhibition increases with increasing resistance of cellulose to

hydrolysis (refer to figure in Section 5.1; 172). Cellobiase enzyme is also

inhibited by glucose. Refer to the system model illustrated in Section 5.1 for

additional details.

Since glucose does not accumulate to any great extent in a saccharif iciation and fermentation process, cellobiase concentration and activity have

important influences on an overall cellulase system activity. Traditional

mutation and selection techniques to improve the cellobiase levels in T. reesei

have not been too successful to date. However, cultures of Aspergi 1 1 us phoenicis have shown promise as additive sources of cellobiase to J. reesi

enzyme levels (172). Also, some yeast strains can convert both cellobiose and

xylose ( 186)

.

.f Product Inhibition: Ethanol . With reference to the model system

illustrated in Section 5.1, conventional fermentation processes or combined

saccharification-fermentation processes for the production of ethanol are limited by the inhibiting effect of ethanol. This inhibition decreases both

rate of ethanol production and the volume of microbial cell mass when the final

ethanol concentration reaches 10-15 percent V/V. In order to reduce or eliminate ethanol inhibition, ethanol must be removed from the fermentation

broth as it is being formed and/or fermenting microbial strains more resistant

to ethanol must be developed.

Traditional distillation approaches are too energy intensive for practical

consideration in future industrial biotechnological process engineering

approaches. Biochemical/microbiological solutions to the problem of ethanol

tolerance in yeast may be complicated by the relationship between ethanol

tolerance and altered lipid composition in yeast plasma membranes (162).

Further research is needed in this area.

A number of alternative technical approaches are currently being pursued in

order to reduce ethanol and product inhibition in bioconversions. Some of these

77
approaches include vacuum extraction, cell floculation, two-phase

bioconversions, immobilization of microorganisms, etc. (Table 5.7).

Continued efforts are also needed to establish the kinetic relationships of

growth, substrate use, and both sugar and ethanol formation during saccharification and fermentation in order to better understand process variables and to

improve/optimize process design and control.

.g Conversion of Pentose Sugars to Ethanol . The technology and industrial

process parameters for the conversion of glucose (Cg) to ethanol using

Saccharomyces cerevisiae yeast strains and the bacterium Zymomonas mobi 1 is are generally well understood. Yeasts, however, are generally capable of

metabolizing only a small range of carbohydrates to ethanol.

Rosenberg has recently reviewed research on the conversion of pentose

sugars to ethanol (110). For improving the process economics of converting

biomass residues to ethanol, conversion of xylose (Cg), which makes up about

one-third of the potentially fermentable sugars in cellulosic biomass, is

essential. Although some recent progress has been noted, additional research

efforts are needed (174, 204).

Identification of a yeast that can simultaneously convert Cg and Cg

sugars and/or can metabol ical ly work in concert with other microbial strains

would be prefered because of the wealth of industrial experience with yeast, as well as the rapidly accumulating knowledge base about yeast genetics and the

application of rONA techniques to yeasts. Such an approach is being pursued at

USDA to transfer into S. cerevisiae the two genes of P. tannophilus required for

producing the enzymes used in converting xylose (Cg) to xylitol and then from

xylitol to ethanol. S. cerevisiae could then potentially convert both Cg and

Cg sugars in the fermentation process step (184). Another approach is to

transfer into yeast the one gene from E. coli that expresses the enzyme xylose

isomerase. This enzyme converts xylose to a ketone sugar that can be fermented

by S. cerevisiae ( 184)

.

Table 5.8 illustrates some of the current research activity on the

conversion of Cg sugars/xylose to ethanol.

78
Table 5.7

Examples of Technical Approaches to Reducing Ethanol/Product Inhibitions

Approach Organ i sm Reference

Mixed culture (different

substrate- product

inhibition characteristics)

S. cerevisiae NCfC 87

+ NRRL-Y-11, 572 (sake yeast)

187

Aqueous two-phase

bioconversion

T. reesei

+

S. cerevisiae

188, 189

Continuous vacuum extraction

and cell recycle

Zymomonas mobilis 190, 191

Ultrafiltration (capillary

cross flow microfiltration)

Z. mobilis 192

Multistream ethanol feed in

multistage tower fermenter

Candida util i

s

193

Fermentation coupled to

liquid-liquid extraction

S. cerevisiae

( immoDi 1 i zed cel Is)

194

Settler —cell floculation

and recycle

Z. mobilis 195

Drying ethanol using organic

cellulosic residues — 196

Thermostable enzymes from

thermophilic organisms

Bacillis stearothermophi 1 i

s

197

Immobil i zation

0 calcium alginate

0 calcium alginate

0 ion exchange resins

0 boros i 1 icate-gl ass fibers;

polystyrene beads

0 coentrapment of Sepharoseimmobilized ^-gl ucos idase in

calcium alginate (+yeast)

spheres

rDNA (engineer microorganism with — 174

rapid enzymatic activity + microorganism resistant to ethanol

toxicity)

S. cerevisiae 131, 198, 199

1 . mobi 1 1 s 200

S. cerevisiae 201

Z . mobi

1

1 s 202, 203

Talaromyces emersonii

*Biotechnol. & Bioeng. 24: 1461-1463, 1982.

79
Table 5.8

Approaches to Improved Conversion of Sugars/Xylose to Ethanol

Substrate Organi sm Culture Environment Reference

wood chip hemi- Candida

cellulose hydro- sp. XF 217

lysate

aerobic/ anaerobic 205, 206

wheat straw hemicel 1 ulosePachysolen

tannophil is

aerobic 204, 207,

208

xylose

D-xyl ulose

Candida tropical i

s

aerobic

Schizosaccharomyces pombe aerobic/anaerobic

209

210

.h Simultaneous Sacchar if i cat ion -Ferment at ion . To improve bioconversion

process economics by reducing capital costs and production costs of ethanol, a number of technological approaches are in progress to achieve process intensification. One approach involves either simultaneous saccha.rif icationfermentation or the combining of saccharif ici ation and fermentation process

steps. In this biological system both cellulose and hemicel lulose would be

hydrolyzed to Cg and Cg monomers followed by the direct conversion of these

sugars to ethanol. Since ethanol has less inhibitory effect on the hydrolysis

of cellulose than cellobiose, the saccharification process would be improved.

However, since there are different optimum reaction temperatures for enzymes in

the two processes and different microorgan ism are required for Cg and Cg

conversion, a coupled system may be difficult to optimize (172).

In spite of the limitations. Table 5.9 gives examples of attempts to couple

these two bioconversion process steps.

80
Table 5.9

Technical Approaches to Simultaneous Saccharification and Fermentation

Substrate Saccharification + Fermentation Reference

saw dust T. reesei S. cerevisiae,

S. car Isbergenis,

Candida brassicae*

111

cel 1 ulosic Clostridium thermocellum Z. mobi 1 is* 211

materi al

s

Acetivibrio cellulyticus

wood chips. Themomonospora sp. Clostridium thermocellum* 162

pulp mill

wastes

soda floe. Clostridium thermocellum

corn stover C. thermosaccharolyticum or C. thermohydrosul f ur icum 58,

162, 174

cel 1 ulose Thievlavia terrestris (anaerobe - Z. mobilis)* 182

bal 1-mi 1 led cellulolytic mesophile Clostridium saccharolyticum sp. 212

pulp, paper,

cellulose powder

straw Trichoderma viride Saccharomyces uvarum Pachysolen tannophilus

213

*Conversion of Cg sugars only.

Using rONA techniques, scientists at Helicon are working on the isolation

of genes from Escherichia adecarboxyl ata for the conversion of cellobiose to

ethanol via glucose. Their work includes attempts to get expression of these

genes in an E. coli system and on the characterization of the enzyme system

(184). The ultimate goal of this research would be to express the genes in Z.

mobi 1 is

.

81
.4 Technologies Important for Bio-Industrial Processes

As indicated in previous sections of this report, developments in certain

technologies will be essential to the commercialization and market success of

products developed via biotechnological processes. Technologies that will be

critical to the industrial and, ultimately, commercial success of organic

chemical products synthesized by biological routes include biocatalyst , separation and purification, process monitoring and control, and rPNA

technologies . The following discussion will illustrate examples of research and

developments in these technologies and their potential application.

.a Biocatalysts . A catalyst increases the rate of a chemical reaction but

does not effect reaction equilibrium and is itself usually unchanged by the

reaction. It does this by lowering the activation energy necessary for a reaction to proceed. Catalysts are used throughout the chemical industry. A

few examples of catalysts used in industrial and technologic! al ly important

processes include those used in the manufacture of sulfuric acid and ammonia and

the "catalytic converters" used to convert carbon monoxide to carbon dioxide and

nitrogen oxides to nitrogen and oxygen in automobiles. Enzymes are proteins

which act as catalysts (biocatalysts). Some enzymes facilitate very complex

reactions or series of reactions. By selection of suitable enzymes, one can obtain very specific reactions to transform large complicated bio-organic

macromolecules such as cellulose to simple organic feedstock molecules (e.g.,

ethanol, acetic acid, etc.). In effect the choice of a specific enzyme

promotes the reaction path from among the many possible reactions. Examples of

their use are the conversion of whey to glucose and galactose and the hydrolysis

of cellulosic materials to ethanol.

The ecological backgrounds and biochemical diversity of microorgan isms

should make them important sources of a wide variety of biocatalysts (enzymes)

for biochemical conversion of raw materials to higher value added organic

chemicals. In spite of this potential, only about 20 of the approximately 2500

known enzymes currently have commercial significance. The current annual market

value* of these 20 enzymes is about $125-150 million and they are used in the

pharmaceutical, food processing, and chemical industries (214). Many

*The projected market value of bulk enzymes is $500 million by 1986 (Chemical

Week, May 26, 1982, pp. 44-47).

82
extracel lular enzymes could become articles of commerce if they could be

produced in sufficient quantities to make biochemical conversion processes more

economical than chemical synthesis ( 215 ). However, commercial production of

additional enzymes will require continued development of microbial growth and

isolation procedures, developments in applied genetics, improved/real time

process control, and implementation of continuous process operations.

Commercial development of industrially useful enzymes is a long and capital

intensive process. Some of the approaches that, individually or in combination,

have been and could be used to increase enzyme yields are listed below ( 215 ):

Synthesis of Enzyme

0 Optimize culture conditions: temperature, pH, carbon source, nitrogen

source, aeration, agitation, induction of inducible enzymes, and

maintenance of the secretory phase.

0 Genetic approaches (such as recombinant DNA) to remove restrictive

control mechanisms.

0 Applied genetics approaches such as mutation and selection, rONA, gene

amplification, and gene fusion and use of cells with specialized

cytomembrane structures.

Secretion of Enzyme

0 Addition of chemicals to make the cell wall/membrane complex more

permeable ("leaky").

Rate of Reaction

0 Optimize substrate concentrations, pH levels, temperature, ionic

strength.

0 Obviate end product inhibition of enzyme.

0 Immobilization of enzymes/cells to extend use (re-use) of biocatalyst,

and facilitate removal of products.

83
Use of enzymes in organic chemical synthesis offers some important advantages over traditional methods of chemical synthesis. For example, enzymes are catalytic, highly reaction specific (even stereospecific), produce exceptionally

pure products, and allow synthesis to be carried out under milder reaction

conditions (216, 217). Enzymes also possess some qualities that have limited

their industrial usefulness and reduced their economic potential. For example,

enyzmes operate effectively only in a relatively narrow range of reaction

conditions (aqueous solutions, moderate temperatures, pH sensitivity; 216).

Also to use expensive enzymes once in reaction mixtures such as batch

fermentation vessels has additional industrial economic disincentives:

0 the initial cost of the enzyme;

0 the need to remove the enzyme from the product;

0 extended reaction times to reduce enzyme costs; and

0 the need for large reaction vessels with concommitant high capital costs

(218).

Thus, due to the incongruity between the relatively limited conditions

associated with chemical reaction processes and the "delicate" nature of biocatalysts, enzymes had only limited applicability as catalysts in industrial

processes until the early 1970s. Because of newer approaches which make enzymes

insoluble while retaining their catalytic activity (analogous to in vivo association of enzymes with cell membranes, mitochondria and other cell organelles),

industrial use of enzymes via immobilized biocatalytic (enzymes and microbial)

systems has begun to expand (132, 219-221). Such enzymes are adsorbed, co- polymerized, covalently-bonded, or otherwise attached to an inert waterinsoluble material. This mechanism of attachment permits them to be separated

physically from both substrate and reaction product, and also allows them to be

recycled or reused easily. Immobilized enzymes have significantly enhanced

processing and manufacture (for example, Cheddar cheese) because they can be

integrated into a continuous process and can be recovered for recycling (222).

In addition to use of immobilized biocatalysts to improve process economics

and throughput, development of mathematical models to predict biosystem process

behavior and to describe quantitative relationships between process variables

will be essential to the success of optimization strategies for biotechnological

84
processes that are eventually used. As one starting point in developing process

models, thermodynamic information is required to predict equilibrium (yield) and

the concentration of enzyme-catalyzed reaction products; the heat absorbed or released from the reaction; and the conditions, such as temperature, fluid

forces, pH, and substrate concentration, necessary to maximize a chemical

process (for example, the enzyme-catalyzed hydrolysis of cellulose and

hemicel lulose to lower molecular weight products which can be converted by

fermentation to commercially useful chemicals). A detailed knowledge of the

thermodynamics and thermochemistry of enzyme-catalyzed reactions can be used to

optimize biochemical processes and to minimize the side reactions which give

unwanted by-products.

In order to develop predictive models of biochemical processes and to

select the appropriate reaction pathways, equilibriun data are necessary. Data

banks of evaluated thermodynamic data will be necessary to predict the equilibrium, yields, and heat 1 iberated/ absorbed in enzyme-catalyzed industrial

reactions. These include heats of formation, entropies, and heat capacities of

organic molecules. After the selection of the appropriate reaction, the rate of

the reaction will depend on enzyme concentration, temperature, pH, metal ion

concentration, etc. Therefore, chemical kinetic data will also be necessary for

choosing the proper industrial process conditions.

Currently, at the laboratory scale, corn residues (corn waste) have been

converted to both ethanol and acetic acid in significant yields by Clostridium

thermocel 1 urn (an anaerobic bacterium). An understanding of the thermodynamics

and energetics of the reactions under various conditions will be essential for

the implementation of such a process on an industrial scale.

.1 Immobilized Enzymes . The basic enzyme immobilization methods are adsorption, cross-linking, entrapment, microencapsulation and covalent

attachment (69, 217). The operational advantages of immobilized enzymes are:

0 reuseability and increased operational stability of enzymes;

0 possibility of both batch and continuous process operation modes;

0 rapid termination of reactions;

0 greater variety of engineering designs for continuous processes;

85
0 reduced cost of operation; and

0 potential for efficiency gains in multiple enzymatic reactions (218-220,

223)

.

Table 5.10 illustrates chemical and physical methods employed for the

immobilization of enzymes.

Table 5.10

Methods for Immobilization of Enzymes

Chemical methods —(Covalent bond formation-dependent)

Attachment of enzyme to water-insoluble, functionalized polymer

Incorporation of enzyme into growing polymer chain

Intermolecul ar crosslinking of enzyme with a multifunctional,

low molecular weight reagent

Physical methods --( Noncovalent bond formation-dependent)

Adsorption of enzyme onto water-insoluble matrix

Entrapment of enzyme within water-insoluble matrix

Entrapment of enzyme within permanent or nonpermanent

semi permeable microcapsules

Containment of enzyme within special semipermeable

membrane devices

Source: Reference 219.

Table 5.11 reviews the advantages and disadvantages of some immobilization

methods, as well as prospects for their industrial utility.

There are some disadvantages to enzyme immobilization. A major one is that some enzymes are partially inactivated by immobilization. The key to the

science (or art) of enzyme immobilization is to balance the loss of enzymatic

activity that occurs upon association with support materials with the long-term

86
Table 5.1 1

Advantages and Disadvantages of

Seme Methods for Immobilizing Enzymes

Method Advantages Df sadvantaqes

Enclosure in semi permeab le membrane sacHigh retention

of initial

act i V i ty

.

No long term

stabi 1 izat ions of activity. Clean

stream needed.

Entrapment in polymeric

mater i a 1 , e. g.

,

cel 1 ul ose acetate

Easi ly used,

1 ong 1 i fe (with

suitably long

lived enzyme).

Some enzyme

inactivation on immobi 1 izat ion.

Adsorption to charged supportsEasi 1 y done.

High enzyme

act i V i t i es immobi 1 ized.

Often pH

optimum change

ach ieved.

Hydraul ic properti es of supports less

than ideal.

Impermanent

enzyme-support

1 i nk.

Absorption into

porous supports

Easi 1 y done.

H i gh act i v i t i es achieved. Good

hydraul ic propert i es.

Di ffusion of

substrate/

product to/ from

enzyme di f ficult.

Enzyme can "leak"*

Supports costly.

Absorption or adsorption plus cross- 1 i nki ng

Good retention

of activi'^y in use. Sometimes greater

temperature

stabi 1 i ty.

Initial act i v i t i es

’ owered.

Most supports

costly, need

re-use.

Cova lent

attachment to

act i vated

support

material

(3ood retent i on

of activity in use. Good nydrauiic

propert i es. Sometimes greater

temperature

stabi 1 i t"/.

Possible leakage

of activating

chemical.

Source: Reference 218.

Commerc i a I Use

Uh certain

Yes. Glucose,

isomerase, etc.

Yes. Gl ucose

i somerase,

penici Min

acy lase

Lhcertai n

Yes. 61 ucose

i somerase,

gl ucoamy lase

Lhcertai n

87
retention (half-life) of activity (218). For economically feasible industrial

processes, extended enzymatic activity half-life is more valuable than high

initial enzymatic activity. To improve enzymatic half-life activity and the

diffusion of substrate into the enzyme (active site)--polymer support system,

additional research is needed. Some examples of promising leads includes the

use of magnetic fields to influence the orientation of polymers (224), the

application of ultrasound for accelerating enzymatic activity (225), and the

regeneration of the catalytic activity of thermoinactivated enzymes (226).

.2 Industrial Applications of Immobilized Enzymes . To date there are many

more applications of immobilized enzjmies in analytical fields than in large

scale industrial processes (227-230). The largest in tonnage and most widely

known current commercial application of immobilized enzymes is for the partial

i someri zat ion of glucose to fructose in the production of high fructose corn syrup (HFCS; 231). The major application of this technology in the pharmaceutical industry is the conversion (via fermentation with the enzyme penicillin

acylase) of benzyl penicillin to 6-aminopen ic i 1 Ian ic acid for the preparation of

semi-synthetic penicillin (220). Table 5.12 summarizes industrial process and

pilot plant developments in the application of immobilized enzymes.

Although a variety of approaches can be used to acquire and install an immobilized enzyme system in a commercial operation, the approach most commonly

used is to obtain the complete catalytic system from an external supplier, such

as Corning Glass Works, Monsanto, American Cyanamid, R.J. Reynolds, or Union

Carbide (232). Experience has demonstrated that a close working relationship

between system supplier and buyer is essential for the tailoring of the system

to meet the user's specific needs. This experience may also allow biocatalytic

system suppliers to forward integrate and to become producers of commercial

products

.

From a commercial immobilized enzyme system user viewpoint, objective means

are essential to compare the industrial performance of different catalyst

systems or similar catalytic systems from different sources is essential. Means

for objective evaluation is often unavailable and the data that are available

are difficult to compare. Thus, comparative evaluation of competing systems is

88
Table 5.12

Application of Immobilized Enzymes in Large-Scale Industrial Processes (I)

and in Pilot Plants (P)

Process*

Company

Anheuser-Bush

Cl i nton Corn ( USA)

Car-Mi (USA)

Corning (USA) U.S. Army Natick Center (USA)

Denki Kagaku (Japan)

Diamond Shamrock (USA)

Gist Brocades (Holland)

Gulf Oil (USA)

Hameen Peruna (Finland)

I Cl (England, USA)

Lehigh Uhiversity (USA)

Novo (Denmark)

Sanmatsu (Japan)

Snam-Progetti (DeBi; Italy)

Tanabe Seiyaku (Japan)

Va I i o ( F in I and)

CPC (USA)

Amstar ( USA)

Suddeutsch Zucker (WG)

a b c d h

*(a) synthesis of L-aspartic acid with the use of immobilized microbial

cells containing aspartase

( b) synthesis of L-malic acid with the use of immobilized microbial cells

containing fumarase

(c) production of 6-ami nopen ici I Ian ic acid with the use of immobilized

pen ici I I i n acy lase

(d) resolution of racemic mixtures of amino acids (produced by chemical non- enzymatic method) with the use of immobilized aminoacylase

(e) production of D-amino acid derivatives with the use of Immobilized

hydantoinase

( f) production of g

I ucose- fructose syrups with the use of immobilized glucose isomerase

(g) use of immoblized lactase for production of non lactose milk frcm milk sugars, and

for production of glucose/galactose syrups from whey

(h) production of glucose from partial hydrolysates of starch with the use of

immobilized glucoamylase

(1) production of glucose and/or ethanol from cellulose with the use of immobilized cel I u lase

(j) production of g I ucose- fructose syrups from sucrose with the use of immobilized invertase

Source: References 183, 216.

89
extremely difficult (231). Fundamental information on and standardized methods

to evaluate biochemical, mechanical, hydraulic, etc. characteristics of

processing parameters is essential. This would allow the development of

performance data on a more unified basis. It would also be possible to

establish kinetic requirements for multiple key process parameters for the

efficient engineering design of commerical reaction systems. Examples of data

needed include enzyme activity reaction half-life, catalytic packing density,

external and internal mass transport efficiencies, reactor contact efficiency,

and residence time distribution (Table 5.13; 231). Because of proprietary

positions, however, enzyme producers and enzyme system suppliers may be

reluctant to advocate standards in this area.

.3 Immobilized Cells . Serious work on immobilized cell research began in

the early to mid-1970s only after the impetus given by advances in immobilized

enzyme technology. Most of the research on immobilized cells (microbal or plant) has described techniques for entrapping cells in gels, although other

procedures such as covalent attachment, adsorption, and cross-linking leading to

insoluble aggregates have been used (131, 233). Some examples of cell

immobilization procedures are given in Table 5.14.

90
Biochemical

Characteristics

II.

Mechanical

Characteristics

4-

j2 in (H C N U o —

4- o o "O

(/> +• «4» 3 5 ® c

:s CO lu

a> o c N Q 0 n to © c 3 c o 4- T? to c »— <5 C i. <n 4- O to

(0 x: 3 »— c

0) > o 4- o c U> 0 o o ”0

J- on

u

Q. o 0 C o

*0 c 4* (0 to

u

u o c/) •— 0

cn "5 o

>- 4-U % c0

Ci 4- * CO > 4- » © S«ir C s r ® <*3 •mm 4- Q. 0 (0 E o (0 0 N >- *P O o *— 1. •— •—

o

1. (0u

O t- 4- o —

C/)

Q <n < c.

a o cL t ©

(0 IL. (TJ to

© 0 0 > ® O' c 0 © O. » 0 4- c (n o o '4- C3 c c

<J -P

©

O 3 0 s © .C s

•—

u U)

u (/)

u

— -a 4- 4- £

+- 0) CL

© 4- y>

© (0 •— (T3 CO

u c o g t © *3 "n •mm to im c c

in s

**

O 0 (0 3 u o 0 X

© 4-

©

Q.

Q CO o a 0- (0 0. z CO < QC (/) mJ z

1. *o>

<N

m 'O x| CM in VO CO ou

Q.>.

fOuoo

1.os o<

•oc

<0 E o c >- CL _ *— 0 0 4- g 1 1- 4- o 0 o /X, <0 C > o • — 0 u (0 C 0 MX Sx* 0

(D 4- 0

8,

T3 4- 4-

C •— >- 0

-J 0 >- 0 0 4- 0 u 0 C c MX % 0 c <0 a: 0 o — X CO 0 n c CO MX s Q. 0 O 0 "O E O 3 0 0 3 0 o Ck o 0 > E 4- b ^

4- 4- 0 0 UJ xz 4- C

(0 c (0 0 Q

X E 4- MX 4-

C 0

4- L. O) to 0 c. 0 XM 0 s 4- 00 4- MX 0 2 O L. •X E 0 3 C

>• to

— •— Lu $ >- L.— 0

(0c

>. o

-H

—

— 4- > fO u

a T?c e +-NO

C o &

'4- 0 o © CO LU

_ UJ c o 0 3 h- 1 0 MX § 8 "O s, C

!5

1.0 © 0 0 E L. c 0 0 0 4- fe © 0 0. 0 L.

•fe lx 0 tx c 1 o O 0 CL XM >- X 4* L. 0

1x1 LU Z cc Q. CO 0. z on

tn VO QO Ov o CM

m

(N0C0

L.010o

cn
Table 5.14

Cell Immobilization Procedures

Covalent binding

Hydroxya'i kyl methacrylate ( gl ut aral dehyde)

Carboxymethylcel 1 ulose (carbodi-imide)

Entrapment

Polyacryl amide

A1 ginate

Cel 1 ulose- tri acetate

Agar

Carrageenan

Chi tosan

Collagen (gelatin)

Polystyrene

Urethane

Nylon (micro-encapsulation)

Adsorption

Amon-exchange resin

Dowex 1 DEAE-cel lulose

Ion-exchange cellulose

Polyvinyl chlor ide and

porous bricks

Cross-linked pectate

Metal oxide

Bioadsorption: concanavalin A

Cross-1 inking

Gl utaral dehyde

Albumin and gl utaraldehyde

Gelatin and gl utaral dehyde

Source: Reference 233.

For further developments in this field, it will be necessary to understand

the nature of the cells (viable-resting or growing, non-viable, permeabi 1 ized)

once they are associated with a matrix (220, 233, 234). Although relatively

simple reaction systems are employed currently for both immobilized enzyme and

cell systems, immobilized cell systems (prokaryotic and eukaryotic) offer

greater potential for developing reusable, multi-enzyme, complex reaction

systems (153, 233-235). Thus, immobilized cells, exemplif^'ed by their use in

water purification and pollution control systems, may have a broader range of

industrial applications than immobilized enzymes.

Generally, most of the methods of immobilization employed for enzymes can also be used for cells; however, much greater care must be taken to prevent

92
inactivation of desired cellular activities (218). Also many of the same process design considerations noted for immobilized enz,ymes in Table 5.13 are

also applicable to immobilized cell systems.

Table 5.15 highlights some advantages and disadvantages of methods for

immobilizing whole cells, as well as the potential of the methods for using

multi-enzyme systems. The best known immobilized cell process is that of the

Tanabe Seiyaku Company. In this process, E . col i cells having a high aspartase

activity were trapped in polyacrylamide. They were subsequently treated to

disrupt the normal cell permeability barrier and, as a result, they stereoselectively converted ammonium fumarate to L-aspartic acid (132).

Some specific advantages of immobilized cells over immobilized enzymes for

continuous biocatalytic activity in industrial processes include (132):

0 high yields of biocatalytic activity are achieved on immobilization;

0 enzyme extraction and purification processes and costs are eliminated;

0 immobilized cells have higher operational stability;

0 actual cost of the "enzyme" is reduced;

0 intracellular enzymatic machinery of immobilized cells generally

remains intact;

0 immobilized cells allow use of intracel lular and generally unstable

enzymes; and

0 immobilized cells appear well suited to multi-step biocatal ic

conversions and coenzyme regeneration.

Immobilized cell systems have some disadvantages such as leakage of

catalyst and support system, production of secondary products, potentially high

capital costs, and fewer industrial successes. However, once particular

93
Mo+!iods

for

Imrriobi Iiz1

ng

Whole

Cel

Is

Use

wl

th

mult

iMethod

Advantages

Pi

sadvantages

enzyme

systems

Enclosure

in

semipermeab

le

membrane

Gentle

Fouling

of

membranes.

Uncertain

No

stabilization

of

activity.

0>o c © c

+- x:

©t © 0 ©t © u © o U1 oi U) Ul to (/I i/) o 5 ©

>-

©

©

>-

©

>-

©

>-

©

>- 5 No c

(0U)uo

>-cO

>.NC

LU

0)oe

>-NCo oo > to •H v- O) U 0 © Q — T3 O L

Co

©

© cx >- c

to o cx

— 0 —

T5 o 2 to 4- 2 © to -H 1 8, ©© ©> u

4-

© >-

to

©

C o © jx: t3JD

C

© 0 cn

u©©

© ... O) c ©©

©

C

g CO

© to 4- c o © C © oi 4- c © c 4- u c © c _ © © © © E

*—

©

—

©

© o — >

L. 3 +- .c 1 E u •— — © to u 1 <0 4“ © O 0 to 2. — to O © 0 c © U © .c 0 L. E © 0 © o (J CO o — o CL cr

(.0 (0 LU 0o

-a 'g >- — c — o

tn

—

<0 +• 0 N 0 —

•H 2^ t3

(0 o oc

+-

(0 • . ^ <D — ^ C — > O —

*0 (TJ

O

(/) (D

©Co

"O > o ^ — U7 ^ — 0 U 3 S

(0 (T3

—

-O

©

O >. I ^ o c CO ° "8 ^ 8) © <0 +-

1/1 c a — 4- X ® — — T3 = o — C © to to o o ® © c © S

"®

©

Q = o o o — UJ </) o UJ (3 — coo ©©>— 1 t3

(0c

4- o • ©

—

© c C o to k. > o 0 © © 3 4- — 4- 4- 4- C <0 *o © • t © © c "O © U L c © © 4-

to

®

© a.

—

C 2 o C 1. ^ ^ 5

,'lI • I'

'll O !jI] ©©

•Kuu

>* +*• — (O o +- o. ©o c © c © © © *o c

4- to

©

© c C 0 E 4- © m — © © Ol 8, g 3 — C © ©

O. >- w*

L

©L OlU

L.

©

O

—

© 0

4-

a < < o CJ

C 1 1 1 1 1 UJ CN KO in

•o

to

©

to t t0

4-©>

or 0 CL JZ

Cl Q. + 4-

o. 3 U MB 3 (0 ©

to

to o to T3 3 4- — © o —

Ol i. 4-©

t_ 0 C

a

© CL 0

JTg

U 0 .C

© 0 4- 4- u L.

O

C

©

L

©

O

4- •— 4-

© O) cx

4- 4-

© a.

C c ©© 3 0 O E c to •— 4- *—

4- 4-

C 4-

— 4- Q. CL © t_ 1 3 U L. 0 to 0 o O © a. to jr

to to > Q. 0 4-

*o J3 0 3 u < < o (0 o 7

CN©Vc©

L.©

M-o

CO

;!
immobilized cell systems are evaluated regarding their potential for industrial

use, it will be possible to lower capital operating costs of industrial

biochemical processes by reducing the size of the reaction and/or fermenting

vessels and by increasing the throughput of the substrate stream. The

realization of the potential benefits of this technological "tool" will depend

on identifying the appropriate role for it in an industrial process (for

example, in the conversion of cellulosic material to commodity organic

chemicals, in a specific biotransformation to produce a higher value added

product, or in the replacement of a unit process step in a complex chemical

synthesis)

.

.4 Research Opportunities . The following lists some areas of research

opportunity for both immobilized enzymes and cells that could increase the

potential industrial usefulness of this technology:

Immobilized Enzymes- -

0 production of membrane associated forms of water soluble, labile,

enzymes; .

0 stabilizing larger molecular weight enzymes by modification of the

surface amino acids, by cross-bridging of the active site with

neighboring molecules, by combined thermochemical and photochemical

linkages, by designing semi-synthetic enzymes, etc.;

0 immobilization and/or microencapsulation of expensive and labile

coenzymes for recycling or continuous use;

0 understanding physicochemical reactions for stabilizing enzymes to

permit higher product yields;

0 CO- immcbi 1 i zing series of enzymes for sequential biochemical reactions;

0 developing kinetic models for individual reaction rates to achieve

steady state in a complex reaction system;

95
0 reversing catalytic loss (enzyme rejuvination) ; and

0 immobilizing enzymes that will operate effectively in complex and/or

"dirty" streams.

Immobi 1 i zed Cel 1 s

0 understanding how dissolved oxygen levels affect immobilized cells and

their nutrient use;

0 operation of immobilized whole and permeabi 1 i zed cell systems under

harsh industrial conditions;

0 operation of immobilized cell or cell systems in substrate streams

(slurries), and under solid state fermentation reaction conditions;

0 genetic manipulation for amplifying cellular synthetic pathways useful

in specific industrial conversions at the expense of other "wasteful"

pathways and enzymatic reactions; and

0 control of decay of biocatalytic activity.

In addition, modification and/or tailoring of polymers as biocatalyst

support devices for--(a) improved process throughput and substrate loading, (b)

enhanced biocatalyst orientation, half-life reactivity and stability, and (c)

better coupling with product separation processes--could facilitate industrial

development of more "solid phase" biotechnological approaches to industrial

synthesis of organic chemicals (236).

5. Immobilized Cell Technology and Fermentation . Research in immobilized

cell technology is beginning to affect bioprocess used in the fermentation of

sugars to ethanol as a means to increase cell density, to reduce the volume of

fluid used in traditional fermentation vessels, and to facilitate development of

continuous processes.

96
Entrapment of cells in relatively non-toxic polymers such as polyacrylamide, collagen, agar, alginate, carrogeenan, polystyrene, cellulose

acetate, etc. has been the method most widely used for the immobilization of

cells. Calcium alginate and carrageenan have been used successfully to

immobilize yeast cells for the production of ethanol. Listed in Table 5.16 are examples of some of the research activity and developments in this area.

Table 5.16

Research Activities Related to Im.mobilized Cells

Sugar

Immobi 1 i zat ion

Polymer Organi sm Substrate

(g/L)

Ca-alginate S. cerevisiae glucose

(100)

Ca-alginate S. cerevisiae cane molasses

(175)

Ca-alginate Z . mobi 1 i

s

glucose

Ca-alginate S. cerevisiae glucose

(30)

Carrageenan S. cerevisiae glucose

(200)

^Reference 200.

% of Feed ?^ax. Ethanol

Sugar Productivity

^ Use (g ethanol/L/H)'’ Reference

80 4.3 131

83 21.3 198

97 71 200

94 21 199

100 40 237

Members of the Japanese Research Association for Petroleum Alternatives

Development (Ajinimoto, JGC, Kansae Paint, Marnzen Oil, Sanraku-Ocean ) have

reported recently the completion of a pilot plant for the rapid, continuous

conversion of biomass to ethanol via yeast cells immobilized on radiation cured

polymers (238).

In addition to these studies, a variety of other approaches for stab'lizing

whole cells in polymer matrices are being explored regarding their utility for

larger scale bio-reactor systems. Table 5.17 gives a sampling of experimental

efforts in progress.

97
Table 5.17

Experimental Efforts to Stabilize Whole Cells in Polymer Matrices or on Surfaces

Immobi 1 i zation

System Organi sm Substrate Reference

Ca~alginate + cross

"linking for stability

(a. -polyethleneimine

and gl utaraldehyde;

b. -carbodi imide and

N-hydroxysuccinimide;

c. -periodate and

polyethyleneimine)

S. cerevisiae 239

Ca alginate P. tannophilus xyl ose 208

Ca alginate K. fragilis whey 240

Crude egg white

+ gl utaral dehyde

C. acidophil ia -- 241

Polyurethane foam and gel -- -- 242

Ch i tosan E . col i L-serine 243

Induced bacterial

filamentous growth

E. coli

S. typhimurium

, Baci 1 lus species

— 244

Ion-exchange resins S. cerevisiae gl ucose 201

Titanium activated

microporous inorganic

supports

S. cerevisiae sucrose 245

Fixed film reactor with

treated (gelatin,

glutaraldehyde)

Rashig rings

S. cerevisiae glucose 113

Continued advances in cell immobilization technology coupled to innovations

in biotechnological processes will improve the economics for future use of such

bio~systems in the industrial production of organic chemicals.

.b Separation and Purification Technologies . As fermentation is central

to biotechnological processes, the ability to separate specific components from

highly complex fermenter slurries is essential to the success of industrial

fermentation processes.

I

98
The expanded potential for use of continuous fermentation and cell

recycling processes in biotechnology offers cost reductions in comparison with

batch fermentation, because the equipment may be smaller and more energy

efficient for an equivalent throughput and fewer production delays are encountered. The realization of this potential, however, will require

innovations in separation, extraction and purification technologies, since:

0 process technologies developed will depend upon the physical and chemical

properties of the fermentation broths; and

0 these properties are difficult to measure in terms of quantifying the

mechanisms of the process (246).

In terms of process economics, the cost of recovery procedures for high

value added organic chemicals products developed through biotechnologies will

not be as critical as those procedures developed for recovery of commodity

organic chemical products from fermentation processes. If bioprocess substrates

can be economically or politically as desirable as petroleum or coal and if

microbial reactions are as economically and technologically attractive as chemical substrate conversions for producing commodity organic chemicals, new

separation technologies using methods based upon low energy and high throughput

per unit of capital cost are required (246).

The subsequent paragraphs in this section will highlight some important

separation and purification technologies, some developments in the field, and

the potential applicability of these technologies to industrial biotechnological

processes for the production of organic chemicals.

.1 Membrane Technology . Membrane techniques or combinations of membrane

techniques may have application in continuous alcohol removal and fermentation

cell recycling. Membranes that are relatively permeable to solvents and

impermeable to most other solutes present in the fermentation broth may allow

enrichment of alcohol concentration to levels sufficient for recovery from

biological systems with relatively low alcohol tolerance (162).

99
Technical interest in processes of mass transfer through membranes has

j increased substantially in the last decade. Table 5.18 lists technically

important membrane separation processes and relevant properties, and, of these i processes, ultrafiltration, hyperfiltration, dialysis and electrodialysis are

^

presently of the most technical significance (247). ,

It is not possible here to discuss the advantages and disadvantages as well

as applicability of each technology listed in Table 5.18 in this section of the | report. Rather, this section of the report will illustrate the potential

applicability of some of these technologies, such as ultraf iltration , to

separation and purification processes in industrial biotechnological processes.

I Ultraf iltration has important industrial applicability in the purification

'

and concentration of biological molecules due to its ability to minimize

denaturation and decomposition losses of labile, high value added products. A

review by Michaels summarizes recent developments and limitations in the use of

ultrafiltration (248).

Additional research and development in several areas of ultraf i 1 tration

technology will be essential in order to understand biological, chemical,

' physical, and hydraulic design parameters and before considering the broad

application of this technology to large scale biotechnological and fermentation

processes. Some general areas for technology improvement include:

I

0 modification of membranes to reduce or eliminate fouling; I

0 improved fermentation broth conditioning techniques; ,

i

K

I 0 adaption for high volume, aqueous stream concentration and purification;

0 modification of the polymer chemical characteristics;

0 improved selectivity of membranes for separations;

.

100
CO

inO

(O

<DOou

CLco

(0us. COc

<T3 to

Q.s

(0oco0

4- c y

L

U

O v2 0 5

4-0

4-0 L

O o 0 +- »— o y C c c 0

MW 0

4- 0 0u c0 oc

•(- 0 3 O

in *0

y

a Q* • o 4- 0 0 o in •— b) L C

4- •0

O 4- 0 % C C L 0 0 0 0 0 0 4- u L 0 u L, 3 3 •0 3 3 0 i

(0 0) 0 4- 0 0 0 0

(/) in L.

C 0 0 0 0 CD 0 0 0 b,^ 0 u

a. b.

C

“6

u

a. fe. 3 C O

u 0 CL 0 y

L. 0 y

•“

y

y •“ 0 4- 4- T3

U_ «!- 4- 0 0 4- 4- 0 c 0 1. y 0 0 4- C 0

CT +- •f- 4- 4- +“

C 4-

C SQ 0 Cu 0 0 = JL 0 O 0 4“ 0 0 » © 0 > L. U y y L L. •0 y y

•0 *o c 0 0 -0 0 c

1. >- >•

O >. 0 c Q £ y 0 £ o — 00c0 0 L 0c 0 E 0 0 0 0 0 —

(/)

u 0 C £ 0. 0 3 c 0 C £ 0 u 2

*" 0 0 L. 1. £ £ 3 3

X) £ 0 ^ £ E >• 0 0 0 0 0 4- £ c 3 0 — ifc E 0 0 0 0 C

(/> 3 — 1. 0)

L 0 *0 0 0 Q. t ^

!o 30

-g c0 a

u

O 3 1_ 0 JC £ U 0 £ y U o 0 0 Q Q. X 0 a.

u

Cl

>* Q. 0

8, 0 0

3. 0 3 4- t- y y 0 c c 3 0 0 0 y 0 0 0 y C 2 0 0 u i. >— •— 0 3 C C 4- 4- u y C u w 0 0u i i

4-

X0 y

8,

4- >- o 0 £ E 0 0 4- — MW £ >K

er

10 £ c 0 E 0 -r 3 02

in0 5r

O

§r 0 O E ^ £0 0 0 C C

C 0 O 0 co

4- o o 0U 3 0 0 3 t

(0

— 0 0 0

Q. 4“

Q 0) 0)Q 0 0 y 0 L. U 0 u

CO 0 0 0

*0 £ •0 “0 C y y c c o 0 0 0 0 0 O) c c 0 0 c c — c o 0 y y 0 0 c

Q.

— o c 0 0 i_ i. 0 0 —

O 0 3 3 4* 3 3 4- —c 0 »- 4- y y y u 0 0 4- (0 ^ ©

qI

y •0 L. —

*0 0 0 0 *0 c c 0 0 o 0 » c C *— •— 0 0 o 0

4- >* *— l_ 0 0 — 0 4- L 1. 0 >- 0 £ c

4- -1- «— L. 0 O — 0 0 0 0 E —

(/) 0 Q. 4-

(/) 4- 0 u 0 •0 0 0 0 t. 4" 0 ^ © y L. 0 — y N 3 E 0 4- o. 0 0 0 0 or l- u — — 0 — ©

CL 3 X

Q LU CL e> wJ 3

p«-

CNJ 0oc0

i_00

tr0o

1.3O

CO
0 deactivation of certain contaminating enzymes; and

0 removal and/or concentration of particulate solids such as cells/cell

debris from fermentation broths (246, 248, 249).

The potential for hollow fiber ultrafiltration membranes in larger scale

industrial processes is illustrated by the following two examples. First,

Monsanto has developed a proprietary line of prism hollow fiber separators.

These separators, currently being used to remove useful hydrogen from certain

wastes, may have potential in separating and purifying fermentation products

(250). Second, RPI has also developed a hollow fiber ultrafiltration process.

It is currently being considered for use in the trapping of viruses and plasmids

from waste control or separation processes, but may also be applicable to large

scale fermentation process product separation (251). Possible barriers to the

successful implementation on a large scale of either of these technological

approaches are membrane fouling and fragility.

In addition to hollow fiber ultrafiltration, tangential flow filtration

could potentially improve continuous biomass-convers ion-to-ethanol processes by

removing toxic materials, growth inhibitory substances, spent cells, as well as desirable reaction products while recycling viable cells to the fermentation

vessel (252).

liltraf il tration processes could also be coupled to immobilized biocatalyst

(enzyme/cell) system reactors for the continuous: (a) flow through of substrate

into the catalyst, (b) conversion of substrate to product, and (c) diffusion and

convection of the reaction products out of the device (162, 248, 253, 254).

Advances in our understanding and application of applied genetic techniques

to the induction of filamentous growth of industrially important microorgani sms such as E. coli and Bacillus species could facilitate confinement (or

immobilization) of microbial cells on membranes and which then could improve

flow rates in continuous processes (244).

102
Liquid membrane (LM) technology may also have a potential range of

applications to large scale biotechnological processes. LM technology involves

those processes in which there is a simultaneous extraction/re-extraction

process involving a selective separating phase or membrane. LMs are made by an emulsion of two immiscible phases and then dispersing the emulsion in a third

phase, the continuous phase (255). This technology offers a new means for

encapsulating materials and separating mixtures through a permeable membrane,

although no full scale commercial plants using LM technology are currently

on-line (247, 255).

LM technology may be particularly applicable to processes involving large

volumes of fluids, for example, fermentation processes and liquid-liquid

reaction systems (256). The principles of encapsulation of enzymes have been

investigated and the results show the potential applicability of enzymes and

this technology to organic chemical synthesis (255, 257). In addition, research

on the conversion of cellulosic materials to ethanol indicates that LM technology has considerable potential: (a) for continuous extractive fermentation

of ethanol; (b) to recover and reuse enzymes, coenzymes and microorgani sms ; (c)

to increase the operational stability of biocatalysts; and (d) to reduce product

inhibition, reaction volumes and operational costs (188, 194).

Two additional examples illustrate the range of possible applications of

membrane technology to biotechnological processes for the production of organic

chemicals. First, General Electric and the University of Pennsylvania are currently working to improve process ultraf i Itrat ion technology by developing a membrane reactor that will separate the reactant from an aqueous solution until

the reaction takes place and to couple the reaction process to product

purification (258). The membrane reaction is a sandwich of porous polymer

membranes with a membrane on one side of the sandwich containing immobilized

liquid organic solvent selective for the reactant and the other membrane has the

catalyst. Thus the product stream is separated from the feed stream. Second,

Engenics is considering use of bundles of capi 1 1 ary membranes on which

microorganisms can be grown in high densities (258). Some product purification

would be achieved by selective passage of products through the capillary

membranes and thus separate the products on the basis of molecular weight or of

hydrophobic or hydrophilic nature.

103
.2 Chromatography . Materials harvested from fermentation processes are usually aqueous. Traditionally, the first separation stage involves centrifugation. The second step involves ultrafiltration or microf iltration . This is

followed by a purification step that hopefully will give a high yield in a short

time at minimal cost.

Large scale liquid chromatography (LC) may offer possibilities for the

isolation and purification of high value added proteins, for example, interferon

and pheromone bioinsecticides from highly aqueous fermentation broths. Although

large scale applications (e.g.,>1000 liters) of LC have met with only limited

successes in the past. Waters Associates believes that their approach using

prepackaged cartridges of polyethylene or Teflon, where the pressure is applied

radially to the cartridge, will eliminate spaces between particles and between

the bed and the wall (259). This would eliminate a major problem of uniform

packing. In related technical developments. Oak Ridge National Laboratory

researchers are developing a pressurized continuous LC device that uses a slowly

rotating bed or sorbent material, fixed multiple feed points, and fixed

withdrawal locations (259).

A major industrial concern remaining with these and other new large scale

LC techniques that are being developed is how well these techniques will operate

over extended time periods and under industrial conditions.

In addition to these LC techniques, high performance liquid chromatography

(HPLC) either individually or coupled with affinity chromatography could evolve

into an industrially important technique for the rapid separation and purification of enzymes, peptides and other important biological molecules (260).

Another chromatographic technique that may have a broad range of industrial

applications in the separation and purification of organic chemicals and biologically important macromolecules from crude plant/animal tissue extracts is

countercurrent chromatography (261). This technique has already been applied on

an experimental basis to the separation of gl ucose/ fructose mixtures (262).

.3 Supercritical Gas Extractions . There is considerable research activity

underway to examine the potential of dense gases at or near their critical

points as an alternative to liquid solvents in the extraction of organic

104
compounds. Supercritical gas extraction depends upon unusual properties of

gases in the "critical region" where no phase boundries between gas and liquid

states are displayed. When increased pressure is applied to certain gases such

as CO

2

, usually an inefficient solvent, the extractive potential of CO 2 increases as well as its density. With higher density and temperature, the

solubility of organic compounds in CO

2 increases. Above 31°C only a supercritical phase exists where liquid CO

2

becomes a highly selective solvent, for

example, in the extraction: of caffeine from tea leaves and green coffee; of

aromatic oils from crushed roasted coffee; of pyrethrins (esters) from gound

pyrethrum flowers; of nicotine from raw tobacco; of tetrahydroxycannibinol from

marijuana; of oils from sunflower seeds, soybeans, rapeseeds, and peanuts; of

resins from air dried hops; and of cocoa butter from cocoa mass (263-266).

With advances in biotechnology, the potential exists for producing

industrially useful organic chemicals from biomass via aqueous fermentation

processes. In these processes, the selective extractive properties of super

critical CO

2

extraction may be particularly useful in the separation and

purification of such organic chemicals as:

0 low-to-medium weight oxygenated organic compounds, such as esters,

“ ketones, alcohols, ethers and aldehydes, and

0 low molecular weight non-polar organic compounds, such as alkanes,

alkenes, terpenes, as well as certain polar compounds, such as acetic

acid (264).

In addition, this technology may also be applied to lignocel lulose extraction

(267).

Major barriers that will be encountered in the commercial application of

this technology include: (a) a fundamental understanding of the thermodynamics

of the process, (b) the mechanics of rate processes involved, and (c) the design

of commercial scale operations (266).*

*nMR measurements at high pressures could aid our understanding of process

dynamics and intermolecul ar interactions in supercritical gases

(Science 216: 1179-1184, 1982).

105
.4 Monoclonal Antibodies . Monoclonal antibodies have obvious commercial

potential in clinical laboratory diagnostic testing and in health care. In addition, monoclonal antibodies have potential both as probes of enzyme

structure and function and as means for the rapid isolation and purification of

enzymes or proteins from crude extracts, such as those encountered in dilute

fermentation broths (268, 269).

In related developments in the commercial production of monoclonal antibodies, Damon Biotech used a microencapsulation technique that had previously

been used in controlled drug delivery systems (liposomes; 270, 271). The Damon

Biotech method developed is an in vitro model for producing pure monoclonal

antibodies by encapsulating specific hybridoma cells in porous carbohydrate

capsules and exposing the capsules to growth medium that allows the hybridomas

to produce monoclonal antibodies within the capsules. At the end of the growth

period, the nutrient medium is washed from the capsules, the microcapsules are opened with heparin, and the monoclonal antibodies are separated from the

hybridoma cells by centrifugation (272, 273). The monoclonal antibody producing

hybridoma cells are then recovered, encapsulated, and reused.

This encapsulation technique may also be applicable to large scale

fermentation and biocatalytic systems, analogous to proposals for trapping

enzymes in micelles (257), or the immobilization of microbial cells in liquid

membranes ( Micrococcus denitrif icans used in the reduction of nitrate to

nitrite; 274). Development in encapsulation technology could also potentially

be coupled with sedimentation field flow fractionation for the separation of

bio-macromolecules (275).

.5 Biological Separation/Genetic Engineering . An approach that potentially could be used in specific circumstances in fermentation systems to

separate the desired product from the reaction system would be to genetically

engineer the compartmental i zation of the desired product within the producing

organism. In industrial scale-up developments in the production of human

insulin proteins, the genetically engineered E. coli strain employed morphological ly demonstrated the accumulation of insulin proteins intracel 1 ul arl

y

within inclusion bodies (276). In a related development, ICI researchers

106
identified a microorganism, Alcaligenes eutrophus , that can concentrate the (PHB)* aliphatic polyester polyhydroxybutyrate * in intracellular granules (277).

In certain specific cases, these inclusion bodies or granules could be used as means to facilitate the isolation and purification of the desired product from

other intracellular components.

Alternatively, rDNA techniques can be used to insert DNA information

signalling sequences into a microorganism to trigger the extr acel 1 ul ar secretion

of a desired product (278). This approach not only would facilitate subsequent

product separation and purification, but also would allow such microbial cells

to be immobilized and used in continuous processes.**

.6 Dry Separations . Although more closely associated with efforts to reduce

particulate pollution from industrial sources and to remove water impurities,

"dry" separation techniques, such as electrostatic, electromagnetic, and

screening, may have some specialized applicability in industrial

biotechnological processes used for producing organic chemicals. These

separation methods differentiate between particles on the basis of such

properties as size, shape, electrical characteristics, magnetic characteristics,

and density (279, 280). Possible applications of "dry" separation techniques

include the following:

0 recovery or removal of trace minerals;

0 mechanical separation of biomass fractions based on size differences (if

problems of moisture can be overcome);

0 di alectrophoretic separation of specific cell populations (281); and

*Possible example' of future "non-traditional" commodity chemical (biopolymer/

plastic) with additional small volume, high value added applications resulting

from PHB's piezoelectric and biodegradable properties (European Chem. News,

May 3, 1982, p. 15).

**Another approach would be to manipulate genetically the production of certain

intracel 1 ul ar metabolites in microorgani sms having no cell walls. Intracellular metabolites would be released from the microorganism when cells were

placed in a hypotonic solution (Biotechnology Letters 4: 217-222, 1982).

107
Regarding this latter point, a more fundamental understanding of microbial

magnetotaxis and of the synthesis of magnetosomes by certain microbial species

may prove useful in considering future separation technology alternatives (286).

In addition, application of magnetic polymers as supports for immobilized

enzymes or of magnetic affinity chromatography techniques may also be useful for

the rapid isolation of proteins and enzymes from batch and fluidized bed and/or

continuous reaction vessels (287-289). Separations of cells from reaction

vessels could also be accomplished by means of antibodies or lectins associated

with magnetic microspheres that would bind specifically to cell surface

receptors (290, 291)

.

An alternative to both electrophoresis and chromatography in separation and

purification techniques that also may be applicable to biotechnological pro- cesses is electromolecular propulsion (EMP) or the chemoelectronic mobilization

of chemical species in low conductivity fields (292, 293). Although currently

applied only to lipoprotein analysis and to purification of gold, silver and

platinum ores, this technique could induce the movement of non-polar molecules,

such as aromatic hydrocarbons, in a separation process.

f The preceeding discussion illustrates only some of many opportunities for

the potential application of a broad range of innovative separation and purification technologies to large scale industrial biotechnological processes. For

this potential to be realized, however, considerable fundamental research is

needed on the integration of these technologies with industrial process design

parameters and variables.

.c Process Monitoring and Control

As industry expands its use of biotechnological and fermentation processes

for improving product yields and system productivity, greater demands will be

made on process control and monitoring functions and on the automated, real-time
and continuous assessment of process parameters. Information generated from the

development of and/or improvements in more sophisticated computerized process

control and monitoring systems could serve as the basis for development of

predictive models of biotechnological processes and of process parameters.

Areas for improvement include:

0 technical developments in, evaluation and performance comparison of, and

standards for control and monitoring instrumentation and techniques;

0 fundamental understanding of the applicability of on-line sensing,

measurement and analytic techniques; and

0 availabilty of critically evaluated data on key process parameters.

Information and developments in these areas will assist in establishing

reproducible and consistent manufacturing conditions, in quantitatively

improving product quality, in reducing human errors and raw material waste, in

increasing process accuracy and energy efficiency, and in increasing process

productivity.

Although there have been advances in automation of off-line measurement

techniques, these techniques are relatively slow in comparison to on-line

methods that involve measurement directly on or within a biologic system (294).

However, many on-line sensing and measurement systems provide only an indirect

measure of the process paranieter under consideration. Table 5.19 illustrates

some examples of measurement approaches currently being used to monitor biotechnological processes. Since major objectives of capital intensive industrial

biotechnological and fermentation processes are system productivity and product

yield, a barrier to controlling such process for improving process efficiencies

is in the continuous quantitative assessment of substate and product concentrations. However, sensing systems for direct and continuous measurement of

substrate concentration, for example, biomass, and resulting process conversion

products and product concentration are generally lacking for large scale

industrial biotechnological processes (295).

109
TABLE 5.19

Approaches to Measurement (and Control) of Biotechnological Processes

Process Parameters

Temperature

pH

CO

2

O

2 concentration and transfer

Cel I mass

Ce I I g" cwt h

Cel I metabo 1 1 sm

Nutrient uptake

Ionic strength

Cel I morphology - pop u I at i on

di f ferent i at ion

Measurement Approaches

o cooling requirements

o flow calorimetry

o dynamic calorimetry

o glass-reference electrode

o base addition

o off-gas analysis—infrared absorbance

o dynamic assessment: tubing method, mass spectrometer (and gas chrcmatography)

o off-gas analysis (gas flow rate,

inlet O

2 concentration,

outlet concentration)

o dynamic assessment (air flow

temporarily stopped and change in dissolved O

2 measured over time)

o mass spectrometer

o oxygen uptake rate

o cummulative CO

2 production

o total culture fluorescence

(NACH- - NADH)

o electrode system

o |otal amount of ATP

o flow vi sccmeter

o power reqLTiements of impel ler system

o scattered light o substrate consunption

o production of CO

2 o rate of base addition (load cells) o q Lent i tat ion of O

2 demand

o respiratory quotient (RQ) o electrical conductance

o enzyme probe

o dialysis membrane probe

o electrical conductance

o cytof I uorcmetry

Source: Reference 294.

no
The following discussion provides some examples of technological

developments that may improve quantitively the continuous and more direct

on-line measurement of process parameters.

Advances in immobilized enzyme technology have aided considerably in the

design and industrial applicability of enzyme electrodes as biosensors. The

advances have also indicated the potential for use of multi-enzyme systems to

expand the analytical capabilities of enzyme electrodes (283, 296). Such enzyme

electrodes would allow the rapid, specific and sensitive analysis of various

components in complex mixtures. In addition to enzymes, immobilized (or

permeabi 1 i zed) prokaryotic or eukaryotic cells in electrodes may extend analytic

capabilites beyond sensing biochemical substances, and, via cell regenerat ion

,

may increase the operational life of biosensors (297). Immobilized whole cel''

membrane reactor sensing systems could also be employed to measure substrate

diffusivities within biosystems (298).

Both immobilized cell and enzyme electrodes in association with a thermistor could improve industrial microcalorimetr ic measurement capabilities

(299). Developments in flow microcalorimetric techniques may also potentially

expand industrial process control capabilities by providing: (a) predictive

methods for screening polymers, (b) measurements of interactions of biological

molecules, (c) information on surface properties in purification of proteins,

and (d) data on the activity of immobilized biocatalytic systems (300).

NMR techniques could be applied to the monitoring and measurement of

specific nutrients or other components in complex fermentation broths (301,

302). Advances in semi permeable membrane technology may offer sensing

alternatives for the on-line measurement of fermentation product concentration

(303). In addition to these more macro-sensing technical possibil ites,

developments in X-ray laser and microdensitometry and microf 1 uor imetry

technologies could potentially provide information on biochemical activity and

other functional parameters on individual cells in complex biotechnological

processes (304, 305).

Ill
These examples represent just a sampling of the many avenues being

investigated to improve the monitoring and control of industrial bio-processes.

As industry begins to scale-up bio-processes over the next several years, its

demands for better measurement techniques will serve as a stimulus for technological innovation and developments in this area.

d . Genetic Engineering

In addition to traditional industrial microbiological tools such as mutation and selection for improving product yields from bioprocesses, recent

research and developments using new applied genetics-genetic engineering tools,

such as recombinant DNA (rONA), protoplast fusion, and gene amplification via

plasmids and bacteriophages, have expanded industrial options for fermentation

and other biotechnological processes.

Preceeding sections of this report have illustrated several examples of

genetic engineering and rONA research focused on product yield improvements in

the conversion of cellulosic materials to ethanol. Some of these molecular

manipulations attempted to increase microbial ethanol tolerance, to improve the

thermostability of fermenting mircroorgan i sms (metabolic pathways -enzyme

systems), to transfer genes for multi-step enzyme systems for more direct

conversions, etc.

It is unlikely in the near term that NBS research activities will include

genetic engineering of microorganisms . However, it is important to realize in

planning Bureau research activities that genetic engineering will be an important industrial tool. In addition, genetic engineering of industrial

microorgani sms will have important influences on industrial scale-up considerations and on the design of industrial biotechnological processes. Some

examples of these influences include (51):

0 strain stability;

0 type of fermenter-fermentation process used;

112
0 nonsubstrdte nutrient requirements and costs;

0 separation technologies complementary to microbial properties;

0 operating conditions such as cell immobilization and cell recycle; and

0 efficiency of substrate utilization.

Thus, the future success of biotechnology related research and service

activities at NBS will require the cooperation and intergration of many

scientific disciplines. Further, a basic working knowledge at the Bureau of

molecular genetics and of genetic engineering techniques will be essential in

the long-term for understanding and for being responsive to industry needs in

biotechnology within the framework of the Bureau's mission.

113
}i I
6. DISCUSSIONS WITH INDUSTRY AND TECHNICAL GROUPS

.1 Purpose of Discussions and Listing of Meetings

An important element in the process of anticipating future industry needs

for NBS services in biotechnology and of guiding NBS competence building efforts

in this area is establishing a dialogue with industry and with individuals

knowledgeable in the biotechnologies. Such dialogue provides an opportunity to

obtain information on barriers or problems that industry has or will have in

developing products via biotechnological processes, and on how these problems

may relate to traditional Bureau infrastructure services.

Based on the Bureau's projections regarding the importance of biotechnology

in the future production of organic chemicals and of biocatalysts in the

economics of biotechnological processes, the initial Bureau focus for discussions with industry was the organic chemical industry. In addition to

discussions with individuals at firms in the chemical industry, discussions were

also held with individuals from technical associations, academia, other

industries using or developing biotechnological processes, and a high-technology'

investment firm. Table 6.1 provides a listing of the individuals with whom affiliations.* meetings were held as well as their *

.2 Summary of Industry Views on Needs in Biotechnology

The following discussion provides a summary of comments by individuals with

whom NBS staff met from January through April 1982. Summary comments are grouped under two major headings. Standards and Measurement Needs and

Fundamental Science and Information Needs . These summary comments reflect

external preceptions of future industry needs in biotechnology and of the

relationship of these needs to possible roles for NBS in this area. Sections 8

*NBS participants varied from meeting to meeting. With the exception of the

meeting with CPC International, the author of this report participated in all

meetings. Other NBS participants included Drs . Peter Heydemann, William

Kirchhoff, Merrill Hessel , Stanley Abramowitz, Elio Passaglia, and Robert

Goldberg

.

115
00 4- 00 X O

<u 3 X oo <3 • o o

i/i e • »> t r> 3 03

O)

X

X 00 CD 5>,

X 4- E (T3 O 00 3 on O on o CD 3 O

+j •<- o 3 3 3 on

O 3 -o -p

oo i- S- > E O o 3 CD E

O) 4-> Q. 3 3 O •r* 3 O o 3 c 4- oO CD i- X o >— 3 CD O X <D 3 I— 03 Q. O S_ O X 03 5^

+J 3 >> 3 1 i- CL-r- oJ 3 X

X

CC

O CD 03 3 o X 3 03

O • t— •(— >0 O E >, >) O •f— >0 X E d) •" cn CD.— CD CD-'- >5 E CD •" 50 50 4- o o o 3 O O X CD O CDX 3 CD r- CD

CD

O >^r- 3 o 1— 03

O r— 3 3 O

O o o 5o

O CO

OO

X

OO 4-> 1

03 o 3 o CD t— ^—- o c 3 O ti 3 3 C o 3 i- E o o O O 00 X X 3 03 SI X 3 3 i_ X 3 3 03 3 -r- f— c ' Q U X r- OJ o e X X O 3 4- X 4- X o

x: X! E OJ 3 O 3 3 SO 00 3 3 3 U o 3 O 3 o f— >0 4-> X X O X X 3 3 3 X <- on O 4- X 4- X OJ <D M o O CD 00 O O X X XJ O CD 03 00 o O o

-l-> •f— c •>- •r— *r- on -I— •r“ 3 3 3 00 -r- 3 -r- 3 o U_ O) X X X 03 X X X 3 E 03 E o E X

CQc

oo oo

O <70 o CO o O

CQ CM CD LO o CO

C s: oo OO X X X O) CD o CO CD. CM

>4- c 1 t t 1 1 1 1 o o CM CM o ro ro tX X CO •CD CO lO ro CO

O) CL <3^ 1

CM CO CO t— OJ —

o r— CM CM .”4 CM iX

Cd O) 1 1 iX o o c o t— ro CM ro m X CM ro

OJ X-(-> 3 • o O

cr

a lo •r" o o X • ^ 00 d) o

i/i on S- 00 4_) 3

CT> OJ o =t -f- CM

X r—

0) c c 4-> O) o 3 LO o 1— 1— o IT3 3 >) <0 CO o

JD >-> -r— •r—C

X CD

O 3 C_3 T3 OJ 4-> o OJ O "CM X 1— O) fO o > X 1— • 3 X 3

IM 00 < X

O

X • CM

X CT> 4- o 4- 3 CM

00 CM 3 oo -O LO •r“ ro

O o 3 > X

Q.

CC 3 X

O

X 3 > CO

X

X 3: o X -r- CO

fO 03 o ^ o O 3 03 o on CM • CM 4-

O

O CD o2 C3 0O o 3 O .CQ CM 3 O 50 Z 3 Q CM

S- s_ c 3 X 00 >, h—

H

X C_5 CD o X oo o O i. i- 3 3 >1 " <x 3

03

u fO •r— d) fO > Q in c CTi

X o 3 5 O r— C •r— 3 CQ X S •r— 2: X 50 2: 0 0 3 Cl 03 2:

fO s:

X

O O- ^ X

X

X CD

O r— 3 50.— o o 1—1 X r— 3 •' 3 3 O ^ on o 4- 3 X -;«< •r- 3: oo X X >0 •' 03 .-3 d) U OJ oo I— 3 •I— 4-J o 4- 3 e o 4-> 3 S_ X •f— 3 t—^ 3 O CD 3 X X 3 03 X 00 T3 s_ o n: 3 s- 1- • .

X (— X 3 T3 03 O CD 03 Q_ r— o oo s- o CD X o X LlJ -f- UJ U X -f- O 4- ' 3 O -r- O) OJ IT3 O 3 fO • X >- 00 > X > 03 O S- •f- o >

1— s_

C 4-> o U_ 03 3 LO 3 C -X 00 3 X 4_ ro

X 4-

O -a i- •<- o •r- CQ s T3 X O 3 X o on E 3 2: X OO 3 ro

u

"O TD OJ o oo

X • 3 3X0 3 »x o 03 03 E oo

r-H 03 E CM o c «=c CQ on on CJ> UJ X z •—< CM QC CO X QC S O 3 C3C

>.

1.

X 4-> on X 3

00 '—- 3 50 CD

O 3 O) *r" x: 50 3 •f“ 1/1 T3 > 03 X 3 3 x: 4- 3 C •t— C 3 3 >5 3 • 1— on 4- 5 3

-l-> .T3

O f—

O 03 3 CD 3 3 > 1 <T3 o on •r“ CD O E 3 > on 3

oo 4-) 3 u S- 3 C_) 3 on 03 03 3 X

CQ c- 03 Q- 03 o 3 o on Z o; T3 X —I on 5o JS X on S- >> 3 T3 3 — X

O) (T3X 3 4- t

O 3 00 50 4-

i~

C 1— > 3 SO -c •r— 3 3 3

a. S- o i- CD 03 3 X 3 E X cu OJ on 03 on X S- 3 03 03 • r— o

QC CQ IM

X <C o < <=£

Q o: QC CQ

cn CO o X CO c CM CM tX

CM +J O) . . . . . CO O) 4-J c 3 3 3 X CTi O) (O 03 03 03 03 3

s: o 3) >3) >33 3) LiJuan

Menjivar

Bethesda

Research

Laboratories

301/840-8000

biotechnology

process

8717

Grovemont

Circle

engineering

P.O.

Box

6009

Gaithersburg,

MD

20877
CD (

4-> c 00 -a 1— 00 c (U CU OJ >0

00 0 cn

u

M

u 00 00 •t— 0 c •»— 0

t/1 OJ 00 4-) r— CU r— “O •1

(n 0 00 fO 0 •t— c X

-M ra 0 a.' ^ 4-J c 0 X 03

00 E sa 00 CU 00 f— cn 00 0 . CU 0 C2. • 0 0.) E E 0 c • r 0 E 03

s- •r- 00 s_ E 0 s_ 03 OJ •1— 00 cn E E cn

O) JO -a r— Q. >0 OJ E JU> s_ I 

X c •t— 0 0

4-> 0 fO

M 00 w- 0 c OJ fO "C t— c • ^ 0 0 • n C s- •r“ O) OJ • rv c • r •r“ 0

»—I

>, • <4- •r- >0 CU O) . •- JO

X c ’ r* >0 c >) s_ cc CD C E cn > E cn 03 cn X X <4- 0 0 • r OJ 0 cn T3 c CO • . ^ cni (U 0 r— c >> u 0 r— r- >> .E (

c O) 0 0 >n c c X I— Q. r— X CU 0 -t-> cn U 0 • r— M 0 .—-

cn 0 CU

u 0 0 4-> cn ^—. c rx3 0 c S_ ’r“ 0 0 •>— c X c . c

00 sz 0 JE OJ f— 00 •r— r— 00 CO 0 X 0 X 00 CU *r“ - 0 C 0 •f" 0 a; r— 00 XI 0 s- (/) •t—• 0 S- 0 OJ s- TD O) OJ E c OJ c JO 0 c cu OJ c OJ 03 CU E C3 OJ r— +-> E fO 4-> 0 c= E -E > u fO 4-J CU 4-J >0 4-> O) 0 i_ ISI cn 0 cn E 0 0 0 c 0 cn 0 00 0 M r— S-

•r— •r* OJ

C S- •r— c E r— •1— O) 0 !_ E •r— OJ •f—

C O!

Ll_ <+- O) 0 CU JO E 4-J 0 Q. 0 X s_X O! 0

crc

C\J CO <D 0 CO

X »—1

00 oo 0 0 1—1 on cn CM X 0 •30 00

(U CM on cr<

X E 1 1 1 1 1 1 ^ 0 CO CD ch CM

T3

X onm

X CO

OJ

X CD

m

X

C CU c r— CM CM cm CM

•r— CU r—) 0 0 0 0 4-J 1— cn CD CM cn X c0 i- u <v

4-> >) E CU t-J E

1—

H

CU E E 03 . 0 E CU X

CO E 4-> E E 0 X (

X 0

CU

C •r— 0 E 0 X r— 0 s_ 4-> E CJ r** X •r“ CU 03 0 03 >,-r- CU o2

03 4-J 4-> 1-H T3

X CU E E > 1— fC E 0 E X CO 0 cn CU 00 CO N O) on E 0 (U 4-J ct 0 E E OO

C_J 0 • 0 • CM DC ro 0 =3

X c/)

C CD <c 0 U_ 3 E c*_ (iJS •>- 0 Oo

fO txj c u O

on

cr.

CO

O)

CJ o c: fc o ~i

cu +J o •>- o o

».o

C r-l CU nO

4-J

X

X rc 1— 00 c •f— o> C30 E E

03 CJ 0 rO 4J • f— t-H 0 (U •* CU 0 4-J E CU X 00 0 0 -c 00 E E X f—• S ! E 1— E •r-* E r— 4-J 0 4-J 03 1—^

O)

X <c E -CD •r—X 4-J E fO

X 00 4-> 4-J 0 1 CU 0 3 O rO cn E 4-J OJ

r 00 E 4-J CD 4-J X E CC c: C CD E OJ E Q !0 CU t—1

CU f—X

X 03 0 ‘»—X CU -f— E <4- • E 1 f— •p- 0 X 4-; E • E

T3 0 <4- 0 E s 0 0 o 4-J 0 00 E E X 0

TOX

Q • E 0 rTD CO 03 0 O) • OJ < C_J X X 0 CO 2: IT r-H 3 2: X X X ^ (T3 U +J

s_ (y> ra

<v '—

00 <T3 O) +j

QC c

OJ

s- s- -M O) C Q.

O)

X

O LxJ E

O) 2:

OJ LU o o c c o o

Q_ -•-> 3 cn Q C

O)

Sro G.

Om <U CO O '=0-

00 a)

00 -r- fO u >-

r- c/-> z: CD C «' cn fo 00 c > c co c

S-o

C_> CO C_J

cn -—,

•r— •r— 03 E

00 4-J NJ

X E CU 03 r—

—

4-J E E n— 0 X

X 03

CU CU CU CU >0 CU 0 0 C_)

X > cn E f— 4-J >» E 00 >0 E U to CU s. •f— E CU 00 CU -0 CU to {/I 0 •<- 03 2: E • CU a*

4-J

M CU •1— E s E CU E -S«C E CD 3 X 03 E X s E -r- 0 <4- 0 DT X T3 cn • • 0 4-J 03 X X 0 CU X CU 0 E -r— C_3 . 1— . X E Q CU E E X 03 •c Q r— 03 C/0 2: X • f—

CU r— 03 E E E lEI E C/0 3 to E X lO 00 CJ X <0 03 03 4-J 0 X X CU X 03 TD

CU •r~~ 03 4-J -r- ‘n~ E E C4- >— ^ . E r— E - 1— E CU E E C4- 03 -a 1— I— CU 03 >4- CU (U —J CU E CU 1— 03 E X E Cf- E CU / t X .SkC • t— X E X 03 X 1— s

CU 0 03 0 E f— -r- 0 to f— 0 0 . 0 X 0 - 1— 0 03 X s: X X X 2 3 X 0 C-J X 2: 3 X c_> X 3 X 3 X f—4

LD cno t—4 r—4

CM CO fX

CU . . • . . . 4-> X E S- E E E 03 CU 03 X X X 0 X c =£ c

Apr.

26

Zachary

S.

Wochok

Corporate

Research &

Development

314/694-8643

biotechnology;

immobilized

L.

Edward

Klein

Monsanto

Company

enzymes;

agriculture

and

F.

B.

Zienty

800

North

Lindbergh

Blvd,

plant

cell

research;

health

George

Glover

St.

Louis,

MO

63167

care

research

Ned

Seigle
and 9 of this report will describe possible NBS responses to industry needs

should the Bureau decide to develop certain capabilities in this area in

addition to its on-going competency development efforts.

.a Standards and Measurement Needs

.1 Raw Material Substrates/Feedstocks . Lignocel lulosic materials and

hydrolyzed starch are the two biomass feedstocks that are being seriously

considered as substrates in industrial processes for the production of commodity

organic chemicals. Reference standards for these and other substrates are inadequate. Usually, only a functional definition of a substrate is available.

For both of the above substrates, but especially for 1 ignocel lulose materials,

basic information for the development of standards is limited. To develop SRMs

for 1 ignocel 1 ulose and major chemical components of 1 ignocel lulose (cellulose,

hemicel lulose and lignin), these feedstocks need to be characterized on a molecular level and appropriate assay systems need to be developed. In addition, questions of 1 ignocel 1 ulose source heterogeneity, of source lot

variability, of feedstock storage, and of pretreatment and isolation methods

need to be addressed in developing SRMs. Municipal and bio-wastes will probably

not serve as feedstocks for industrial production of organic chemicals via

fermentation. However, according to some industry spokepersons , NBS experience

in developing standards in this area could assist in developing standards for

1 ignocel lulosic materials or other substrates used as feedstocks for biotechnological processes. On the other hand, some industry spokespersons indicated

that because of the unique, specific, and usually proprietary substrate/

feedstock requirements for an industrial process, feedstock standard reference

materials may not be required.

.2 Biocatalysts/ Enzymes . Most of the enzynes used in commercial processes

are obtained from microbial sources, are extracellular, and are impure. Thus, one of the major problems in the commercial use of enzymes in biotechnological

processes is that one must deal with product (enzyme) functionality rather than

with product composition and homogeneity. This problem will become even more

acute as enzyme or biocatalytic systems assume greater industrial process and

commercial significance in the future.

118
In general, buyers of commercial enzymes, with the possible exception of

those in the clinical chemistry diagnostics area, are not sophisticated

regarding the possible benefits of standard methods for measuring enzyme

activity. Although standardization of enzyme activity appears reasonable in

academic laboratories, this capability and the extensive supporting literature

has not yet been transferred effectively to the commercial sector. A possible

exception may be the producers of "enzyme tools" (restriction endonucleases,

ligases, etc.) for rDNA research.

Currently, there is considerable variation between commercial enzyme

producers not only in the methods of enzyme production (for example, different

substrates and microbial strains) to produce the same enzyme but also in the

analytic approaches to measuring enzyme activity. For the buyer of "bulk

enzymes," this makes difficult the accurate comparison of the relative cost of

enzymes of specific activities from different sources. According to some industry spokespersons, examples of enzymes where standards are needed include

carbohydrases, proteases, lipases, and pectinases.

Standards of enzyme activity would be helpful to buyers by establishing the

equivalency of products used in commerce. However, it is uncertain whether the

respective proprietary positions of enzyme producers, mostly foreign owned

companies, would limit the market impact of NBS development of standard assay

procedures and SRMs in this area.

.3 Microorganisms and Cell Cultures . Currently, the American Type Culture

Collection (ATCC) and USDA's Northern Regional Research Laboratory are major

organizations responsible for serving as repositories of microbial cultures and

for maintaining culture collections. Additional manpower and resources are required to maintain and to expand these culture collections. This will become

even more critical in the future as patented rDNA microorganisms are deposited

in these culture collections and as microbial cultures, both wild type and

genetically modified, become more important as articles of commerce.

According to industry spokespersons, the standardization and characterization of microbial strains in culture collections is vital to industry plans

for expansion into and investment in biotechnology. Some industry spokespersons

119
suggested that the responsibil ity for support of this "national resource" should

reside in the Department of Commerce, since this activity would provide generic,

infrastructure support to the new biotechnology industry.

Also, as microorgani sms become more important as articles of commerce,

buyers will need some assurance that they are obtaining not only the microbial

strain they are seeking, but also the appropriate passage of the microorganism

that will express certain well documented morphologic and biochemical characteristics, and, possibly with rDNA organisms, verifiable genetic sequences.

According to industry spokesperson, well characterized and standardized measurement and analytic methods for testing and certifying the similarities and

differences between microorganisms at the molecular level will become more

important in the future. For example, methods will be needed for evaluating the

degree of homology between gene sequences of patented rDNA procaryotic or eucaryotic cells involved in patent disputes.

In addition to microorganisms, advances in cell culture technology (animal

and plant) will provide additional industrial alternatives for the production of

organic chemicals. The establishment of cell culture repositories (cell banks),

the characterization of cell cultures in cell banks, and the on-line availability of cell culture data will assume increasing importance in industry's

plans for developments in biotechnology. Investment of resources and development of standards and of evaluated data sources in this area will also be

required

.

.4 Proteins and Nucleotides . As the biotechnology industry grows, the

commercial demand profile for specific enzymes (for example, endonucleases), for

peptides (for example, as synthetic nutrients), and for oligonucleotides (for

example, as gene sequence linkers and probes) will shift from academia to

industry. Currently, a number of manufacturers produce these products, and

little has been done in the way of developing measurement methods or standards

for assuring tbe buyer of product purity, equivalency, etc. In addition,

analytic techniques and measurement methods for assessing these newer articles

of commerce are varied. Existing techniques for comparing products from

different firms are either complex or lacking.

120
.5 Biopo'lyners/ Polysaccharides . l^ith increased interest in applications

of biotechnology, microbial biopolymers/polysaccharides will have increased

importance as articles of commerce as lubricants, food additives, surface

sealants, controlled substance release vehicles, surfactants in tertiary oil

recovery, etc. However, progress has been slow in developing measurement

methods, standards, and evaluated data sources for polysaccharides including

standarized nomenclature and analytic and measurement techniques for batch

comparisons. To overcome these barriers limiting the expanded role of

polysaccharides as articles of commerce, efforts in the following areas are needed: (a) critically evaluated information base on biopolymer/ polysaccharide

characteristics including thermodynamic properties, (b) standardized measurement

techniques; (c) tests for product purity and for certification of product

equivalency; and (d) SRMs.

.6 Public-Private Sector Discussion of and Consensus on Standards . In addition to establishing closer working relations with industry scientists and

with technical associations such as the Industrial Biotechnology Association

(IBA) and the American Society for Microbiology (ASM), it was suggested that

some other actions could be taken by the Bureau to facilitate the public-private

sector consensus process necessary for the development of effective measurement

methods and standards in biotechnology. For example, NBS and NSF could

collaborate in a workshop or a workshop series on the necessity and advisability

of developing standards for biomass or other biotechnology raw materi al s . (The

Bureau has already offered technical support on standards in this area with the

ASTM. In addition, the Bureau could offer technical support to such groups as the Enzyme Ad Hoc Committee or other voluntary standards organizations. Bureau

scientists, in cooperation with international technical organizations, have

begun discussions on approaches to developing international standards in this

area, for example, biothermodynamics —Inter Union Commission on Biothermodynamics of the IUPAC-IUB-IUPA3.)

.b Fundamental Science and Information Needs

.1 Enzymes . .a Enzyme Character ization . Industry spokespersons believed

that the Bureau's base of expertise in physical and analytic chemistry.

121
microcalorimetry, materials science, instrumentation and catalysis would be

helpful in providing a fundamental science base for better understanding of the

parameters of enzymatic reactions and of biocatalyst immobilization in

industrial biotechnological processes. The extent to which enzymes or biocatalysts are used in biotechnological processes will depend on the purity of

the enzyme and on the development of appropriate measurement methods. Examples

of types of information that would be useful to industry in characterizing

enzymes under different conditions include activation energies, molecular

conformation of the active site, molecular and thermodynamic aspects of binding

to and cleaving of substrate, kinetics, efficiency of enzyme systems, molecular

attachment to and catalytic properties in association with polymers (immobilized

systems), and evaluations of isolation and measurement methods.

In addition to benefiting existing bioprocesses, tne development of basic

information on enzymatic processes, on enzyme active sites and conformational

relationships, and on models of enzyne action will be of importance to

industry's development of modified or synthetic enzymes tailored for specific

and possibly novel industrial bioprocesses in the future. Fundamental

information on enzyme structure-function relationships will assume even greater

importance to industry in the longer-term future, since rDNA technique will be

able to modify specific enzyme molecules and, thereby, change enzyme function.

.b Enzyme Catalogue . In addition to Methods in Enzymology , an information

source or catalogue of known enzymes that would centralize critically evaluated

data on catalytic reactions; thermodynamic and kinetic reactions; enzyme

properties, activities, and characteristics; standardized assays and measurement

methods; international units for the conversion of specific substrates;

methodology to determine purity; properties of isoenzymes; enzymes requiring

cofactors, etc. would be useful in the long-term for the application of enzyme

technology to industrial biotechnological processes. Such an on-line

information system would be particularly helpful in developing predictive models

and in evaluating chemical and biological process alternatives for product

synthesis. For immobilized biocatalysts, some industry spokespersons indicated

that handbook information on diffusibi 1 ity and orientation, on rates of

reaction, on biocatalyst half-life and support systems, and on approaches for

catalyst rejuvenation would also be important for optimizing industrial

processes

.

122
.2 Fermentation . A major industrial consideration in fermentation

processes is the amount of water in the reaction vessel. Economically feasible

industrial bioprocesses for the production of organic chemicals will require

fermentation process improvements so that reactions can take place in reduced

water environments. In addition to the general lack of critically evaluated

information on reduced water and/or "solid substrate" fermentation processes,

there is also, according to industry, a general lack of information on properties of "slurry" streams of biomass substrate on fermentation process

parameters (flow characteristics, power for agitation, shear forces, mass

transport of gases, thermodynamics and product heats of formation, cell density,

foaming), as well as on subsequent separation of the product from the reaction

vessel, on cell recycle, and on biocatalyst recovery.

.3 Separation and Extraction . Bioprocesses usually involve producing

desired end-products in lower concentrations than those obtained via other

process routes. In scaling-up, substantial difficulties will be encountered in

separating desired products from the fermentation milieu. The traditional

distillation approach is quite energy intensive, and, therefore, expensive.

Thus, newer separation technologies that are amenable to the scale-up a/id other

requirements of industrial processes will have to be developed. According to

industry spokespersons, considerable input from materials science research will

be required for developing improved and durable commercial separation processes,

as well as processes that will allow fermentation process steps to be coupled to

and be compatible with chemical synthetic steps. Evaluation of newer separation

and extraction procedures will be essential prior to industry investment in

capital intensive biotechnological processes. There will be a need for accurate

data on thermophysical properties of alternative separation approaches, on phase

diagrams of constituents of mixtures, on properties of various solvents, and on predictive models of properties of separation approaches and of possible range

of applications. Membrane, liquid chromatography, super critical CO

2 were

some of the separation technologies mentioned by industry spokespersons that may

be applicable to large-scale industrial bio-processes.

123
.4 Process Monitoring and Control . Development of real-time, continuous

process monitoring and control instrumentation and probes will be important for

the expansion of industry use of biotechnological processes (for example,

continuous fermentation and cell culture). Although there are a number of

indirect approaches to measuring such process parameters as O

2

, CO

2

, pH,

viscosity, diffusivity, etc., little has been developed for the continuous

measurement of biological products and secondary met abol ites in complex

fermentation broths, for monitoring microbial cells, and for monitoring plant

and animal cell cultures. Examples of instrumentation development needs

mentioned by industry spokespersons include: (a) non-specific biomass sensors,

(b) substrate sensors with high sensitivity and rapid response times, (c) and

sensors that can measure production of a specific chemical in a complex reaction

environment

.

.5 Chemical Engineering . In addition to the specific chemical engineering

examples cited above, general experience in chemical engineering will be

essential to the successful scale-up of industrial biotechnolgoical processes.

According to industry spokespersons technological improvements in biotechnological process design and control will be important to the economic feasibility

of bioprocess approaches for the production of organic chemicals. A fundamental

understanding of critical scale-up parameters, of the kinetics of production,

and of fermentation processes will be required.

124
7. NBS RESEARCH CAPABILITIES RELATED TO BIOTECHNOLOGY

With the possible exception of activities in (1) clinical chemistry, (2)

analytical measurements, and (3)denta1 biomaterials development and evaluation,

NBS is not involved in areas specifically related to biotechnology . However,

the Bureau does have fundamental expertise and capabilities in the non- biological sciences that may relate to long-range industry infrastructure

support needs in biotechnology.

To determine what this expertise is and where these capabilities reside in

the Bureau, an assessment of current NBS research activities was undertaken.

The objective was to identify areas of fundamental research strength and

traditional NBS missions and roles that could possibly relate to industry needs

in biotechnology. Table 7.1 summarizes the results of this assessment.

Biotechnology-related research capabilities are dispersed throughout the

technical centers of the Bureau. Organizational ly, the National Measurement

Laboratory (NML) contains most of the Bureau's fundamental research capabilities

in the non-biological sciences that potentially relate to: (a) future industry

needs in biotechnology, and (b) the development of a core of biotechnological

expertise. Specifically, the Centers for Chemical Physics, for Analytical

Chemistry, and for Materials Science provide a base of scientific, measurement

and standards expertise in physical chemistry, organic chemistry, analytical

chemistry, polymer sciences and related fundamental disciples, as well as instrumentation technology that appears, based upon discussions with industry

(Section 6), to be particularly important in industrial biotechnology process

development

.

Potential also exists within the National Engineering Laboratory (NEL),

specifically the Center for Chemical Engineering (CCE), to transfer to

industrial biotechnology problems some of the expertise they have and are acquiring in flow dynamics, modeling of unit operations, slurry and continuous

process technology, process control sensing systems, and separation technology.

Examples of industrial problems relevant to CCE expertise include: (a) characterization of and process engineering design requirements for slurries of

biomass substrate, and (b) measurement and evaluation of alternative extraction

125
o — o 2: I i 3 3 0 O Q) 3* I 0) ~ I 3 O —

Qt

W

H> (9 “2 2, ^ ^ o 3 0-0 ® ® "! ? 2 8 o — ®

g- s ^ — 3 o o

Vi (C —o O Q> il— O o n ^8§

-!o ^

S'

® Q) *0 3

a- 0) «< — - *<

(A o —

-t, tfl S s

•n Q.

-f—

n

O 3* ® -1 v> a

•• o

-+• ® 5- i

i^ ?!

CO 3 — w O ® ® -» — 3 ® ® 0. t o S 3 ® — O — — 3 O — -o39 O

—

(A (A O I i — *0 3* 3*

(A

— (A 2. ^ -S — O 03—1 » O -f (9. O o

(A oS ^08 *1 ® *0

•+

— •< 9 g

X3 “ o^ c <A (A ® O

CA

—

J ^ ^ o — 0 1 o 1 - = o o o m — 3 s o» 3* 3 O S ® O ® N 3 -f ® o T •< % -? CA o g i - o ^ s

-+•

®

®

®

®

® *0 (A (A 3 o

-I “J

® • ® < 3. o5

i. =? ® •<

®—

® 3 —®

9r

a 3 — - © a o n O — -f © ^ a c O a ® — — 5 -1 » CO ® ’*•' 2 • © 3

CL

O3 O I s =

-4> 3 T ®

2. 5! 2,«-+ 3 o & ® c —8 3. i n ® ®

® 0

-f o ® ® c n — o* o ® o

3* tA 3*

—

® COt § -f.

O-> 3 CD

®

O3* 2

S- ^

Of® 3 -N X s 3 ® C CO -f a. ® ® '< ^ o ' S' 8 ^ ®3® ® — 3 • o -+

O'

O 3 ^ g

D. 3 — O ® -f

CO

—< o 3

CA

—

-f "< — CO S 3 3 3 - - — O — 3 3 *0 CO 3* o o . 8S

o. . « — o << ® 3 CA

CA

—

—

®

C CA 7 33 — 3 ® — < »< ® (0

(O

—' —

S " 3 (A J s*o I— 8

*o c2 0

:o0 i 0Q Z® s? 2? I3C 0®c 0

S’

X03

*8

aN

®

s- <

O'8 T3®n 3®1 1 8 2 CO a —

-<

® > > :» 2 T O ® CD O -f <A </» 3 -> ® I CA < 3 <

-I

—

O

®

Q 3 3 -» "O 3 ® “! -f *< ® omzm2o

3J5 = ?2n — ©

-f— CD OS

u) ;d® O CO ^ T3 LA 3 A •< O (A ^ O > 3- 3 o i 2- ^ CA 2 » 2. 5 o • — m o 3 3* o I m5sm2zo — 3 ® O’ ^ Z®

*0a r' ® — 3

NATIONAL

MEASUREMENT

LABORAT(RY

NATICH^L
NATI(>^L

MEASUREMENT

LABORATCRY

NATIONM

ENGINEERING

NBS-wlde

Abs.

LABORATORY

Research

NBS

GENERIC

RESEARCH

CAPABILITIES

Phy.

Rad.

Cham.

Anal.

Matl

.

Activity

Complementary

to

Core

Bl

od’echnology

,

Qiant.

Res.

Phys.

Chem.

Scl.

Chemical

o

o

• ^

o

3

s t

t s -P u 0

* o g O O to •r ^ t-

•o c C lA 3 a c O 2

^

>

X X X X

— m

— O) o ® ^53

^ s - 0 3 D <0 .5 C —

V' c O

(9 iCI ^ C

Ct 7. ^ X

O)cX

e

o

.C —' —

I 5 I

S S' — X

lA (0 —

— U W1 •— .O >0 Q < S

i o

0 5j ^

E O0 g — lA D O (A3 i s O c

L. * 9 8 8

£ <A U .8 c o> 2

>.

lA *0 «A 0

- 1 O w 0 V. Ov

^ e >* w — «> U L. 0 O ® t lA u — i= v> (D o

E ® g C <A 0 c o ® o >» 6

— C .C .c £

X O O h- Q.

y 3 — u

§

-oo < z
NATIONS

MEASUREMENT

LABORATCRY

NATIONAL

ENGINEERING tc 3 I § § *CT -J U UJ

• o

o

i/i w

'a §

c ^ . < o w

U» O <0 a

c p

SO w

(/> 8

* UJ— o t 5 -j <j « St;:

S 5 J2 ^ ® l:

2 o O H co

>*u o

— 1_ —

» (O > </)

I ® — ID

</) u> 4. Q S 9 u V. 2 ir < <

Ui t f

o S 5

s I

9

<0 1.

4-

1j I/) Z *i IIN

C —

9 9O

9 ^

> O 9 U O <Q

C E — 9 — £ o o

Si

3 _

4- 00

® s.U 4- O 9

e

1.

tn o Z i/j 5

9§

^ 9 t i/t O —

9 — ^

9 E \ E 9 C £ O

9 U —

y —

o -o

«} 9

4- C l- — o O)

4- — 9

s s ^

o» — ^

- u O

9 +- —

B 2. 9 <.0

^ S

z i3

U 9 9

0 0 4-

O 9 — — o *0 ^

tf> w ^

s ^

^ ^

°

4- c c «n

ui 9 o e 9

>» i — o — ul 9 4- — 4- W 9 WJ U

3 A 3 9 9 s 5. n 9 u o

O 9 9 — u

E z ^ *0 O.

e— (/I u —

9 <j> 4- >* 3 S

</) <

9 « N € 8, 111

I CTO O) S £ :=

o

L. 4-S

u^ 1 = — 9 —

>

>» 9 ^ U A ^

9 1- —

5 i — >- — *0

E E

>. u — 9 O j=

a. 4-

r 8

®

-g

VI 9 9^ u) 0. 9

O

I 9 —

> U 4-

O

c

O --

(A —

9 9

9 CO S !D

X XXX

9 4- 9 4- C — 9

9 .* (A % » c e 9 c

» 9 — — — o ^ T? > > o. w, £ O u u 9 u

(T 5 3 3 J) 5

g* £

— 9

i 3 5 -

>• — w 9

<C lA

>*- T3 — O

-0 £

s i

— w 3 c

9 (A O ^9 3 V) t t

4- 9 2 8- — Q. ^ ^ 99 —

<AO— <A E 9

>. —

I 2

c tj 9 9 £ —

4- 9 o <n o o

9 w

u

9 9
•

*Post-doctorB I

**Based

upon

task

Information

s/stem

keywords,

No

tasks

categorized

as

07

(biotechnology). f :: I ::: a c

'+ 3 ^ o’

2. ! X CL. — ® 3 —

(O

—1 i ^ ? 2. z X O — 3" S O O o I » o Jo A 3

CO (/) 2. s I i

-fO Cu Qt 2

U)

® *<

——

-f

n o o w — — a :j

Q>

—

— \ O

® CO 0 o 3- •

Q»3 •• 1 §

U1 ot

(/)

V) i s i 1 w ® c

o»n o ^ 3^0o :i 8 O Q O

2L -8

X

— « ® o 3 3 (A T3 *3

S’ ? ^ » •+

V)

® 8 I

2.

a. 3 ® *< (O

C

— 3 ^ (fl o

“a

® < 2 i 3 T 2. 8 I 2.

(A 3 > > 33 2 "» O ® CD ® - V» to ® — ® I V) < ® < ? S 3 V)

*o 3 Of O T - '< 0s ? = ? — ®

**

TMa

g < ® — ® o r“ 3' — §— m O CO oS O 3§ 2. S 2. m O 3 IT £. § ^ 8

..

n >

CO52mz H m iio o

Table

7.1

(continued)
techniques for the separation of specific biological molecules from relatively

aqueous reaction mixtures. In addition to the CCE, the Center for Electronics

and Electrical Engineering has expertise that could contribute to bioprocess

sensing systems and to the on-line electronic control of industrial

biotechnological processes.

Although the proceeding discussion illustrates that the Bureau has several

bases of technical strength that could relate to industry needs in biotechnology, it is also apparent that there are a number of gaps in the Bureau's

fundamental capabilities related to biotechnology.

Should the Bureau decide to invest its resources to provide infrastructure

support to industrial applications of biotechnology, in addition to on-going

competency development efforts, NBS needs to acquire or develop scientific

capabilities in the following areas: enzymology, molecular biology/biophysics,

micrcbi al/cel 1 biochemistry, and bioprocess engineering. In order to effectively link existing in-house Bureau scientific expertise and capabilities with

future biotechnology needs of industry related to the Bureau's mission, the

Bureau will need to begin developing, over the next few years, core expertise in

fundamental biosciences. This will allow the Bureau to: (a) sustain the

dialogue that has already begun with industry in biotechnology, (b) accurately

and effectively tailor Bureau responses to industry needs, and (c) develop the core science base for collaboration with and, to some degree, aggregating

existing Bureau research capabilities related to biotechnology. This competency

development process will also provide the in-house technical credibility

necessary for a sustained Bureau presence in this area.
8. DEVELOPMENT OF NBS KNOWLEDGE BASE AND CAPABILITIES RELATED TO LONG-TERM

INDUSTRY NEEDS IN BIOTECHNOLOGY

In this section of the report, NBS scientists and technical staff* identify

nine possible NBS research initiatives and services that might be undertaken to

provide infrastructure support for long-term industry needs in biotechnology.

In general, these technical descriptions by NBS staff focus on two areas: (a)

fundamental science (knowledge development) opportunities and possible roles for

NBS; and (b) possible areas where NBS knowledge base and services could be

applied to long-term industry needs in biotechnology as these needs relate to

the Bureau's mission.

The identification of these technical areas was influenced by industry's

perceived needs in biotechnology in relation to the mission of the Bureau

(Section 6), and by existing Bureau scientific and service capabilities

(Section 7). Bureau scientific staff in providing input to this section of the

report were influenced not only by industry needs and by the scientific opportunities in areas related to industry neeos, but also by Bureau resource limitations and competing demands on existing resources. The summation of these

influences is reflected in a cautiously optimistic assessment by Bureau

scientific and technical staff.

It is emphasized that the contributions which follow are only examples of a range of possible technical areas where NBS scientific and technical staff might

contribute to meet long-term industry infrastructure support needs in biotechnology. Further, these technical descriptions by Bureau staff represent a cross-sectional view of Bureau responses to industry needs for applying the

outputs of a dynamic technology. Only through continued interactions between

Bureau scientists and industry researchers will industry be better able to

articulate its measurement infrastructure support requirements in biotechnology,

and NBS be better able to define its role and tailor its response in this area.

* Stanley Abramowitz, Robert Alverez, Sherman Fivozinsky, Ruth Haines, John

Herron, Morris Krauss, David Lide, Cedric Powell, Stanley Rasberry, William

Reed, Dennis Reeder, Leslie Smith, and George Uriano.

131
Decisions to allocate Bureau resources to any of the following technical areas must, of course, be made through the Bureau's competitive progran and budget

process where comparison with other priority areas can occur.

.1 Predictive Models

The structure of bio-organic molecules, such as the active site of enzymes,

might be analyzed and even predicted by various molecular quantum mechanics

models (306). Ab initio quantum mechanical methods are still not adequate for

such large systems (>100 atoms), but the ab initio methods are used to determine

the interaction energy parameters and to investigate the reaction mechanism by

studying model systems (307). A vast amount of calculating has been done with

emphasis on hydrogen-bonding interactions. There is still a need for an accurate yet inexpensive method for calculating the van der Waals interactions.

The inclusion of pseudopotentials permits the inclusion of heavy atoms which

play an important role at the active site of many enzymes. Accurate pseudopotentials that also include relativistic efforts were recently developed and

their use will certainly become more widespread.

Using the hydrolysis of polysaccharides by lysozyme or the hydration of

CO

2 by carbonic anhydrase as guiding reactions, quantum mechanical methods

could be developed at NBS for applications to the following problems:

0 Determination of the energy surface describing the transition complex for

reaction at the active site of an enzyme by parametri zi ng the ab initio

energy surface for models of the active site and incorporating parametric

representation into classical methods of calculating the conformations of

ground state enzymes in order to extend methods to the reaction complex.

0 Determination of the effect of solvation on changing the character of a gas

phase reaction by using the multi-configuration self-consistent-field

method including analytic evaluation of gradients with particular attention

given to (a) the "catalytic" effect of metal ions, and (b) systematic

calculation of reaction surfaces for classical organic reactions to provide

background for catalytic calculations.

132
0 Critical analysis and evaluation of present methods of calculating conformations, energies, and spectra of intermediates important in those enzymatic reactions that possibly will be of industrial process significance.

0 Development of new methods for calculating the van der Waals interaction

between molecules.

Development of these fundamental research capabilities and data resource

within the Bureau will allow, in the long-term, for the exploitation of

predictive models in evaluating on a molecular level biocatalytic reactions that

might be useful in industrial biotechnological processes.

.2 Microanalysis of Complex Mixtures in Bioprocess

Use of highly sensitive analytical techniques utilizing enzymes as reagents

has been a slowly developing field until the past four years when the techniques

of immobilization of enzymes began to be used extensively (230, 308, 309).

Enzymes that were normally free or water-soluble were immobilized or made

insoluble by either entrapment in a matrix or by covalent bonding of the enzyme

to a solid phase. Enzymes thus bound showed retention of activity for long

periods of time. For example, a single sample of immobilized glucose oxidase

has been used for several thousand determinations of glucose in samples.

Widespread use of immobilized enzymes and a proliferation of automated

instruments that utilize this technology are expected to be of particular

importance in the field of clinical chemistry in the next five to ten years

(310, 311). Limitations and technological barriers are mainly related to the

understanding of the mechanisms of optimal binding of the enzyme to solid phase

supports and to practical ways of further protecting the activity of the bound

enzyme.

The role of NBS relative to this technology could be several fold. First,

from an analytical standpoint, gaining familiarity with immobilized enzymes as analytical reagents is an imperative. One cannot understand the problems of the

133
technology until an intimate working knowledge of the strengths and pitfalls of

the use of these reagents is gained. Second, improvements in reagent sensitivity and selectivity can be made if fundamental knowledge is gained on the

mechanism of enzyme-substrate interactions, the conformation of the enzyme

active site, and the proper amino acid residues that may be used in covalent

binding for optimal preservation of activity.

Of the current applications of immobilized enzynes, one of the most

interesting is in the use of enzyme electrodes. The coating of electrodes with

an organic polymer containing immobilized enzymes yields an electrochemical

sensor that may be useful in applications where sample preparation has been a major problem. By eliminating extraction and clean-up steps, better analyses

will be feasible.

Development of more reliable sensors, understanding conditions for

immobilizing enzymes, and providing materials definition for reagents required

in the binding steps are all applications that will be needed by industry in a variety of biotechnological applications.

.3 Thermodynamics of Bioprocesses

One of the fundamental questions faced in all chemical processing is that

of knowing the energetics of the chemical reactions involved. Traditional work

in thermodynamics has focused on hydrocarbons, inorganic materials and aqueous

solutions of simple electrolytes. Bioprocesses on the other hand involve

complex molecule and electrolyte interactions. Today there are several

calorimetric, theoretical, and separation techniques available which can be

successful ly modified and used to obtain the thermodynamic properties important

in bioprocesses. Four of these are summarized here.

Biological calorimetry is presentl y mostly involved with scanning

calorimetric studies of reactions. These measurements are made on complex

reacting systems and do not in general yield precise and accurate heats of

formation (A^pH) of biological substances (312). Precise and accurate of

134
biological materials are needed. In particular measurements of heats of

combustion and heats of solution (d^H) are needed so that heats of

formation are available for the aqueous phase. Key measurements of purine and

pyrimidine bases and nucleosides could be made at NBS so that correlation

methods for estimating these parameters could be developed. Similar

correlations could be made for heat capacities (Cp) and entropy (S) so that

equilibria could be predicted for various temperatures. This activity once

developed could be coordinated with a data evaluation effort to extend the 1971

NBS compilation of thermodynamic properties of compounds containing carbon,

hydrogen, nitrogen, oxygen, phosphorus and sulfur.

During the last decade NBS developed state-of-the-art apparatus for

microcalorimetry. These techniques were applied to problems as diverse as heats

of cell division, heat output due to the internal resistance of batteries, and

enzyme catalyzed reactions of serum. Today microcal orimetry is being used for some biothermodynamic studies. Equilibria of enzyme catalyzed reactions could

be studied at NBS by using the methods of microcalorimetry. This technique

could also be extended to the measurement of rates of enzyme catalyzed

reactions

.

With the advent of gas chromatography (GC) and high pressure liquid

chromatography (HPLC) the solubility and partition coefficients of trace

organics in the aqueous and condensed phases can be measured. In other programs

at NBS these techniques have been developed for environmental concerns. These

techniques could now be extended by NBS to biological systems. Measurements

made at other than room temperature will allow the determination of heats of

solution by this method, an important quantity for industrial biotechnological

process design. Comparison with established (time consuming) calorimetric

studies could be made in order to develop rapid techniques for measuring heats

of solution.

Gibbs Energy maps are of importance in determining product yields for

biochemical reactions. These maps are contour diagrams which show the

composition of complex mixtures as a function of pH and metal ion concentration.

To date all these maps have been produced assuming ideal solutions (313, 314).

135
Using expertise already established at the Bureau in the aqueous thermodynainics

of electrolytes and coupling this expertise with recent developments at NBS on

an equilibrium approach to the computation of activity coefficients, thermodynamic properties and compositions of complex mixtures could be constructed

using these maps to more accurately reflect the non-ideal solutions found in

biochemical systems. Specifically, the maps for the several energy transfer

reactions of importance to biochemical systems are among the ones which could be

constructed.

This information base would serve as a valuable resource to industry

applications in biotechnology by allowing development of predictive models of

alternative biotechnological approaches and by improving industry predictive

capabilities on process economics.

.4 Chemical Kinetics of Enzyme Reactions

The chemical kinetics of enzyme reactions is a well developed discipline in

terms of measuring overall enzyme rate constants (or enzyme activity) and

determining the effect of pH and temperature on reaction processes.

Studies of the detail of the processes, for example the nature of the

transient intermediates or specific configuration transformat ions at the enzyme

catalytic sites, are made extremely difficult by the requirement that we monitor

what is happening in some small isolated part of an extremely large molecule

possessing thousands of individual bonds. There is thus a requirement for

si te- specificity and transient analysis capability.

At the forefront of the methods being applied to this problem is resonance

Raman spectroscopy which has been successfully used to detect enzyme-substrate

transient species, and which has great promise for carrying out time resolved

studies on these transient intermediates (315, 316). Another method which has

potential is the method of extended X-ray emission fine structure spectroscopy

(EXEFS). Extended X-ray absorption fine structure (EXAFS) spectroscopy has

already been applied to biochemical systems (317). It provides information on the structure of the enzyme system. EXEFS provides more direct information on

136
the environment of the valence electrons and thus potentially provides direct

chemical information concerning the active site.

A third method potentially applicable to enzyme systems is the laser

induced lanthanide ion luminescence probe technique (318). This approach is

applicable to systems containing metal ions and involves substitutions by the

lanthanide ion and its laser induces luminescence. The luminescence spectra

provide information on the nature of metal -coordi nated water molecules,

characterizat ion of bonding sites, and other data on enzyme-substrate

interations

.

Strengthening existing NBS competences to apply and to extend these

techniques and to provide time-resolved site-specific chemical information on enzyme-substrate interactions will be a valuable resource for industry to draw

on in optimizing biotechnological process. As an initial step in enhancing NBS

capabilities in this area, laser enhanced Raman spectroscopy and the EXEFS

methods could be used to study systems ranging from simple organic compounds to

enzyme catalyzed reactions.

Along with the development of experimental strategies, the Bureau could

begin an overall evaluation of the status of chemical kinetic data compilation

needs in this area.

.5 Surface and Interface Characterization

The characterization methods of surface science are being widely utilized

for a large variety of problems of both scientific and technical importance

(319). Surface analysis, in particular, is now an integral part of many

technologies and industries (e.g., catalysis, coatings, corrosion, semiconductor

devices, computer, automobile, communications) for many different applications

(e.g., failure analysis, quality control, process and device development).

Surface properties and processes are similarly important in many areas of public

concern (e.g., defense, energy, health, environment).

137
Surface analysis has been used in recent years to characterize chemical

treatments of teeth and wool fibers; similarly, surface analysis has been

utilized for assessments of implant materials (e.g., the presence of toxic

metals on the surfaces of plastics) and for determining corrosion mechanisms and

the durability of implanted prostheses. It is believed that these types of

applications will grown and be extended to a wider range of materials (e.g.,

composites, rubbers, adhesives, plastics), properties (microbial corrosion,

stress corrosion, biocompatibility, toxicity, adhesion, wear, immunological

response), and processes ( compatabi 1 i ty, controlled porosity, artificial

organs)

.

There is a growing interest and demand for improved methods of interface

characterization (320). For many scientific and technical applications

including those listed above, it is desirable to measure the composition and

other properties of solid-solid, solid-liquid, and solid-gas interfaces with the

sensitivity and specificity now available for surface characteriztion (i.e.,

usually a solid-vacuum interface). Ideally, interface characterizations should

be obtained in situ or non-destruct ively as artifacts can frequently occur when one phase is removed to expose the interface region.

Bureau expertise in surface and interface characterization techniques could

be applied to several problems in industrial applications of biotechnology. One area where surface and interface effects are critical is bioattachment. A

variety of physical, chemical, and biological methods are being applied to study

the attachment of bacteria to solid surfaces, molecular film adsorption on solids from a bacterial culture, adhesion of cells to collagen and plastics, and

the adhesion of proteins to polymers. Development of new and evaluation of

existing methods for their industrial process applicability appears essential.

Another area of high potential impact is the characterization of lipid

membranes. Theory has been developed recently to describe the interaction of van der Vjaals molecules with dielectric films. From this it might be possible

to determine how the transport of macromolecules through membranes could be

controlled and/or facilitated. This could in turn lead to improvements in the

separation of specific biological molecules from complex reaction mixtures.

138
.6 Separation and Analytic Techniques

The main method currently being used in biotechnology for separating and

analyzing protein ana recombinant DNA fractions is gel electrophoresis and

associated techniques. Recent developments in this area have created a great

deal of interest in establishing data ban<s based on the information that can be

derived from these separations.

Examples of recent developments in this field include the reporting of new staining methods that have extended the sensitivity of the analytical gel

techniques to detect proteins and peptides at sub-nanogram quantities, with

detection limits nearing that of autoradiography. Better computer algorithms

and advanced concepts of graphic display have helped to handle the large amount

of data generated by gel electrophoresis, particularly two-dimensional methods.

Future developments in analytical gel technology may include the interfacing of high-performance liquid chromatography (HPLC) to analytical gel

separation and detection. The newer HPLC gel-filtration columns have

capabilities of performing initial separations of proteins. Further separation

of components in HPLC peaks by the combination of isoelectric focusing and

polyacryl amide gel electrophoresis in the presence of a detergent, should

provide another order of separation, characterization and measurement of

important biomolecules.

NBS has a possible role in the research and development of these

techniques. Capabilities are being developed in order to establish protein

marker compounds that will help standardize the two-dimensional electrophoresis

technique. Use of HPLC in the materials definition phase should provide an additional dimension to the separations procedures.

Application of these results could provide a basis for standardi zation of

results between laboratories involved in biotechnology-related research

activities. Extension of the data to include sequencing of recombinant DNA

would require minor reconfiguration of the electrophoretic equipment and

development of staining methods to detect DNA/nucleot ide patterns.

139
.7 Separation Technologies -- Membranes

Efficient separation techniques developed for industrial biotechnological

processes will be critical to the economic success of these processes. Exisitng

separation methods will have to be tailored to the separation requirements of

large-scale industrial process (for example, the production of commodity organic

chemicals via biotechnological processes, or the coupling of biological and

chemical synthetic process steps). Membrane separation methods are attractive

for industrial processes of this type, and advances in membrane technology will

have a significant influence on the development and implementation of commercial

processes

.

NBS has considerable expertise and fundamental research capabilities in

polymer science, microscopy, structure determinations in polymer systems, and

synthetic polymers. A possible opportunity for the Bureau could be to use this

base of expertise for developing a fundamental understanding of membrane

measurement and characterization techniques in aqueous solutions in order to

better understand the effect of processing variables on membrane structure and

performance. Information generated in these areas would begin to provide a scientific basis to otherwise empirical approaches in membrane separation

technology.

In addition to these approaches, NBS could develop fundamental research

capabilities for studying; (a) the permeability of polymer films, (b) the

molecular structure and morphology of membrane structures, (c) membrane

selectivity and performance, (d) aging processes in membranes, and (e) the

interrelationships between these research areas.

Development of information in this area would be of particular interest to

industry in evaluating separation technology alternatives for and in the design

of biotechnological processes.

140
.8 Standard Reference Materials (SRMs)

If past NBS experience with developments in other emerging technologies

(for example, analytical chemistry) and industries of national importance (for

example, health care-clinical laboratory testing) serves as a reliable guide,

the need for Standard Reference Materials (SRMs) in the industrial applications

of biotechnology is certain.

Because of the dynamic state of this technology and of the rapidity of

technological advances in this area, it is not clear at this time what specific

SRMs should be developed to meet the long-term needs of industry in its

application of biotechnologies. To a considerable extent. Bureau development of

SRMs in this area will depend upon development of Bureau scientific and

technical capabilities and expertise in areas related to biotechnology, as well

as upon outside scientific expertise to guide SRM program efforts and resource

investment decisions.

Several initial steps, however, could be undertaken by the Bureau in order

to identify and evaluate industry's specific needs for SRMs in biotechnology.

These could include:

0 an education effort by NBS with prospective industry clients to explain the

need for accurate and reliable measurements and SRMs as one vehicle for

improving industry process operating efficiency and also for improving

quality control over intermediate and/or final products;

0 encouragement by NBS for industry participation in setting SRM planning

priorities in biotechnology; and

0 evaluation of the applicability/compatibility of existing (or currently

planned) Bureau SRMs with measurement technology needs of industrial

bioprocesses (for example, temperature; calorimetry; pH, basimetry,

acidimetry; sugars; CO^ and O2 off-gas analysis; rheology; and light

scattering)

.

141
In the longer-term, a compatibility must exist between the measurement

technology and the SRMs that are developed in biotechnology. Since measurement

technology specifically applicable to industrial process applications in

biotechnology is either lacking or only in its formative stages, SRM development

must follow development in measurement technology areas. Specific industrial

biotechnology SRM needs will become more apparent when: (a) measurement systems

needs to be "functional", (b) the optimization of process parameters becomes

critical to efficiencies and yields, and (c) well-defined trade specifications

are developed for end products as commodities or articles of commerce. However,

some possible targets of opportunity for Bureau SRM development could include

SRMs applicable to the following:

0 measurement of the initial and half-life activity of enzymes and enzyme

systems used in industrial biotechnological process;

0 determining the "dimensional specificity" of industrial bioprocess

filtration techniques using aerosal/part iculate dimensional metrology;

0 development of standard feedstocks/substrates or of organic constituents of

substrates used in industrial bioprocesses;

0 development of standard reaction pre-tests;

0 determining the purity of products (for example, proteins, peptides, and

polysaccharides) from biotechnological processes and quantifying impurities

within products; and

0 filling in the existing gaps for thermometry SRMs covering temperatures at

and just above room temperature (for example, 3-5 new fixed point SRMs

might be developed covering the range from 29.77°C, the existing gallium

melting point SRM, up to about 85°C; two SRMs are currently under

development that would provide fixed points in this range).

142
.9 Standard Reference Data (SRD)

As industrial biotechnological processes become more operational, the need

for critically evaluated data will be essential in order to predict the

effectiveness and the efficiency of the processes.

The Bureau’s Standard Reference Data Program supports a number of major

data base development activities which could match expanding industrial

biotechnology concerns (321). Existing programs in chemical thermodynamics,

chemical kinetics, transport properties, phase diagrams, as well as other areas,

could be expanded to supply important input to the industrial use of

bioprocesses

.

The SRD program would significantly expand its substance coverage to enable

response to information requirements on the complex chemicals and processes

associated with projected industrial applications in biotechnology. SRD outputs

could consist of compilations of selected properties of important pure

substances, predictive methods for properties of mixtures and other

hard-to-measure data, and computer-readable data bases for use in industrial

bioprocess models.

In tne long-term, as industrial bioprocesses are further developed and

biological mechanisms become well characterized, the SRD Program might expand

its scope to include evaluation and compilation of more purely biological data.

The temporal progression of an SRD activity directed at industrial

biotechnology evaluated information needs might be viewed in three steps:

0 Initially, selected compilations could provide evaluated data in the

traditional areas of SRD expertise which overlap important biotechnology

needs. Rate consiants of enzymatically-controlled reactions and thermal

and transport properties are examples.

0 Predictive methods could be constructed to provide needed properties which

have not been measured over the required range of process parameters; for

143
example, the thermodynamic properties of substrates for

enzymat ical ly-control led reactions and properties of the mixtures found in

process streams.

0 Comprehensive computer-readable data bases could be made available for use

in major industrial bioprocess predictive models.
9. NBS ROLE IN SUPPORT OF INDUSTRY NEEDS IN BIOTECHNOLOGY

This section of the report outlines a number of steps the Bureau could

take to provide infrastructure support to industry needs in biotechnology. The

selection of these steps for possible Bureau action takes into consideration

biotechnology developments and trends (Section 5), industry perception of needs

in relation to NBS's mission (Section 6), existing Bureau capabilities (Section

7), and Bureau scientific and technical staff ideas on research initiatives and

services that could be undertaken or developed to provide infrastructure suppori

for industry needs in biotechnology (Section 8).

Table 9.1 provides a framework for possible Bureau action and identifies a range of scientific, measurement and standards opportunities for NBS that are specifically related to long-term industry needs in biotechnology. To implement

the action items identified in Table 9.1, considerable, but not exclusive,

reliance would be placed on expanding and/or refocusing existing Bureau

capabilities in physics, chemistry, material s/polymer science, chemical

engineering, measurement techniques and instrumentation, standards, and

evaluation and management of technical information. To complement existing

Bureau capabilities in the short-term, staff expertise in enzymology would be

required. In the intermediate to longer-term, expertise in molecular biology/

biophysics and microbial/cell biochemistry and eventually in bioprccess

engineering would be required.

To effectively address many of the problems related to industry needs in

biotechnology, interdisciplinary scientific approaches would be necessary.

Should the Bureau decide to implement part or all of the framework illustrated

in Table 9.1, Bureau management would have to provide a climate that would

encourage interdisciplinary cooperation and collaboration among Bureau

scientists and technical staff.

145
tn to

(0

m

O 6 1£ to u fA U) s

j) £ <J to

U) Q. "O £ 0 0 0 0

u> 4- “—

u 4- L. 4-

u

*o

Cu 0 ^ c U

A 3 0 o 1_ 0 >- 0 +- >. o 0 >• U "O 0 B h. 0

t.

b0)

A 4- U) o u <D *oc o ^ s ® 3 0

(/) s c

4- 0

0) i E o 0 3 0 E

i. to c *o 4- !o

•H 3 -o ><- Q. 0 0 0 $ 0 *o iS S' C s U) +. C £o 0 £0 Oc 0c 00v 3 *0 0 U e U1 (J o

h.

jC 0 > >- 0 o T > V. •• L. S’ 9 4- o U) cn u 0 4- 4- 0 U1 o o c O 0 0 (0 C —> 0 0 0o k. k L, 0 § cO 0 "o k 0 3 — *o £ c *— o 0 Cl 4— E

CL > C

•H •** « *+• 0 — 4- 0 E 0 u Q. ^ A c

§.

O *Q

OO to “O 0 0 0 0 u 0 o c 1. 4- o. ^ 4“ 0 X3 0 t 0 0 0 0 0 a. CL c o -H 0 c MTD — $ if o

(/) c © ^

>• tA N ^ C 3 © U>

to c

to

22

to i 0 8 0 © 3 4“ 0 B. y

(J s O'

u 0 c — I C •* MB •— 3 0 0 o •B 1 4- A 4- -o c 8 "O CL b. O) 0 >- © — — * 4* CL E

(0 *o 4- E (0 V. u L. — O (Cc 0 £ 0 o 0 0 Q. 0 u 5 > — c L. b

4-A 4- L. MB 0 0 4- MB

CL _0 0 £ 0 C £ 0 0 4- <0 to w# 4= C 4- C £ 0 0 s- MB >. a.

C tj

4- 8- 0 0 o >-

Q <0 £ L. c L- to 0 <0 0 O

®

>- 10 0 4- L.

N t_ >B S’

MB 4“ $ 0 o S A

—

4- t.

U t. 1 &

4-

C 2 u to 3 C 0 5 0

s.

U t 0U 0 O

to

a

u

a.

u 3

T3

O'

y0 O)

CE O (0 to ® 0 0 § 4- O) X 0 C c 0 >4— u 4- 0

•t-i- 0£ A *0

to 4-

— 0 c 0 c0 c MB -Cy

L.a. 0 0c 0 S c0

Q. L.

tO s

4- *o

——

S’ 0 a. to LU 4- 0 L. 0 0 0 4- B. tO 0 4- CL 4- 4* 0 0 0 u

-i

4- 5- UJ 4- 0 0 E

MB 4* 0 MB B* BB (0 BM (0 *o 0 o to 22 c © 0 o — 43 X 0 3 y 0 © 0 MB t

BM 0 -o

to < CX 3 o t t 0 0

h)

B. (0 c > L. 0 o 0 £ g — < 4- •— 4- U 0 S

JZ •B> 3 y

a

MB

y

X

*D

Q §

LU 0 to

O 0 B. 1 0 > •o

y t

4* o 4- 4- 3

t. LU >- > O) c 4-

h. MB 0 — 4- 0 u 4- 0 X 4- 0 0

*£ 0 B 0 Q LU 0 0 L> 4= CL o C 0 0 4- MB MB 0 k C 0 4-

•o Z H*

m 0 £ to a. lO 0 4- MM 0 *o £ U 0 MB E Q c < LU 1 "O 2 4- 4- -o 0 44 C o MB 0 o <0 £ 0

S’ to

CM c0 U o

to

C

u

C0 0 0u

c:0 0”0 s

43

h.

O 0

iC 0>

a.£ o E 3£

to

— 1 — 0 CM

2.

to c 0 0 (0 4- •B 0 A !E •B

-o 0c & 3 & 0b 0 £ CL

a 0

£14- 0 0 8- 00 0 t;

X0

8-

X0 c O • MB C 0 o to to MB •— C MB n 0 — E 0 o O 0 0 o> 0 to c 4- 0 u 0 "8 c 0 t

MB -o L. 0 4- 0 < > • > 0 o 4- > 4- 4- o > ««B 0 0 4“ C > “O • » 4- 0 o 0 0 0 •B. C 0 o 0 0 0 0 0 0 u H- A 0 ® c

(3\ 4-

O

Q £ Q £ toO 4-

O Ib *» c 4-Q CL

X Q.

w LU £ Q 0 0 Ci

CD

O

O 0 0 0 o 0 O c0 C 0

Hb

U— >* c u -X c 0 U 05 0 C £ c C • 0 y 0 MB

S’ 0 L. £ 0 C to B.

C

y Ib 1 o

f=

JZ 0 MB 0 0 MB 0 0 4* y — CL u 4- -C c 4-

4" L.

A to 0 C

O 0 4- t 0 c to •B O E 3 "O o MB — MB 0 MB

4- 0 C to •o ^ o jC 0 0 BB

a

4- •— to

C

u o 0 0 c (3. t o CL 0 to 0 0 4- g

•B 4- £ 0 1- 0

*y

y

4-0 tO -o L. i (O

®o to 0y

4-0 k

a 3 0

to

N (0 0 S L.

C o 0 4- c MB 0 0 2 a

4- «B 4- o > 2 to o T3 y >. to to C 1. y 0 >. 0 2 Ib 0 u to n *o 3 0

2.

t. LU

-S

t- 4- S 0 *o 5

to 1b *o BB

O 4-

A l- LU to o. to 0 b 0 O') 0 to 0 >- c O 3 0 MB \b o c S c C 0 3 to 0 I— Q. 0 o c 0 a

a.

O 0 0 *o 0 k s

CM 0 4- JDC > g 0 0 BB 3 c — 1 to •B. 0 0 0 4- E c 4- 4- 0 MB 4-

X B» (0 V. 2 k

y

U 0

M-» MB

to «B» •o 0 £ to >* 0 >- -o

C 4- >M JC 1.

u c 4- MB c O *—

C 0 3 4- 0 0 0 to c 4- >v 0 ® BB 0 0 0 MB B

to ’5 o to

C BB S’

-m— 0 4- MB MB 1b 4- 0 to 0 MB to to 0 u % C o. 0 CL 0 0 C 4- 4* «B,. 0 U *o E — *o to •— MB MB s c 4* M 0 0 y c y b 0 0 1. 4- 0 n y U)

MB c 1. 0 05 0 0 MB

y 0 0 MBy *o

y

y 0 c c O 0 4- 1b — Ib y 1b c •B *o 4- 1b MB c 0 1. — 0 Lb to 0 0 CL c c 0 u E a 0 0 4- O) 4- 4- u o *o E © 0 *o © 4- 0 4- 4- <b 0 3 3 C 0 MB A c 4- a. 4* X 0 MB C <5 N O' (0 — 4- 3 0 0 y A 3 0 y LU "O 3 — to o O O o O O 0uc0

1. ^ 0 s s- q: 0 to

tX ^

"O0 O ^ 0 0c0 0 i- Q -O 5 8, 0U 0 —

o.

©

©

Ul "O

u E

•ou© ^ 1 § 0 to

+“w

to0 0Q — © 0 q: >

LU
</) •0 10 0 c 1 u c — <4

U0 < 0 Z 0 9 Z o0 0 0

4-C

•g 10

ech of 80

£io0 0 05 ^ O

.2

*0 •mm 0 o 4- •mm 4- 0 u

— 0 ^ 0 0 0 C .Q 0 0 —

V) 4- E 0 0 8- 2 o 4- 4-

— c C "O — 0 J3 5 m >»

0) CT0 u)

;/) o3 ? 4- 0 0 ? 1 0M

4-0U. 0 (T3 4- *U) C 0 0) S if "5 0 « c 0 0? — 0 C N o — mm S’ 6 COO 0 i. —• ^ u 3E — 9 a o 0 (A (n 9

>. Ql 0 .o

b S' g- Cl. 0 0 c c c S

••

•? 13

Q c

4»

a .c 3 0 o* e ^ 0 £ 0 0 0 1 CO S 0 o 0 i § 0 0 s E 0 c u

g

X •o

fe ^ C 0 0 i. 0 0 0 — < •— 4 £ 0 0 4- E C 0

UJ UJ Its a. h. h. 3 u -C U) 4 ^ 0 pC > c 0 0 0 c >-

SI

U o 4- 0 ^ 0 < u o 4- .O • « 0 u 0 0 c

iTt f b

g

lu 0 O l_ 0 c ® -g

«i» 0 0 >» 0 0 0 O -t- o 0 o 8- o o 1 s +. w •0 C o — U 0 u >. 0 0

tTi 0 u o 0 3 0 >

0)

(A 0 0 0 S’ t;

u.o 0

-o 0 5 *0) >0a

-rU

C

-s ^ a 0t 9 1 ^ 0 >0O 0

>•0 o0

L.

— 3 0 0 4* 2 5 O O'

b0

"S

4- >0Q oC 00

L. 0 o 0 o O 0 0 C U 1c0c 0 0 0 » 1 t_ 0 U *o 1 3 c 0 c 6 0 3 I U 4- 0 0 4- C 0 0 0 O C u C 0 u 0 >• U 0 £ 3 0 4> c M — 3

4- «) o N 0 9

U. £ i 0 4- 4- o «a> e •0 C 4- 0 4- 0 mmm 0 0 _ 0 0 9 c o s O" mZ 0 b u w O M 0

•M 0 £ •“ 0 0 — 0 u ^ 't: 0 C 0 0 a. U

C 0 — 0 u u: 0 c ifl 0 c X U 0

c/? 0

4-§ e e a 3 § 2 S i 0o 0 3 4» 0 O 0 c • 0 4- o UJ % 1 0 •4

:i3 0 0 E :: 0 0 e 1- L. -o c u 0 . *a u 0 U

ST

<D 0 c 0 — 0 L. V) e 3 10 10 0 0 £ £ 0 0 ^ Ll! c 0 U) 1 ^ O JZ 3 0 a. a o H- O’) 0 0 1- 3 0 0 X £ 0 C 0 0 0 < J

'J) 4- i

<0 O. u — b. 0 8(D —

® 9 0 O — 0

-o

a s b Q i

UJ *o

C

a § U1 g 0 *0 9

CL O*

— 0 U

LU >- I a

T3 c S 0 eb o Q §

LJ CM •a 0 a"

C 0 0 .Q

r try 0 0 0 O 0 .c i ^ < UJ 1 0 0 0 >* o a. S

•i 0 4- 0 0

h— CM

M

u -o 0 0 c c o S

.o >.

a

C £ 0 >

VI Z c 3 o JC

— 4* 0 O) s — s z •

yo 1/1 0 0 0 o L. £ *o o o u. 4- a* 0 c 0

dU 0 c a S

<T

O 3 — fO (/) — >. u 3 S’ 0 o c 0 — 3 C ? 0 U 5 S’ 3 3 3 — £ 0 C U o u

-s 0 0 0 mm

® >»

X 0 0 .C 4- < 0 S 0 0 0 0 5 1 > — S 0 > U U 0 8

i: 5 on 0 ® 0 0 0 0 0 0

CO CL 4-

C z > z a. 0 Q Q.

— •o 4- -O

O 0 0 0 o 0 o •

•0 0E c 0 0 't: 0 c u ^ o 4- w — *0 0 C

«**0 C >. 0 4-0 0 a^ C 0 (0 0 0 b 00 0 6 o O) CD c0 e0

(0

—

U 3

U)

— o 3

O’

M

— c c •—

U 0 <j s u 0 3 8

JO Cl

— o N .C 30 0 —

•“ 4- 0 *o

— o 0 > 0 c J3 4- c c. — s § 0 0 — E 0 o

13 > L- o 4- 04» 0 •a >* 0 o U c u >. s

4-

C i

CO <0 0 0 0 4-

U -0 0 3 —

>• ••

b o • 4- 0 u 4- 4- 0

UJ 0 U o >* 0 .c 0 o 0 0 — 4- K* UJ t/i U) c 0 4- £ 4- J= 3 0 e — 1 >- c 0 <0 4- E 0 0 o U .O CD

W- 0 0 (/) 0 u

O 0 £ c. E 4- 3 JC 0 c S

CN s c c ^ -g 0 c 0 U 0 O

U 0 1 0 0 0 1. >• U — 0 0 c PM ^ X b u — •0 0 0 *a 0 0 0 '*• -o o mm

CO i ^ c E JZ

Q o ^ m ^ Q Qm 0 o 0 o E 0 0 — 0 E 0 Q. 4- 0 o — 0) c. o L. 0 0 0 0 s s

•0 c 0) 0 o CU c 0 mm >.

O S c ^ c .c c 0 0 0 0

SI •H ? S’ c '4> 0 C 4- 0 s I — O 0

3- 0 <J) 3

•• 0 E 00 *0 a. 0 4- 5 i > S’

-o ““ = 0 c — 0

>. 0 u

u o C >%

C

u c 4- 0 S s 0 0 Q 0 c. 0 0^ a 0 0 ^ C > •0 4- 0 4“ 0 4- >0 4- o CL s ^

•^

•t- cr 5 ^ u

X 0 o 0 0 O

X 0 (Q

O U) 0 a. UJ £ O E 0 .o UJ

U E 0 o 0 0 0 0 Ico

0) co

inUi

U IQ C L. < 4-

4-

C

C

— o

biotechnological

processes
IB 0u

{/) IS ^ £ <

UJ LU 2

Q_5ogQ

<1

il< <

UJ

© I

>* 1 o 4 i © o £ 3 0 c0

*0

©E u C 4- 0 o 0

Q. 0 — 01

_ 01

C 0) 0

£. c 0 0 U

__

u "0 01 ^ 0 0 c u c c 01 o. 0 •1- 0 0 0 0 U u 01 4“ 4- 8 ^

a.

a.

(C0c1©0Q

ST Ss

CD

J

C

©

(/) fe.

— COO E S JD s

rr

1^

t/)

y)

'Oc

fOo ?-

CDO 0 </) N —

«- i/) *0 0 © b

rt: icT = 8 —^ if — O) o o

•A

— ^ o

CL -O

"C 0 < •*- c o — 5 S f0

*oCa ^ 6 o O 7 — o 0 I s 0 Q.

U c U Q. _ z Q U (Id 0 01 3

-0

UC 0 0 0 c 4> C 0 01 w u c 0 (71 ff u 01 a. c 0 >- 0 JO 0 >* *0 c

4-

u 3 0 0 z 0 4- u 0 o Q. Q. to u

*D

ODO - 5 o »*- O

a.

t/) 0 0 b 5

Q. «)

U 5 5r P ? >.5 © 0

•o

©

T3C0

>-N *0 C C

</> 0 0

>-

—

*0 0 © c ~ O 0 — 0 "0

-Htno 0u— "0C

S’ 0 O

Q.U = t

j: E u u 0 ©

4- £ c ^ ° «,* +- O 0 — C 4- — © E — 0 ^

•03

4- c

•0 o © — = t; 0 0

-K

©

CL

CL0

*0C0c — »

-0 >- C ^ 0

U1 S O —t

>-

©

-c CL & ^

-D >.E f b 0 o3U © 4-

Ci U1

©

—

u > 4- -P 0 S -s; i2

""

—

W S 0 0 0 ©

Lij > 5 ^ cr © O 0% £ >- —

*0— 3

4- 0

(/D

O 0 JC ©o U u © u u e -o

O

— -0

—

—

u0 o — — 01 ©

T3

4-^0 U C O ^ o — ^ z ^

Q. 2 0 s =

8-

O -5

—

®

© •(- -t > — o © c — Q 3 ^

tnm

o. E — © O l_ Z S’

_ o — 0 -8 0 0 0 © 0 0. 0 c 0 01 4- >. 0 o 60

Q. 5 0 ©

01c 3U 0 0 Z

L.o

4-0 t

S’

J2 01 >* 0 u Cl o 0 U 0 O 0 L. 0 1= © 5 — •— 0 3 0 £ u — 0

“6 0 4-

Q

u 4- o 0 0 01

01 c 0 E u

U 0 E 4- 3 * 3 O

— 0 0 3 •0

C

n

U "0 5 u © O > U U C N 0 © C ^ 0 01 u 4- 0 0 c 0 — o 0 ~ 01

— 3 c 01 01 0 X 4-

u < 0

S’ -6 u u g s o O) C 0

UJ o: o

01 c ? — 0 c >• o1 C 0 0 c 4- u 0) 0 0 u 01 01 u 0 4- 0 c 0 ... 1 E 0

JZ 01

V 0 01 6 £. c 01 4- £. 0 u •0 0 — •— C.

... *0 t £ U

u c .a 0 u 01 0 0 0 Q. £ 0 0 u 01

C

C

>- 0 0 01 4- 0 © c U 3 d $. 0

4T 0 .0 • • *0 c 01 .c 0) 0 © 4 01 0) £ c c 0 L. c C z c 0 0 ... c U 0 •— 0 — 0 © 0 0 u 0 0 u u >. 3 u u 0 4“ .c 0 0 — 0 u 4 0 4 — U 0 *0 wr c 01 u C 01 4- 0 0 0 01 0 4- >*

.c 0 V. 0 u 01 4-

u 3 ^ 01 >. 0 >* GO u 01 4- 4* 0 U c » .0 0 "0 4 01 c © C © 01 01 u 0 u -P 0 c 4 c 't: ^ 0 a < 4. — u \ 3 c 4- 0) Q C 0 rtj •mm 0 .c c

UJ 4- 3 01 *0 01 0 X 3 4- E 0 — mmm c. 1/1 > 4“ 01 4. $ C 0 •0 0 c 3 £i >- 0 c — 0 3 — a c C 0 01 u 0 — 4 u 0

•0 *0

u

•0c — o

0)

—

— 5 t — >. O *S 0)0 0 0 u N

>*

—

—

u

— 0 0 t- 0 o — 0

01

u

>- 0 f -6 o 0

>»u

01 § 5r +g s

O'

— ^ S’ o —o ^ z — o — 4- 0 01 O *0 — O

*0-2Oc

jcO)c ©o

>.u0u

o. ^ O o

Q.fcO2

01CU00c* 0) c©

t3

01 0 © u U ^ O *0 O ^s © —

i: t 3 ©

0* Q- <^ X0

2.X©

01o © Q. c 4 $ 0) 01 0 L. Q. y 3 U Q

01 CT

© .C

Q

u jr £ .c

©

© 0 Q. 0 -0

U ... .D c Om CL ..Ml. a. 4 E 5 4 © u © c u c 0 >- u C 3 £ 0 CL 0 — 05 >* Q. ©

01 4 © 4 0 3 0 ©

S’

Q. 01 01 0 01

© 0 0 Q. 0 c N 0 w 4 0 X 0 01 U © c £ 4 © C C 0 c u ©

S’

© 4 ©© 0) c >. Ol

u 0 0 © 01©

— £ c 01 u © © 0 4 0 C 0 01 *— -0 01 © C c ol *— 01 c u .c 0 0 U — N 0 u

S’

>- c ©

©

© Q. 0 — *— 0) 0 —

©

—* © 4 0 — P 0 .c

—— 0 6 2 U

— £ 4 c u £ c u 4 © u 0 c c 0 Q. — 0 u 1 0 0 0 0 >. 4 c U ... — 4 u 0 — U

'J 0 — .. .c 0 © 0 C

© c u L.U

u •0 0 ©

© *0 0 y© .c 3 © > c 0 3 y 01 .0 0) c 0 4 0 4 Q. CL CL 3 4 a. c 01 © 4 u 0* u © 3 c C 0 © © © X 0 © Q. © © (J c: — 0 u 4 1 1 I 1 C/> u Q © n u 01 Q < *0
Finally, Table 9.1 reflects an implementation strategy based upon a current

but dynamic technology. Therefore, it will be necessary for Bureau scientific,

technical and management staff to evaluate this approach peri odi cal ly versus industry inf rastructure support needs over the life cycle of this technology.

Further, the action items listed in Table 9.1 do not extend the Bureau's mission

into the biological sciences but rather concentrate on N8S strengths in the

chemical, physical and engineering sciences, and on traditional NB5 lines of

communication with the private sector.

149
Ii 115

i'

ivl’ ii
References

1. Bunten-Mines, £., Planning Report 1. N8S Contributions to Biotechnology : A Preliminary Report , NBS, April 21, 1980.

2. Long Range Plan of the National Bureau of Standards , June 1981, pp. 70-72.

3. Coates, J. F., Planning Report 9. Implications of Biotechnology for the

National Bureau of Standards: A Seminar Presentation, NBS, November 16,

mi'7~—

4. Center for Chemical Physics, National Measurement Laboratory, NBS, 1981

Annual Report , pp. 18-19.

5. Center for Analytical Chemistry, National Measurement Laboratory, NBS, 1981

Annual Report , p. 104.

6. Pfann, H. F., Petrochemical Industry Briefing Paper. Supporting Background

Information, Bureau of Industrial Economics, U. S. Department of Commerce,

August 18, 1981.

7. Chemical and Engineering News, June 8, 1981, pp. 30-83.

8. Chemicals Allied Products, U. S. Industrial Outlook, Bureau of Industrial

Economics, Li. S. Department of Commerce, 1982, pp. 91-111.

9. Petrochemical Industry Overview, Chemical Economics Handbook - SRI

International, February 1980.

10. Chemical Products Synopsis, Mansville Chemical Products, Cortland, N.Y.,

11. Faith, Keyes, and Clark's Industrial Chemicals, 4th Edition, John Wiley ^ Sons, New York, 19/S.

12. Lipinsky, E. S., Science^: 1465-1471, 1981.

13. Chemical Week, September 30, 1981, pp. 36-40.

14. Science 1339-1341, 1981.

15. Technology Forecasts, January 1982, pp. 2-4.

16. Schneider, N., E. F. Hutton, Biotechnology Directory, October 1981.

17. Chemical and Engineering News, May 4, 1981, pp. 35-41.

18. Industry Week, September 7, 1981, pp. 65-70.

19. Rudd, D. F., Trevino, A. A., Fathi-Af shar, S., and Stadtherr, M. A.,

Petrochemical Technology Assessment, John Wiley & Sons, New York, 1981.

151
20. Holmer, E. C., Chemistry and Industry, January 5, 1980, pp. 18-21.

21. Chemical Week, January 27, 1982, pp. 36-45.

22. Chemical and Engineering News, December 21, 1981, po. 31-53.

23. Chemical Manuf acturers Associaton News, January 1982.

24. Chemical Week, April 7, 1982, pp. 44-53.

25. Lowe, A., Chemistry and Industry, January 5, 1980, pp. 22-27.

26. Chemical Week, March 3, 1982, pp. 24-26.

27. Chemical and Engineering Mews, February 22, 1982, pp. 11-16.

28. Sheets, K. R. , and Zanker, A., U. S. News & World Report, June 8, 1981,

p. 30.

29. Ottaway, D. B., Washington Post, July 12, 1981, pp. Al, A16.

30. Downie, L., Jr., Washington Post, February 27, 1982, A19, A20.

31. Samuelson, R. J., Washington Post, May 12, 1981, D7, D9.

32. Sheets, K. R., and Benham, J., U. S. News & World Report, May 4, 1981,

pp. 49-51.

33. Bishop, J. E., Wall Street Journal, February 2, 1981, p. 16.

34. Kerr, R. A., Science 21^: 427-429, 1981.

35. Energy Future , Stobaugh, R. and Yergen, D., Eds., Ballantine, N.Y. 1979.

36. Davis, W. B., Houston Law Review 17_: 1005-1013, 1980.

37. Balzhiser, R. E., Chemical Engineering, January 11, 1982, pp. 74-96.

38. Taylor, P., Washington Post, March 2, 1982, D6, 010.

39. Chemical Week, July 29, 1981, pp. 26-31.

40. Chemical and Engineering News, May 25, 1981, pp. 17-22.

41. Hughey, A., Wall Street Journal, February 12, 1982, p. 48.

42. Business Week, September 24, 19'9, pp. 104-122.

43. Joyce, R. M., Science 209 : 89-95, 1980.

44. Schriesheim, A., and Kirshenbaum, I., American Scientist^: 536-542,

1981.

152
45. Schulz, H., Pure and Applied Chem. 2225-2241, 1979.

46. Demain, A. L., and Solomon, N. A., Scientific American 245: 67-75, 1981.

47. Eveleigh, 0. E., Scientific American 245 : 154-178. 1981.

48. ylitos. A., Chemistry and Industry, May 2, 1981, pp. 303-310.

49. Energy from Biological Sources , Office of Technology Assessment, Congress

of the United States, Washington, D. C., 1980.

50. Energy in Transition, 1985-2010. Committee on Nuclear and Alternative

Energy Systems, National Research Council, National Academy of Sciences,

Washington , 0 . C. , 1979.

51. Impacts of Applied Genetics, Office of Technology Assessment, Congress of

the United States, Washington, D. C., 1981.

52. Harris, G, A., Sinnett, C. E., and Swift, H. E., Chemtech, December 1981,

pp. 764-768.

53. Myerly, R. C. , Nicholson, M. 0., Katzen, R. , and Taylor, J. M., Chemtech,

March 1981, pp. 186-192.

54. Morris, S. C. , Moskowitz, P. 0., Sevian, W. A., Silberstein, S. , and

Hamilton, L. D., Science 206: 654-662, 1979.

55. Pimentel, D., Moran, M. A., Fast, S., Weber, G., Bukantis, R.

,

Balliett, L., Boveng, P., Cleveland, C., Hindman, 5., and Young, M.,

Science 212: 1110-1115, 1981.

56. Palsson, B. 0., Fathi-Af shar , S. , Rudd, D. F. , Lightfoot, E. N., Science

213 : 513-517, 1981.

57. Goldstein, I. S., Science 189: 847-852, 1975.

58. Zeikus, J. G., Ann. Rev. Microbiol.^: 423-464, 1980.

59. Chemical Week, February 24, 1982, pp. 47-52.

60. Coover, H. W., Jr., and Hart, R. C., Chemical Engineering Progress,

April 1982, pp. 72-75.

61. Chemical Week, February 24, 1982, p. 30.

62. The Journal of Commerce, February 16, 1982.

63. Chemical Week, December 9, 1981, pp. 42, 44.

64. Neijssel, 0. M., T.I.B.S. 5_: iii-iv, 1980.

65. Wetzel, R., American Scientist 68: 664-675, 1980.

153
66. Emery, A. E. H. , The Lancet, December 19, 1981, pp. 1406-1409.

67. Hopwood, D. A., Scientific American 245 : 90-102, 1981.

68. Dunhill, P., Chemistry and Industry, April 4, 1981, pp. 204-217.

69. Gaden, E. L., Jr., Scientific American 245 : 180-196, 1981.

70. New Potentials and Alternative Bases for the European Chemical Industry

(draft). Commission of the European Communities (Bio-Society),

February 1982

.

71. Genex Corporation, Rockville, Maryland,

72. Kline, Charles H., personal communication, February 1982.

73. Blank, D. H., Rosse, 0. G., and Pfann, H. F., International Trade in

Petrochemicals. Factors Relevant to the North American Trade Agreement.

Bureau of Industrial Economics, U. S. Department of Commerce, Washington,

D. C . , June 1 , 1981

.

74. Chemical Week, March 10, 1982, pp. 36-37.

75. Nature 392 , 1981.

76. Nature 294: 602, 1981.

77. Chemistry and Industry, February 6, 1982, p. 68.

78. The Vision of MITI Policies in 1980s.

79. Look Japan, November 10, 1981, p. 18, 19.

80. Inside R & D, November 19, 1980.

81. Chemical Week, August 19, 1981, p. 66.

82. Business Week, December 14, 1981, pp. 110-114.

83. Genetic Technology News, July 1981, p. 3.

84. Genetic Technology News, September 1981, p. 2.

85. Genetic Technology News, October 1981, pp. 4-5.

86. Business Week, October 19, 1981, p. 62, 64.

87. Chemical Engineering Processes, January 1982, pp. 29-34.

88. Pimentel, L. S., Biotechnol . & Bioeng. 1989-2012, 1980.

89. Brooke, J., The Washington Post, February 4, 1982, A16.

154
90. Business Week, February 8, 1982, p. 49.

91. Chemical and Engineering News, July 27, 1981, pp. 62-67.

92. Chemical and Engineering News, January 18, 1982, pp. 33-34.

93. Science Resources Studies Highlights, National Science Foundation,

December 31, 1981, p. 2.

94. Chemical and Engineering News, July 27, 1981, pp. 46-71.

95. Suckling, C., Chemistry and Industry, January 5, 1980, pp. 14-17.

96. Mansfield, E., Science^: 1365-1371, 1982.

97. Chemical Week, April 4, 1982, pp. 44-48.

98. Mansfield, E., Harvard Business Review, Nov. -Dec. 1981, pp. 99-106.

99. Madsen, A,, Private Power , Quill, New York, 1982.

100. Forrester, J. W., Innovation and the Economic Long Wave, Presentation at

Symposium on Technology, Innovation and Corporate Strategy (London),

November 16, 1978.

101. Mass, N. J., and Senge, P. M., Technology Review, Aug. -Sept. 1981,

po. 56-65.

102. Chemical and Engineering News, October 26, 1981, pp. 20-21.

103. Michaels, J. W., Baldwin, W., and Minard, L., Forbes, November 9, 1981,

pp. 16-1-174.

104. Foster, R. N., Research Management, January 1982, pp. 22-27.

105. Porter, M. E., Competitive Strategy , The Free Press, New York, 1980.

106. Hoch, S. H., Research Management, November 1981, pp. 13-18.

107. Merrifield, D. B., Research Management, November 1981, pp. 13-18.

108. Bicmass and the Earth's Energy Cycle. In Biomass: Applications,

Technology and Production, Cheremisinoff , N. P., Cheremisinol^f , P. N., and

Ellerbusch, F., Eds., Marcel Oekker, Inc., New York, 1980, pp. 1-27.

109. Goheen, D. W., J. Chem. Ed. 544-547, 1981.

110. Rosenberg, S. L., Enzyme and Microbial Technology^: 185-193, 1981.

111. Emert, G. H., and Katzen, R., Chemtech, October 1980, pp. 610-614.

155
112. Rees, J. F., J. Chem. Tech. Biotechnol. 30: 161-175, 1980.

113. Sitton, 0. C., and Gaddy, J. L., Biotechnol. & Bioeng._22: 1735-1748,

1980.

114. Sarkanen, K. V., Science 1^1 : 773-776, 1976.

115. Biotechnology in Canada: Promises and Concerns, Institute for Research and

Public Policy and the Science Council of Canada, September 1980.

116. Moriarity, A., Canadian Research, May 1981, pp. 20-23.

117. Clarke, M. R. , and Sastry, C. 6. S. , Canadian Research, January 1982,

pp. 52-56.

118. Lederman, P. B., Skovronek, H. S. , Oes Rosiers, P. E., Chem. Eng. Progress

71: 93-97, 1975.

119. Rees, J. F., J. Chem. Tech. Biotechnol, 22.- "^58-465, 1980.

120. Phillips, L. G., and Shadel, S. L.. Johns Hopkins APL Technical Digest _2:

63-68, 1981.

121. Coombs, J., Chemistry and Industry, April 4, 1981, pp. 223-229.

122. Chemistry International, No. 6, 1980, pp. 3-4.

123. Chemistry and Industry, May 16, 1981, p. 340.

124. Chemical Week, October 7, 1981, p. 41.

125. Genetic Eng. News, Jan, /Feb. 1982, p. 5.

126. Bull, M. A., Sterritt, R. M., and Lester, J. N., J. Chem. Tech. Biotechnol

3_[: 579-583, 1981.

127. North, W. J., Chemtech, May 1981, pp. 294-298.

128. Ciferri, 0., New Scientist, September 24, 1981, pp. 810-812.

129. Benemann, J. R. , Weissman, J. C. , Koopman, B. L., and Oswald, W, J., Nature

268 : 19-23, 1977.

130. Ghosh, S., Klass, 0. L., and Chynoweth, D. P., J. Chem. Tech. Biotechnol.

3j_: 791-807, 1981.

131. Kierstan, M., and Bucke, C., Biotechnol. & Bioeng. _19: 387-397, 1977.

132. Chibata, I., and Tosa, T., Adv. Appl . Microbiol . _22’ 1977.

133. Bucke, C. , and Wiseman, A., Chemistry and Industry, April a, 1981,

pp. 234-240.

156
134. Sanderson, J. E., Wise, D. L., and Augenstein, 0. C. , Biotechnology and

Bioengineering Symposium, No. 8, John Wiley & Sons, Inc., 1978,

pp. 131-151.

135. Inside R&D, May 19, 1982, p. 2.

136. The Futurist, October 1981, p. 67.

137. Fenical, W., Science 21^: 923-928, 1982.

138. Rochaix, J-D., and Van Dillewijn, J., Nature 296 : 70-72, 1982.

139. Wikstrom, P., Szwajcer, E., Brodelius, P., Nilsson, K., and Mosbach, K.,

Biotechnology Letters^: 153-158, 1982.

140. Calvin, M., Energy Technology Conference, Energy Technology^; 687-700,

1978.

141. Williams, L. R. , Casali, J., and Home, V., Chemistry in Austral i a 48

344-345, 1981.

142. Stewart, G. A., Rawlings, W. H. M., Ouick, G. R. , Begg, J. E. , and Peacock,

W. J., Search]^: 107-115, 1981.

143. Morgan, R. P., and Shultz, E. B., Jr., Chemical and Engineering News,

September 7, 1981, pp. 69-77.

144. Chemical Week, March 24, 1982, p. 32.

145. Chemical Week, April 28, 1982, pp. 40-4-1.

146. Technology Ireland 1_3- 43-45, 1981.

147. Margaritis, A., Bajpai, P., and Cannell, E., Biotechnology Letters 2-

595-599, 1981.

148. European Chemical News, 12: 24, 1981.

149. Biomass Digest 2: 2, 1981

150. Chemical Week, September 16, 1981, p. 48.

151. Fowler, M. W., Chemistry and Industry, April 4, 1981, pp. 229-233.

152. Blair, J. G., Science 82, January-February 1982, pp. 70-76.

153. Brodelius, P., and Mosbach, K., J. Chem. Tech. Biotechnol . 22: 330-337,

1982.

154. Chemical and Engineering News, June 22, 1981, pp. 33-44.

155. Cocking, E. C., Davey, M. R., Rental, D., and Power, J. B., Nature 293;

265-269, 1981.

157
156. Zaborsky, 0., personal communication, March 15, 1982.

157. Hsu, T. A., Ladisch, M. R., and Tsao, G. T., Chemtech, May 1980, pp.

315-319.

158. Righelato, R. C. , Phil. Trans. Roy. Soc. Lond. B290 : 303-312, 1980.

159. Inside R&D, February 10, 1982, p. 6.

160. Hamer, G., Biotechnol . & Bioeng. 511-531, 1982.

161. Technology Forecasts, February 1982, pp. 4-5.

162. Flickinger, M. C., Biotechnol . & Bioeng. _22 (Suppl. 1 ): 27-48, 1980.

163. Atkinson, B., Black, G. M., and Pinches, A., Process Biochemi stry. May

1980, pp. 24-32.

164. Chemical Engineering, January 26, 1981, pp. 51-53.

165. Chemical and Engineering News, December 7, 1981, pp. 35-42.

166. Inside R&D, February 3, 1982, p. 3.

167. Chemical Engineering, June 15, 1981, pp. 62-65.

168. Trivedi, L. S., and Rao, K. K., Biotechnology Letters _3- 281-284, 1981.

169. Chemical and Engineering News, February 1, 1982, pp. 22-23.

170. Hardt, H., and Lamport, D.T.A., Biotechnol. & Bioeng. 24_: 903-918, 1982.

171. David, C., Daro, A., and Monnoye, D., European Polymer J. _16: 1159-1166,

1980.

172. Ryu, D. D., and Mandels, M., Enzyme and Microbial Technology _2: 91-102,

1980.

173. Eriksson, K-E., Pure and Applied Chem. _53: 33-43, 1981.

174. De Young, H. G., High Technology, January/February 1982, pp. 61-67.

175. Yan, J. F., Science^: 1390-1392, 1982.

176. Hagerdal, B., Ferchak, and Pye, E. K., Biotechnol. & Bioeng. 22:

1515-1526, 1980.

177. Margaritis, A., and Creese, E., Biotechnology Letters _3^ 471-476, 1981.

178. Ferchak, P. D., Hagerdal, B., Pye, E. K., Biotechnol. & Bioeng. 22:

1527-1542, 1980.

158
179. Choudhury, N., Dunn, N. W., and Gray, P. P., Biotechnology Letters 2‘

493-496, 1981.

180. Trivedi, L. S., and Rao, K. K., Biotechnology Letters 2^ 281-284, 1981.

181. Inside FJ&D, April 7, 1982, p. 1.

182. Genetic Technology News, November 1981, p. 6.

183. Chemical Engineering, April 19, 1982, pp. 55-59.

184. Genetic Technology News, February 1982, pp. 3-4.

185. Orioli, E., lorio, G., and Santoro, R., J. Mol. Catalysis, H.* 247-251 , 1982.

186.. Maleszka, R. , Wang, P. Y., and Schneider, H., Biotechnology Letters _4:

133-136, 1982.

187. Jones, R. P., and Greenfield, P. F., Biotechnology Letters 3: 225-230,

1981.

188. Hahn-Hagerdal , B., Mattiasson, B., Albertsson, P. A., Biotechnology Letters

3 : 53-58, 1981.

189. Hahn-Hagerdal, B., Mattiasson, B., Anderson, E. , and Albertsson, P. A.,

J. Chem. Tech. Biotechnol . 157-161 , 1982.

190. Cysewski, G. R., and Wilke, C. R. , Biotechnol . & Bioeng. 19.* 1125-1143,

1977.

191. Lee, J. H., Woodard, J. C., Pagan, R. J., and Rogers, P. L., Biotechnology

Letters 3 : 177-182, 1981.

192. Lee, K. J., Lefebvre, M., Tribe, D. E. , and Rogers, P. L., Biotechnology

Letters 2 : 487-492, 1980.

193. Paca, J,, J. Chem. Tech. Biotechnol. ^O: 764-771, 1980.

194. Minier, M., and Goma, G., Biotechnology Letters 3: 405-408, 1981.

195. de Boks, P. A., and van Eybergen, G. C., Biotechnology Letters 2- 577-582,

1981.

196. Ladisch, M. R., and Dyck, K., Science 205: 898-900, 1979.

10,'. Atkinson, A., Ellwood, D. C., Evans, C. G. T., and Yeo, R. G., Biotechnol.

& Bioeng. }

7_: 1375-1377, 1975.

198. Linko, Y-Y., and Linko, P., Biotechnology Letters 3: 21-26, 1981.

159
199. Cho, G. H., Choi, C. Y., Choi, Y. D., and Han, M. H., Biotechnology Letters

3: 667-671, 1981.

200. Margaritis, A., Bajpai, P. K., and Wallace, J. B., Biotechnology Letters 3;

613-618, 1981.

201. Daugulis, A. J., Brown, N. M. , Cluett, W. R. , and Dunlop, D. B.,

Biotechnology Letters V. 651-656, 1981.

202. Chemical Week, December 17, 1981, pp. 19-20.

203. Arcuri, E. J., Worden, R. M., and Shumate, S. E., Biotechnology Letters 2:

499-504, 1980.

204. Genetic Engineering News, January/February 1982, p. 3.

205. Gong, C., McCracken, L. D., and Tsao, G. T., Biotechnol ocy Letters 3:

245-250, 1981.

206. Gong, C., Ladisch, M. R. , Tsao, G. T., Biotechnology Letters 3: 657-662,

1981.

207. Schneider, H., Wang, P. Y., Chan, Y. K., and Maleszka, R. , Biotechnology

Letters 3: 89-92, 1981.

208. Maleszka, R. , Veliky, I. A., and Schneider, H., Biotechnology Letters 2*

415-420, 1981.

209. Jeffries, T. W., Biotechnology Letters _3‘ 213-218, 1981.

210. Wang, P. Y., Shopsis, C., and Schneider, H., Biochem. Biophys. Res. Comm.

94: 248-254, 1980.

211. Saddler, J. N., Chan, M. K-H., and Louis-Seize, G., Biotechnology Letters

3: 321-323, 1981.

212. Khan, A. W., and Murray, W. D., Biotechnology Letters 177-180, 1982.

213. Detroy, R. W., Cunningham, R. L., Bothast, R. J., Bagby, M. 0., and

Herman, A., Biotechnol. & Bioeng. 1105-1113, 1982.

214. Wolnak, B., In Enzymes - The Interface Between Technology and Economics , Marcel Dekker, Inc., New York, pp. 3-10.

215. Wodzinski, R. J., Adv. Appl . Microbiol . 2^: 1-6, 1979.

216. Martinek, K., and Semenov, A. N., J. Appl. Biochem. 2‘ 93-126, 1981.

217. Kindel, S., Technology, November/December 1981, pp. 62-74.

218. Bucke, C. , and Wiseman, A., Chemistry and Industry, April 4, 1981,

pp. 234-240.

160
,_x9. Zaborsky, 0., Immobilized Enzymes, CRC Press, Cleveland, Ohio, 1973.

220. Dunnill, P., Phil. Trans. R. Soc. Lend. B 290 : 409-420, 1980.

221. Immobilized Enzymes for Industrial Reactions , Messing, R. A., Ed., Academic

Press, New York, 1975.

22. Cheryen, M., VanWyk, P. J., Olson, N. F., and Pvichardson, T., Dairy Sci . 58: 477-481, 1975.

223. Legoy, M. D., Gellf, G., Ergan, F., Coequempot, M. F., Garde, V. L., and

Thomas, D., J. Chem. Tech. Biotechnol . _3^: 170-178, 1982.

224. Freyssinet, J-M., and Hudry-Clergeon, G., Nature 289 : 91-93, 1981.

225. Ishimori, Y., Karube, I., and Suzuki, S., J. Mol. Catalysis 12: 253-259,

1981.

226 . Martinek, K., Mozhaev, V. V., Smirnov, M. D., and Berezin, I. V.,

Biotechnol . & Bioeng. 247-251 , 1980.

227. Mattiasson, B., J. Appl . Biochem. _3- 183-194, 1981.

228. Wiseman, A., Anal. Proc., August 1981, pp. 359-361.

229. O'Sullivan, M. J., Anal. Proc., March 1981, pp. 104-108.

230. Everse, J., Ginsburgh, C. L., and Kaplan, N. 0., Methods of Biochem. Anal.

135-201, 1979.

231. Venkatasubramani an , K., In Enzymes - The Interface Between Technology and

Econcomics , Marcel Oekker, Inc., New York, pp‘. 35-52.

232. Weetall, H. H., In Biotechnological Applications of Proteins and Enzymes,

Academic Press, New York, 1977, pp” 103-125.

233. Mosbach, K., J. Chem. Tech. Biotechnol. 3^: 179-188, 1982.

234. Felix, H., Anal. Biochem. 1^: 211-234, 1982.

235. Spier, R. E., J. Chem. Tech. Biotechnol. ^2: 304-312, 1982.

236. Hodge, P., Chemistry and Industry, September 15, 1979, pp. 624-627.

237. Chibata, I., and Tosa, T., T.I.B.S., April 1980, pp. 80-90.

238. Applied Genetics News, March 1982, p. 6.

239. Birnbaum, D., Pendleton, R. , Larsson, P-0., and Mosbach, '<

, , Biotechnology

Letters 3: 393-400, 1981.

161
240. Linko, Y-Y., Jalanka, H., and Linko, P., Biotechnology Letters _3‘- 263-268,

1981.

241. de Rosa, M., Gambacorta, A., Lama, L., and Nicolaus, B., Biotechnology

Letters 2: 183-186, 1981.

242. Klein, J., and Kluge, M. , Biotechnology Letters 2* 65-70, 1981.

243. Vorlop, K-D., and Klein, J., Biotechnology Letters 2’ 9-14, 1981.

244. Tso, W-W., and Fung, W-P., Biotechnology Letters 2^ 421-424, 1981.

245. Dias, S. M. M., Novais, J. M., Cabral, J. M. S., Biotechnology Letters 4:

203-208, 1982.

246. Atkinson, B., and Sainter, P., J. Chem. Tech. Biotechnol . 32: 100-108,

1982.

247. Marr, R. , and Kopp, A., Int. Chem. Eng. 2^: 44-60, 1982.

248. Michaels, A. S., Chemtech, January 1981, pp. 36-43.

249. Smith, I. H., and Pace, G. W., J. Chem. Tech. Biotechnol. 22: 119-129,

1982.

250. Genetic Technology News, January 1982, pp. 3-4.

251. Genetic Technology News, October 1981, p. 3.

252. Rogers, P. L., Lee, K, J., and Tribe, D. E., Process Biochemi stry,

August/September 1980, pp. 7-11.

253. Greco, G., Jr., Gianfreda, L., Albanesi, D., Cantarella, M. , J. Appl

.

Biochem. 2^ 233-245, 1981.

254. Gianfreda, L., and Greco, G., Jr., Biotechnology Letters 2‘ 33-38, 1981.

255. Li, N. N., The Chemical Engineer, July 1981, pp. 325-327.

256. Lilly, M. D., J. Chem. Tech. Biotechnol. 22: 162-169, 1982.

257. Martinek, K., Levashov, A. V., Klyachko, N. L., Pantin, V. I., and Berezin,

I. V., Biochem Biophys. Acta 657 : 277-294, 1981.

258. Chemical Engineering, October 5, 1981, pp. 75-79.

259. Chemical Week, February 27, 1982, p. 53.

260. Larsson, P-0, and Mosbach, K., Biochem. Soc. Trans. 2^ 285-287, 1981.

261. Ito, Y., Industrial Research & Development, April 1982, 108-111.

162

{

I
262. Barker, P. E., and Chuah, C. H., The Chemical Enqineer, August/September

1981, pp. 389-393.

263. Chemical Week, July 9, 1980, pp. 31-32.

264. 266. Chemical Engineering, January 25, 1982, pp. 50-51.

Randall, L. G,, Separation Science and Technology 17_: 1-118, 1982.

266. King, M. B., and Bott, T. R., Separation Science and Technology 17

119-150, 1982.

267. Calimli, A., and Olcay, A., Separation Science and Technology 27_- 183-197,

1982.

268. Clark, R., Lane, 0. P., and Tjian, R. , J. Biol. Chem. 256: 11854-11858,

1981.

269. Yelton, 0. E., and Scharff, M. D., American Scientist 510-516, 1980.

270. Medical World News, August 21, 1978, pp. 47-57.

271. Langer, R. , Technology Review, April 1981, pp. 26-34.

272. Genetic Technology News, January 1982, pp. 1-2.

273. Chemical and Engineering News, January 11, 1982, pp. 22-23.

274. Mohan, R. R., and Li, N. N., Biotechnol . & Bioeng. 17_: 1137-1156 , 1975.

275. Kirkland, J. J., Yau, W. W., and Szoka, F. C. , Science 215: 296-298,

1982.

276. Williams, 0. C. , van Frank, R. M., Muth, W. L., and Burnett, J. P., Science

215 : 687-689, 1982.

277. King, P. P., J. Chem. Tech. Biotechnol. _32: 2-8, 1982.

278. Chemical Week, March 31, 1982, p. 12.

279. Beddow, J. K., Chemical Engineering, August 10, 1981, pp. 70-84.

280. Hirschbein, B. L., Brown, D. W., and Whitesides, G. M., Chemtech, March

1982, pp. 172-179.

281. Chemical and Engineering News, April 26, 1982, pp. 20-21.

282. Munro, P. A., McGregor, P. J., and Waters, R. D., Biotechnology Letters _3*

297-302, 1981.

283. Vadgama, P., J. Med. Eng. & Tech. 5: 293-298, 1981.

163
284. Weeks, M. G., Munro, P. A., and Spedding, P. L., Biotechnology Letters 4:

85-90, 1982.

285. Sada, E., Katoh, S., and Terashima, M. , Biotechnol . ^ Bioeng. 22: 243-246,

1980.

286. Maugh, T. H., Science^: 1492-1493, 1982.

287. Mosback, K. and Anderson, L., Nature 270 : 259-261, 1977.

288. Chaplin, M. F. , and Kennedy, J. F. , Carbohydrate Research 50: 267-274,

1976.

'

289. Poliak, A., Blumenfeld, H., Wax, M., Baughn, R. L., and Whitesides, G. M.,

J. Am. Chem Soc . 6324-6336, 1980.

290. Molday, R. S., Yen, S. P. S., and Rembaum, A., Nature 268: 437-438, 1977.

291. Kronick, P. L., Campbell, G., and Joseoh, K., Science 200: 1074-1076,

1978.

292. Haber, N., Proc. Natl. Acad. Sci . 79_: 272-276, 1982.

293. Inside R&D, March 17, 1982, p. 2.

294. Fleischaker, R. J., Weaver, J. C. , and Sinskey, A. J., Adv. Appl

.

Microbiol.^: 137-167,1981.

295. Cooney, C. L., Wang, H. Y., and Wang, D. C., Biotechnol. & Bioeng. 1^:

55-67, 1977.

296. Taylor, M. J., and Richardson, T., Adv. Appl, Microbiol. 7-35, 1979.

297. Rechnitz, G. A., Science^: 287-291, 1981.

298. Webster, I. A., and Shuler, M. L., J. Chem. Tech. Biotechnol. _31- 226-234,

1981.

299. Mattiasson, P., Larsson, P-0., and Mosbach, K., Nature 268: 519-520,

1977.

300. Steinberg, G., Chemtech, December 1981, pp. 730-737.

301. Moore, D. F., Sandall, J. P. B., and James, A. M., Biotechnology Letters 3_- 675-678, 1981.

302. Steele, D. J., Anal. Proc., November 1981, pp. 489-491.

303. Lee, J. H., Woodard, J. C., and Rogers, P. L., Biotechnology Letters

251-256, 1981.

164
304. Inside R&D, February 10, 1982, p. 1.

305. Chayen, J. Biochem. Soc. Trans. 2‘- 587-589, 1981.

306. Pincus, M. R., and Scheraga, H. A., Acc. Chem, Res, 299-306, 1981.

307. dementi, E., Computation of Large Molecuies, Spr inger-Ver 1 ag

,

Berlin/ New York, 1981.

308. Methods in Enzymology (V. 55 & 56), Academic Press, New York, 1979.

309. Immobilized Enzymes, Research and Development, Chibata, I., Ed., Halstead,

New York, 1978.

^

310. Gray, 0. N., Keyes, M. H., and Watson, B., Anal. Chem. 49: 1067A-1078A,

1977.

311. Fishman, M. M. , Anal. Chem. 185R-199R, 1980.

312. Biochemical Thermodynamics, Jones, M. N., Ed., Elsevier Scientific

Publishing Co., 1979.

313. Alberty, R. A., J. Biol. Chem. m: 1337-1343, 1968.

314. Alberty, R. A., J. Biol. Chem. W: 3290-3302, 1969.

315. Carey, P. R. , Can. J. Spectrosc. _^6: 134-142, 1981.

316. Woodruff, W. H., and Atkinson, G. H., Anal. Chem. 48: 186-189, 1976.

317. Teo, B-K., Acc. Chem. Res. 1_3: 412-413, 1980.

318. Horrocks, W. DeW., Jr., and Sudnick, 0. R., Acc. Chem. Res. 14: 384-392,

1981. ~

319. Powell, C. J., Australian J. Physics (in press).

320. Powell, C. J., Appl . Surface Sci. 143-169, 1978.

321. Technical Activities 1981, Office of Standard Reference Data, NBS.,

Fivozmsky, S. P., Ed., NB8 iR 81-2442.

165
III
Glossary*

Aerobe. An organism that requires air or free oxygen to maintain life

processes

.

Anaerobe. An organism that does not require air or free oxygen to maintain its

life processes.

Amino Acid. An organic compound that contains one or more basic amino groups

(-NHp) and one or more acidic carboxyl groups (-C00H). They may be

polymerized to form peptides (short chains of amino acids) and more complex

forms (proteins) that are joined together in a strictly ordered sequence

which determines the primary (sequence), secondary (chain folding), and

tertiary (3-dimensional) structure. There are 20 common amino acids.

Antibody. A protein that is a component of the immune system. It originates

either normally or in response to a foreign substance or antigen, and is

characterized by a specific reactivity with its complementary antigen.

Antigen. A molecule, usually a protein or polymeric carbohydrate, which when

introduced in the body stimulates the production of an antibody that will

react specifical ly with it.

Biomass. Includes many plant species, various animal wastes, and concentrated

masses of unicellular organisms. It can serve as a feedstock for the

production of organic chemicals.

Biosynthesis. The production of a chemical compound by a living organism by

either synthesis or degradation.

Biotechnology. The collection of industrial processes and of technologies that

involve the use of biological systems. Some of these processes may involve

the use of genetically engineered microorganisms.

Carbohydrates. The family of organic molecules consisting of simple sugars such

as glucose and sucrose, and sugar chains ( polysaccharides) such as starch and

cel lulose.

Catalyst. A substance that alters the velocity of a chemical reaction and

remains essentially unaltered in form and amount at the end of the reaction.

Cell culture. An in vitro method of propagating cells from animal, plant, and

insect tissues; technique can be applied to the production of complex

proteins, polysaccharides, and hormones.

^Sources . Impacts of Applied Genetics , Office of Technology Assessment,

Washington, D.C., 1981; "Summary of Genetics Briefing,"

Congressional Clearinghouse on the Future, April 23, 1980;

Dictionary of Scientific and Technical Terms , Second Edition,

McGraw-Hi I i , 1978.

167
Cell fusion. The integration of the components of two or more cells to become a single cell; represents a pooling of genetic information that would not occur

under natural circumstances.

Cellulase. A group of extracel 1 ul ar enzymes produced by microorganisms and some lower animals that hydrolyze cellulose to simpler sugars such as anhydroglucose

.

Cell membrane. A thin layer of protoplasm, consisting mainly of lipids and

proteins, which is present on the surface of all cells (plasma membrane).

This cell structure is involved in regulating the flow of substances into and

out of cel Is.

Cell permeability. The permitting or activating of the passage of substances

into and out of cells through the cell or plasma membrane.

Cellulose. A polysaccharide composed entirely of several glucose units linked

end to end; it constitutes the major part of cell walls in plants.

Clone. An individual organism derived asexual ly from a single cell through

mitosis. Phenotypic or genotypic homogeneity is usually but not always a characteristic of the resulting population.

Coenzyme. A nonprotein portion of an enzyme that can be associated at the

active site of an enzyme. It is a prosthetic group which functions as an acceptor of electrons or functional groups (for example, nicotinamide adenine

dinucleotide-NAD) .

Cytoplasm. The protoplasm of a cell, external to the cell's nuclear membrane.

DNA (deoxyribonucleic acid). The genetic material found in all living

organisms. DNA is composed of four deoxyr ibonucleotide building blocks

(nitrogenous base + sugar + phosphate). The nitrogenous bases of DNA are

paired; adenine (A) + thymine (T) and guanine (G) + cytosine (C). The base

pairs are joined by hydrogen bonds. The four bases form the alphabet of the

genetic code. The sequence of each base in a linear array along the

sugar-phosphate backbone of DNA determines the information content of the

gene. Every inherited characteristic has its original somewhere in the code

of each individual's complement of DNA.

Enzyme. A functional protein (e.g., protein with a tertiary structure and

active site) that catalyzes a specific chemical reaction without being

affected by the reaction. Enzymes are the active agents in fermentation

processes, and control the rates of reaction in the vast majority of chemical

reactions in all living organisms.

Eukaryote. A higher, compartmentalized cell characterized by its extensive

internal structure and the presence of a nucleus containing the DNA. All

multicellular organisms are eukaryotic. The simpler cells, the prokaryotes,

have much less compartmental i zat ion and internal structure; bacteria and some algae are prokaryotes.

Feedstock. The raw material furnished to a particular process.

168
Fermentation. The process of transformation by enzymes or microorganisms of

large molecular weight organic substrates {or feedstocks), especially

carbohydrates, to lower molecular weight chemicals. It is used in various

industrial processes for the manufacture of products such as alcohols, acids,

and certain dairy products (such as cheese) by the action of yeasts, molds,

and bacteria.

Gene. Basic hereditary unit; a sequence of DNA which codes for a specific

protein

.

Genetic code. The biochemical basis of heredity consisting of codons

(nitrogenous base triplets along the DNA sequence, e.g., ATA, GAT, AGA, etc.)

that determine the specific cjnino acid sequence transcribed and eventually

translated into proteins. The genetic code is a universal code for all forms

of life studied so far.

Genetic engineering. A technology used at the laboratory level to alter the

hereditary apparatus of a living cell so that the cell can produce more or different chemicals, or perform completely new functions.

Gene mapping. Determining the relative locations of different genes on a given

chromosome.

Genotype. The genetic constitution of an individual or group.

Hybridoma. The cell product of the fusion of two different types of cells which

possesses new combinations of properties inherited from both distinct parent

cells. For example, in the prepartion of monoclonal antibodies, one parent

is a cell (lympocyte) which produces a specific antibody. The other parent

is a tumor cell from a myeloma, a cell which can propagate indefinitely

("immortal") in cell cultur*e. Once these two parent cells are combined/fused, a hybridoma (hybrid melanoma) is formed that after cloning

and selection, can propagate indefinitely in cell culture and can product

large amounts of monoclonal antibody. This antibody recognizes only one specific antigen.

Hydrocarbon. Compounds composed of carbon and hydrogen, and are commonly found

in petroleum, coal and gas.

In vitro. Outside the living organism and in an artifical environment.

In vivo. Within the living organism.

Lignocel lulose. A group of substances in v/oody plants consisting of cellulose,

hemicel 1 ulose, and lignin.

Liposome. Synthetic microscopic phospnolipid vescicle or carrier that can encapsulate individual cells, enzymes, antibiotics, etc. and can serve as controlled substance release vehicles and as vehicles for transferring

genetic material into plants.

Metabolism. The sum of the physical and chemical processes involved in the

maintenance of life and by which energy is made available.

169
Mitochondri a. Structures in the cytoplasm of higher cells that serve as the

"powerhouse" for the cell, producing chemical energy.

f^onoclonal antibodies. Antibodies derived from a single source or clone of

cells (for example, a hybridoma) which recognize only one kind of antigen.

Mutants. Organisms whose visible properties with respect to some trait differ

from the norm of the population due to mutations in its DNA.

Mutation. Any change that alters the sequence of bases along the DNA, changing

the genetic material.

Nucleic acid. A polymer composed of DNA or RNA subunits.

Nucleotides. The fundamental units of nucleic acids. They consist of one of

the four bases--adenine, guanine, cytosine, and thymine (uracil replaces

thymine in the case of RNA)--and its attached sugar-phosphate group.

Organic compounds. Chemical compounds composed of carbon and hydrogen, and also

may contain oxygen, nitrogen, and various other elements.

Phenotype. The sum total of the expression of morphological characteristics (and

the manifestation of physiological, enzymatic, etc. processes) of an organism. Represents the output from the interaction of an organism's

genotype with its environment.

Plasmid. Hered i tary materi al that is not part of a chromosome. Plasmids are circular and self-replicating. Because they are generally small and

relatively simple, they are used in recombinant DNA experiments as acceptors

of foreign DNA, and as vectors for insertion of foreign DNA into organisms.

Polymer. A long-chain molecule formed from smaller repeating structural units.

Polysacchar ide . A long-chain carbohydrate containing many molecules of simple

sugars linked together; examples would include cellulose and starch.

Prokaryote. Cells of bacteria or blue-green algae which are character ized as being rather small, having a single chromosome that is not enclosed by a nuclear membrance, and lacking cytoplasmic organelles.

Protein. A linear polymer of amino acids; proteins are the products of gene

expression and are the functional and structural components of cells.

Protoplast. Cell without a wall (for example, certain enzymes such as pectinases and cellulases can dissociate tobacco leaves into living but

wal 1-less pi ?nt cel Is)

.

Protoplast fusion. A means of achieving genetic transformation by joining two

protoplasts or joining a protoplast with any of the components of another

cell.

170
Recombinant DNA. The hybrid DNA produced by joining pieces of DNA from

different sources.

Restriction endonuclease. Enzyme isolated from a varity of bacteria recognizes

and degrades DNA from foreign organisms, thereby preserving the genetic

integrity of the bacterium. In recombinant DNA experiments, restriction

endonuclease enzymes are used as tiny biological scissors to cut up foreign

DNA before it is recombined with a vector. These enzymes have a unique

specificity for one specific nucleotide sequence.

Saccharification. The conversion or enzymatic hydrolysis of more complex sugars

(e.g., disaccharides-sucrose) to simpler sugars (e.g., monosacchar idesdextrose and levulose).

Vector. A transmission agent; a DNA vector is a self-replicating DNA molecule

that transfers a piece of DNA from one host to another, for example, a plasmid

.

171
i'

::

I

'

'

I

\
NBS-114A (REV. 2-80) U.S. DEPT. OF COMM. 1. PUBLICATION OR 2. Performing Organ. Report No. 3. Publication Date BIBLIOGRAPHIC DATA

SHEET (See instructions)

REPORT NO.

NBSIR 82-2549 July 1982

4. TITLE AND SUBTITLE

NBS AND INDUSTRI.AL BIOTECHNOLOGY: TECHNICAL DEVELOPMENTS AND FUTURE ME.ASUREMENi NEEDS

5. AUTHOR(S)

Dr. Thomas C. 0 Brien

6. PERFORMING ORGANIZATION (If joint or other than NBS. see instructions) 7. Contract/Grant No. national bureau of STANDARDS

DEPARTMENT OF COMMERCE

WASHINGTON, D.C. 20234

8.

Type of Report & Period Covered

9.

SPONSORING ORGANIZATION NAME AND COMPLETE ADDRESS (Street, City. State. ZIP)

10.

SUPPLEMENTARY NOTES

I I Document describes a computer program; SF-185, FIPS 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) B1 Ot EChnO 1 Ogy Wl 1 1 be d Significant industrial

technology in the future. NBS's role with respect to this technology is examined; and

infrastructure support requirements needed by future biotechnology based industries are identified in this report. This report also describes steps NBS could take to meet

future industry infrastructure support needs in biotechnology by: examining commodity

organic chemical industry trends; identifying R&D opportunities and barriers to

commerci al i zation of biotechnology products; and evaluating current NBS capabilities in

relation to long-term industry needs.

Report conclusions indicate that: (a) the commodity organic chemical industry will

undergo structural changes in the next two decades; (b) early applications of

biotechnology will be in higher value added products; (c) "traditional" commodity

chemicals synthesized from petroleum feedstocks will be difficult to displace via

biotechnological process; (d) biotechnology offers significant opportunities for

production of "non-traditional" commodity organic chemicals; (e) biomass may become an important feedstock for production of organic chemicals; and (f) existing NBS

capabilities could meet many of industry long-term infrastructure support needs in

biotechnology.

This report provides a framework for possible short- and long-term actions

NBS could take to meet industry needs in biotechnology. Report includes an extensive

bibliography.

12. KEY WORDS (S/x to twelve entries; alphabetical order; capitaliz‘d only proper names; and separate key words by semicolon s) Biomass conversion R&D; bioprocess engineering; biotechnology; chemical industry

trends/strategies; commodity organic chemicals; measurement/eval uated data needs; NBS

research capabilities.

13. AVAILABILITY

I X I Unlimited

I I For Official Distribution.

14. NO. OF

PRINTED PAGES

Do Not Release to NTIS 180

I I Order From Superintendent of Documents, U.S. Government Printing Office, Washington, D.C. 20402. 15. Price

[~Xl Order From National Technical Information Service (NTIS), Springfield, VA, 22161 $16,50

USCOMM-DC 6043-P80
J