Methods of Analysis by The U.S. Geological Survey

National Water Quality Laboratory—Determination of 

Organic Plus Inorganic Mercury in Filtered and Unfiltered 

Natural Water with Cold Vapor–Atomic Fluorescence 

Spectrometry

Water-Resources Investigations Report 01-4132 

 

U.S. Department of the Interior 

U.S. Geological Survey
Methods of Analysis by the U.S. Geological Survey

National Water Quality Laboratory—Determination of 

Organic Plus Inorganic Mercury in Filtered and Unfiltered 

Natural Water with Cold Vapor–Atomic Fluorescence 

Spectrometry

By John R. Garbarino and Donna L. Damrau 

_______________________________________________________

U.S. GEOLOGICAL SURVEY

Water-Resources Investigations Report 01-4132 

Denver, Colorado 

2001
U.S. Department of the Interior

Gale A. Norton, Secretary

U.S. Geological Survey

Charles G. Groat, Director

The use of trade, product, or firm names in this report is for descriptive 

purposes only and does not imply endorsement by the U.S. Government. 

For additional information write to: Copies of this report can be purchased from: 

Chief, National Water Quality Laboratory U.S. Geological Survey

U.S. Geological Survey Branch of Information Services

Box 25046, Mail Stop 407 Box 25286 

Federal Center Federal Center

Denver, CO 80225-0046 Denver, CO 80225-0286 
CONTENTS 

Abstract 1 

Introduction 1 

Analytical method 2 

1. Application 2 

2. Summary of method 2 

3. Sample collection and storage 3 

4. Quality-control samples, sampling methods, sample-collection equipment, 

and cleaning procedures 3 

5. Contamination and interferences 4 

6. Apparatus and instrumentation 5 

7. Reagents and calibration standards 5 

8. Sample preparation 6 

9. Analytical procedure 7 

10. Calculations 7 

11. Reporting of results 7 

Discussion of results 8 

Conclusions 14 

References cited 15

FIGURES 

 1-3. Box plots showing:

1. Results for a series of solutions based on a National Institute of Standards 

and Technology standard reference material preserved with acid dichromate 

or hydrochloric acid 9 

2. Results for U.S. Geological Survey standard reference water samples Hg24 

and Hg26 and two dilutions of a Baker Instra-Analyzed standard preserved 

in acid dichromate 11 

3. The effects of using argon instead of nitrogen as the purge gas in cold 

vapor–atomic fluorescence spectrometry 12 

4. Graph showing linear regression analysis of results for sample solutions 

based on two reference standards (experimental) and National Institute 

of Standards and Technology standard reference material (theoretical) 13

TABLES 

1. Method detection limit and analytical precision for the determination of 

mercury by cold vapor–atomic fluorescence spectrometry 3 

2. Typical instrument operating conditions 7 

3. Average percent spike recoveries for mercury in reagent-water, filtered and 

unfiltered surface-water, and filtered and unfiltered ground-water matrices 14

Contents III
CONVERSION FACTORS 

Multiply By To obtain

Length

micrometer (μm) 3.94 x 10-5 inch 

millimeter (mm) 3.94 x 10-2 inch 

nanometer (nm) 3.94 x 10-8 inch 

Mass 

gram (g) 3.53 x 10-2 ounce, avoirdupois 

milligram (mg) 3.53 x 10-5 ounce, avoirdupois 

nanogram (ng) 3.53 x 10-11 ounce, avoirdupois 

Volume

liter (L) 2.64 x 10-1 gallon 

microliter (μL) 2.64 x 10-7 gallon 

milliliter (mL) 2.64 x 10-4 gallon 

Pressure 

kilopascal (kPa) 1.45 x 10-1 pounds per square inch 

Degrees Celsius (oC) may be converted to degrees Fahrenheit (oF) by using the following

equation: 

oF = 9/5 (oC) + 32. 

ABBREVIATED WATER-QUALITY UNITS

mg/L milligrams per liter

μg/L micrograms per liter

μg/mL microgram per milliliter

ng/L nanograms per liter

mL/min milliliters per minute

L/min liters per minute

IV Contents
ABBREVIATIONS AND ACRONYMS

ASTM American Society for Testing and Materials 

CASRN Chemical Abstract Service Registry Number 

CV–AAS cold vapor–atomic absorption spectrometry

CV–AFS cold vapor–atomic fluorescence spectrometry

HCl hydrochloric acid 

M Molar (moles per liter)

LT–MDL long-term method detection level 

MDL(s) method detection limit(s) 

MRL method reporting level 

HNO3 nitric acid 

N Normal (acid equivalents per liter)

NIST National Institute of Standards and Technology

NWQL National Water Quality Laboratory

s second 

sp. gr. Specific gravity

SRM standard reference material

SRWS(s) U.S. Geological Survey standard reference water sample(s) 

USEPA U.S. Environmental Protection Agency

USGS U.S. Geological Survey

< less than

± plus or minus 

GLOSSARY 

MDL — The method detection limit (MDL) is defined as the minimum concentration of an 

element that can be measured and reported with 99-percent confidence that the element

concentration is greater than zero and is determined from analysis of a sample in a given matrix

that contains the element of interest (U.S. Environmental Protection Agency, 2000).

Contents V
Methods of Analysis by the U.S. Geological Survey National 

Water Quality Laboratory—Determination of Organic Plus Inorganic 

Mercury in Filtered and Unfiltered Natural Water with Cold Vapor – 

Atomic Fluorescence Spectrometry

By John R. Garbarino and Donna L. Damrau 

Abstract

An analytical method using cold vapor–

atomic fluorescence spectrometry was 

developed by the U.S. Geological Survey in 

2001 for the determination of organic plus 

inorganic mercury in filtered and unfiltered 

natural water. This method was developed to 

eliminate the use of acid dichromate

preservative and to provide capability to 

measure ambient mercury concentrations in

natural water. Dissolved mercury includes all 

oxidizable mercury species present in natural 

water that has been filtered through a 0.45-

micrometer pore size capsule filter. Wholewater recoverable mercury includes dissolved 

mercury species and mercury species 

adsorbed to particulate matter in unfiltered 

natural water. Mercury species can include 

elemental mercury, mercury (II), mercury (II)

complexes, various alkyl- and phenylmercury compounds, and other forms of 

mercury. In this method, samples are 

collected and processed according to standard 

U.S. Geological Survey protocols. Samples 

are preserved onsite with 6N hydrochloric 

acid in a ratio of 1 to 100 in a borosilicateglass bottle with fluoropolymer-lined cap. 

Mercury species are oxidized to mercury (II)

by using bromine monochloride; excess 

oxidation reagent is neutralized with 

hydroxylamine hydrochloride. Elemental 

mercury produced after adding stannous 

chloride is purged from the solution with 

ultrapure argon gas into a cell in which the

mercury concentration is measured by atomic 

fluorescence emission at 253.7 nanometers. 

The analytical response is linear up to 125 

nanograms per liter (ng/L) of mercury, and 

the short-term method detection limit is about 

5 ng/L. The analytical variability at 50 ng/L

is about 10 percent. 

This report describes the method and 

compares the use of hydrochloric acid to acid 

dichromate as a field preservative. Ambient 

mercury concentrations in hydrochloric acidpreserved samples stored in borosilicate-glass 

bottles with fluoropolymer-lined caps are 

shown to be stable for at least 30 days. 

Mercury concentrations are stable for at least

5 months after bromine monochloride is 

added to the sample bottles in the laboratory. 

The long-term average percent recoveries at

20, 45, and 75 ng/L in reagent water, filtered 

and unfiltered ground water, and filtered and 

unfiltered surface water range from 89 to 108, 

96 to 103, and 94 to 98 percent, respectively.

INTRODUCTION 

The ambient concentration for dissolved 

and whole-water recoverable mercury is 

generally at the parts-per-trillion level 

(nanograms-per-liter, ng/L) for natural-water

samples. Measurement at this concentration 

level is important because mercury

bioaccumulates in living organisms. The 

previous method (Fishman and Friedman, 

1989, p. 289–291) that used cold vapor–

atomic absorption spectrometry (CV–AAS) 

was not capable of measuring such low 

concentrations. Cold vapor–atomic 

fluorescence spectrometry (CV–AFS) is 

capable of measuring 5 ng/L mercury directly, 

without sample preconcentration. By

Introduction 1
eliminating sample preconcentration, analysis 

throughput is increased while maintaining the 

capability to determine mercury

concentrations at levels typically regulated by

water-quality standards. The mercury

concentration, organic plus inorganic species, 

is determined in filtered and unfiltered 

natural-water samples by using a brominating

digestion procedure that minimizes

interferences. Samples are preserved with

hydrochloric acid instead of acid dichromate 

solution, which has previously been used. 

This change was necessary because shipment

of samples preserved with dichromate

solution is restricted, and the cost of disposing

of chromium waste is expensive. In addition, 

hydrochloric acid sample preservation has 

been shown as effective as sample

preservation with acid dichromate solution. 

This report describes a method for 

determining mercury in natural-water 

samples. It was developed by the U.S. 

Geological Survey (USGS) for use in the 

National Water Quality Laboratory (NWQL). 

It is rapid, more efficient, and can detect

lower concentration levels compared to 

previous USGS methods (Fishman and 

Friedman, 1989, p. 289–291). This method 

supplements other methods of the USGS for 

the determination of mercury that are

described by Fishman and Friedman (1989) 

and by Fishman (1993). The new method was 

implemented at NWQL in April 2001. 

This report provides a detailed 

description of all aspects of the method, 

including the instrumentation, the reagents, 

the analytical procedure, and the qualitycontrol procedures. Mercury concentration is 

reported by NWQL in micrograms per liter

instead of nanograms per liter, the

concentration unit used throughout this report, 

because of current (2001) limitations in the 

data base. 

ANALYTICAL METHOD

Inorganic Constituents and Method, Lab, and Parameter Codes 

Constituent Method Lab 

code 

Parameter and 

method code 

Mercury, water filtered, organic + inorganic, g/L I-2464-01 2707 71890C

Mercury, water unfiltered, organic + inorganic, g/L I-4464-01 2708 71900D

1. Application 

This method is used to determine 

concentrations of organic plus inorganic 

mercury in filtered and unfiltered naturalwater samples. Bromine monochloride is 

used to oxidize elemental mercury, mercury

complexes, various alkyl- and phenylmercury compounds, and other forms of 

mercury. The upper limit of the linear 

concentration range is 125 ng/L mercury. 

The short-term method detection limit (MDL, 

see Glossary) is 5 ng/L mercury (table 1). 

2. Summary of Method 

Concentrations of organic plus inorganic 

mercury in filtered and unfiltered natural-water

samples are determined by using CV–AFS. 

The method is based on U.S. Environmental 

Determination of Organic Plus Inorganic Mercury in Filtered and Unfiltered 

Natural Water with Cold Vapor-Atomic Fluorescence Spectrometry

2
Protection Agency (1999) Method 1631 but 

does not use gold amalgamation for mercury

preconcentration. By eliminating the 

preconcentration procedure, the MDL is at 

least a factor of 10 greater, yet the method 

provides the determination of mercury at 

ambient concentrations. Mercury species in a 

natural-water sample are oxidized to mercury

(II) inside the borosilicate-glass sample bottle

by using bromine monochloride; excess 

oxidant is neutralized with hydroxylamine 

hydrochloride. Mercury (II) is reduced to 

elemental mercury by using stannous 

chloride, and is purged from the solution with 

ultrapure argon gas into a cell in which the

mercury concentration is measured by atomic 

fluorescence emission at 253.7 nanometers 

(nm). 

Table 1. Method detection limit and analytical 

precision for the determination of mercury by cold 

vapor–atomic fluorescence spectrometry

[ng/L, nanograms per liter; MDL, method detection limit] 

Element Mean, 

ng/L 

Standard 

deviation, 

ng/L 

tstatistic 

Degrees 

of 

freedom

MDL,

ng/L

Mercury 11 2 2.492 24 5

3. Sample Collection and Storage 

Sample-collection protocol must follow 

either the procedure outlined for trace 

elements in Horowitz and others (1994) or for 

mercury in Olson and DeWild (1999). The 

level of contamination related to field 

collection and processing for either protocol, 

however, must be established by using an 

equipment blank. Filtered and unfiltered 

natural-water samples are preserved with 

hydrochloric acid to a pH less than 2. 

Samples are stored in clean borosilicate-glass 

sample bottles with fluoropolymer-lined caps. 

Bottles are prepared by rinsing once with 

10 percent (by volume) mercury-free nitric 

acid and three times with deionized water

before air-drying on a laminar-flow clean 

bench. The sample bottle must be rinsed with 

a portion of the filtered or unfiltered natural 

water prior to filling the bottle with about 200 

mL of sample. Every sample must be

preserved with 2 mL of 6N hydrochloric acid 

(HCl); more acid might be required to lower 

the pH to 2 in some matrices. It is important 

that field personnel use 6N HCl prepackaged 

in polypropylene vials that can be obtained 

from the USGS Ocala Water Quality and 

Research Laboratory in Florida (part number 

06910) because the quality control is 

continually monitored. Ambient mercury

concentrations are stable for 1 month in 

preserved samples. Nevertheless, samples 

should be shipped to the laboratory for 

analysis as quickly as possible. 

4. Quality-Control Samples, Sampling 

Methods, Sample-Collection 

Equipment, and Cleaning Procedures 

4.1 Quality-control samples. Collection 

of quality-control (QC) samples is a required 

component of sample collection for waterquality studies. QC samples are collected, 

usually onsite, to identify, quantify, and 

document bias and variability in data that

result from collecting, processing, shipping, 

and handling of samples by field and 

laboratory personnel. The type, number, and 

distribution of QC samples are determined by

the design and data-quality requirements of

the study. Detailed discussion of the types 

and purposes of quality-control samples is 

provided in Wilde and others (1999). 

The primary purpose of a blank sample is 

to identify potential sources of sample

contamination and to assess the magnitude of 

mercury contamination. Field blanks are 

collected and processed onsite in the same

Analytical Method 3 
manner and by using the same equipment as 

for the environmental samples. The field 

blank is an aliquot of blank water that is 

processed sequentially through each 

component of the sampling system. The 

source water needed for blank samples must 

be produced and certified by a laboratory to 

have mercury concentrations that do not 

exceed a specified method detection limit. 

Inorganic-free blank water must be used for 

all equipment field blanks (Horowitz and 

others, 1994, p. 22). 

Replicate samples also may be

collected. The primary purpose of replicate 

samples is to quantify the variability in all or

part of the sampling and analysis process. 

Replicates, environmental samples collected 

in duplicate, triplicate, or higher multiples, are

considered identical in composition and are 

analyzed for the same chemical constituents. 

4.2 Sampling methods. Use methods 

that allow collection of water samples that

accurately represent the water-quality

characteristics of the surface water or ground 

water at a given time or location. Detailed 

descriptions of sampling methods used by the 

USGS for obtaining surface-water samples, 

ground-water samples, and sample-processing

procedures (splitting, filtration, and shipping) 

are provided by Wilde and others (1999). 

4.3 Sample-collection equipment. Use 

sample-collection equipment and automatic

samplers that are free of tubing, gaskets, and 

other components made of metal or that might 

sorb mercury from the water. The best

material for sample-collection and processing

equipment is any type of fluorocarbon 

polymer. 

4.4 Cleaning procedures. Follow the

procedures outlined in either Horowitz and 

others (1994) or Olsen and DeWild (1999). 

5. Contamination and Interferences 

Contamination must be avoided because 

of the inherently low mercury concentration 

found in natural water. The analyst and field 

personnel need to be attentive to potential 

sources of contamination. Sampling

equipment must be cleaned, and samples need 

to be collected as described by Horowitz and 

others (1994) or by Olson and DeWild 

(1999). Every sample must be preserved with 

2 mL of 6N HCl that can be obtained from the 

USGS Ocala Water Quality and Research

Laboratory in Florida. Samples might 

become contaminated during analysis by

carryover from a previous sample that had 

unusually high mercury concentration. 

Contamination from airborne sources can be 

minimized by placing the autosampler inside

a plastic enclosure pressurized with mercuryfree nitrogen. 

Gold, silver, and iodide interfere with 

the determination of mercury by this method. 

Elemental mercury is amalgamated by gold or

silver. The recovery of 2.5 ng/L of mercury

in the presence of 5 to 100 mg/L iodide 

ranges from 100 to 0 percent (U.S. 

Environmental Protection Agency, 1996; 

Bloom, 1995). After analyzing a sample that 

contains greater than 30 mg/L iodide, it may

be necessary to clean the analytical system

with 4N HCl (U.S. Environmental Protection 

Agency, 1999; Bloom, 1995). Most forms of 

mercury are oxidized by the bromine 

monochloride solution; however, the recovery

of mercury bound within microbial cells 

might require additional ultraviolet 

photooxidation (U.S. Environmental 

Protection Agency, 1999). Buildup of 

condensation in the vapor generator or 

fluorescence cell will degrade the analytical

signal. Condensation can be eliminated by

wrapping heating tape around the vapor 

generator and by using a drying tube in front 

Determination of Organic Plus Inorganic Mercury in Filtered and Unfiltered 

Natural Water with Cold Vapor-Atomic Fluorescence Spectrometry

4
of the fluorescence cell. The drying tube 

should be replaced after about 1 year of 

operation. Ultrapure argon (99.998 percent) 

must be used for the purge gas to minimize

the possibility of fluorescent quenching by

impurities. 

6. Apparatus and Instrumentation 

6.1 Labware. Use clean Type A glass 

volumetric flasks to prepare all solutions. 

Store mercury stock solutions in borosilicateglass bottles with fluoropolymer-lined caps. 

The accuracy of all pipets and volumetric 

flasks should be regularly verified 

gravimetrically or by using an automatic

volume-calibrating spectrophotometer system 

before preparing standard solutions. 

6.2 Instrumentation. Several 

commercial automated CV–AFS systems are

available. These systems are composed of a 

spectrometer, mercury-vapor generator, 

autosampler, and computer. The major

components of the spectrometer are a mercury

line source, a quartz flow-through 

fluorescence cell, and a photometer. The 

mercury-vapor generator is composed of 

several peristaltic pumps and a series of

computer controlled, time-actuated solenoid 

valves that introduce sample and reagents, a 

gas-liquid separator, and a membrane dryer 

tube that removes moisture. The autosampler

and computer automate sample introduction, 

data acquisition, and quantitation. 

7. Reagents and Calibration 

Standards 

All solutions used in this method must be 

verified to have mercury-contaminant 

concentration, after the prescribed dilution, that 

is less than the MDL. ASTM Type I reagent 

water (American Society for Testing and 

Materials, 2000) must always be used to 

prepare solutions. Heating each compound 

used in this method, with the exception of the 

hydrated stannous chloride, in a muffle 

furnace at 250 C for 8 hours will volatilize 

mercury impurities. All solutions must be

stored in designated borosilicate-glass bottles 

with fluoropolymer-lined caps. All 

calibration standards are stable for 1 month. 

7.1 Argon gas is used as the purge

(carrier) gas. Its purity must be at least 

99.998 percent. 

7.2 Nitrogen gas is used as the sheath

gas. Its purity must be at least 99.998 

percent.

7.3 Hydroxylamine hydrochloride

(NH2OH·HCl) Chemical Abstract Service 

Registry Number (CASRN) 5470-11-1. 

7.4 Potassium bromate (KBrO3) 

CASRN 7758-01-2. 

7.5 Potassium bromide (KBr) CASRN 

7758-02-3. 

7.6 Stannous chloride (SnCl2·2H2O) 

CASRN 10025-69-1. 

7.7 Nitric acid solution, 0.4 percent (by

volume). Add 4 mL of concentrated nitric 

acid [HNO3, 16 M, specific gravity (sp. gr.) 

1.41] to 500 mL of water in a 1-L volumetric 

flask. Bring to volume with water. This 

solution is used to rinse between sample

analyses. 

7.8 Hydroxylamine hydrochloride 

(NH2OH·HCl) solution, 15 percent (by

weight). Dissolve 30 g of NH2OH·HCl in a

200-mL volumetric flask that contains about 

10 mL of water. Bring to volume with water. 

Solution must be prepared weekly.

Analytical Method 5 
7.9 Bromine monochloride (BrCl)

solution. This solution is used to oxidize

mercury species in a sample to mercury (II). 

Dissolve 10.8 g of KBr in 1 L of concentrated 

hydrochloric acid (HCl, 12 M, sp. gr. 1.19) in 

a borosilicate-glass bottle. Place a clean 

magnetic stir bar in the bottle and stir for

(NOTE: This process produces copious 

quantities of free halogens, such as Cl2, Br2, 

BrCl.) When all of the KBrO3 has been 

added, the color of the solution should change

from yellow to red to orange. Loosely cap the 

bottle, and stir another hour before tightening.

7.10 Stannous chloride (SnCl2) 

solution, 2 percent (weight-to-volume ratio). 

This solution is used to reduce mercury (II) to 

elemental mercury. Dissolve 20 g of 

SnCl2·2H2O into a 1-L volumetric flask that 

contains 200 mL of water. Slowly add 30 mL

concentrated HCl, mix, and bring to volume 

with water. Allow solution to equilibrate 1 

hour before use. Solution must be prepared 

daily. 

7.11 Primary mercury stock solution, 

10,000 mg/L. National Institute of Standards 

and Technology (NIST) standard reference 

material (SRM) 3133. This reference material 

is stable until the NIST expiration date. 

7.12 Secondary mercury stock solution, 

1.00 mg/L. Pipet 100 L of primary standard 

solution into a 1-L volumetric flask, and 5 mL

of BrCl solution, and bring to volume with 

water. Solution is stable for 1 year. 

7.13 Tertiary mercury stock solution, 

2.0 g/L. Pipet 2.0 mL of secondary stock 

solution into a 1-L volumetric flask, and 5 mL

of BrCl solution, and bring to volume with 

water. Solution is stable for 1 month. 

7.14 Calibration standard 1, 0 ng/L. Add 

1 mL of BrCl solution to a 200-mL volumetric 

flask and bring to volume with water. 

7.15 Calibration standard 2, 5.0 ng/L. 

Add 0.5 mL of the tertiary mercury stock 

solution and 1 mL of BrCl solution to a 200-

mL volumetric flask and bring to volume with 

water. 

7.16 Calibration standard 3, 10.0 ng/L. 

Add 1.0 mL of the tertiary mercury stock 

solution and 1 mL of BrCl solution to a 200-

mL volumetric flask and bring to volume with 

water. 

7.17 Calibration standard 4, 25.0 ng/L. 

Add 2.5 mL of the tertiary mercury stock 

solution and 1 mL of BrCl solution to a 200-

mL volumetric flask and bring to volume with 

water. 

7.18 Calibration standard 5, 50.0 ng/L. 

Add 5.0 mL of the tertiary mercury stock 

solution and 1 mL of BrCl solution to a 200-

mL volumetric flask and bring to volume with 

water. 

7.19 Calibration standard 6, 100.0 

ng/L. Add 10.0 mL of the tertiary mercury

stock solution and 1 mL of BrCl solution to a 

200-mL volumetric flask and bring to volume 

with water.

8. Sample Preparation 

Samples and calibration solutions are

processed identically. Mercury species are 

oxidized to mercury (II) by adding 1 mL of 

BrCl solution (see section 7.9) to 200 mL of 

field-acidified, filtered or unfiltered, naturalwater sample. The sample is digested for at

least 12 hours at room temperature prior to 

Determination of Organic Plus Inorganic Mercury in Filtered and Unfiltered 

Natural Water with Cold Vapor-Atomic Fluorescence Spectrometry

6
analysis. If the yellow color disappears, add 

more BrCl solution to fully digest the sample. 

The sample is completely digested when the

yellow color persists for at least 12 hours. 

Record the total volume of BrCl solution 

added so that the final mercury concentration 

can be adjusted for dilution. Mercury bound 

within microbial cells also might require

ultraviolet photooxidation (U.S. 

Environmental Protection Agency, 1999). 

Some highly organic matrices might require 

longer oxidation times at an elevated

temperature. The sample bottles can be

placed in an oven set at 50 C for 6 hours 

when elevated temperature is required. 

Nevertheless, samples should be analyzed as 

quickly as possible after the oxidation 

procedure. 

Just prior to analysis, 10 mL of BrCloxidized sample is pipetd into an autosampler

borosilicate-glass test tube. Adding 40 L of 

hydroxylamine hydrochloride solution (see 

section 7.8) neutralizes excess BrCl. Allow 

about 5 minutes for the reaction to reach 

completion. The disappearance of the yellow 

color indicates that all the BrCl has been 

destroyed and no traces of halogens remain. 

9. Analytical Procedure 

The analytical procedure for the method 

is fully automated. After calibration 

standards and unknown samples have been 

processed by using the procedure outlined in 

section 8, they are loaded into the 

autosampler. The system introduces a given 

volume of sample and SnCl2 solution (see

section 7.10) with a series of peristaltic 

pumps and solenoid valves. The mercury (II)

present is reduced to elemental mercury that

is purged from the reactor into the 

fluorescence cell with argon gas. The 

mercury concentration is determined by

relating the signal peak area for the samples 

to the peak area for the calibration standards. 

Refer to the NWQL Standard Operating

Procedure IM0348.0 (D.L. Damrau, U.S. 

Geological Survey, written commun., 2001), 

Zellweger Analytics Inc. (1996; 1997), and 

Wendt (1997) for details of the operation and 

maintenance of the instrumentation used in 

this method. Typical operating conditions are 

listed in table 2. 

Table 2. Typical instrument operating conditions

[kPa, kilopascal; lbs/in2

, pounds per square inch; mL/min, 

milliliters per minute; L/min, liters per minute; s, second] 

Parameter Setting

Mercury-vapor generator flow rates 

300 mL/min 

2.75 L/min 

Argon at 276 kPa (40 lbs/in2

) 

Purge (carrier) gas - - - - - - - - - - - - - 

Nitrogen at 276 kPa (40 lbs/in2

) 

Dryer gas - - - - - - - - - - - - - - - - - - - 

Sheath gas - - - - - - - - - - - - - - - - - - 250 mL/min 

Mercury-vapor generator timing 

18 s

170 s

Sample delay - - - - - - - - - - - - - - - - - - 

Wash period - - - - - - - - - - - - - - - - - - 

Load period - - - - - - - - - - - - - - - - - - - 45 s 

Spectrometer 

1,000 

6.50 

Ratio 

Calibration range- - - - - - - - - - - - - - - - 

Fine gain - - - - - - - - - - - - - - -- - - - - - 

Mode switch - - - - - - - - - - - - - - - - - - 

Integration time - - - - - - - - - - - - - - - - 0.25 s 

10. Calculations 

No calculations are required except

when samples are diluted and dilution factors 

are applied. 

11. Reporting Results 

Mercury concentration is reported by

NWQL in micrograms per liter (g/L) instead 

of nanograms per liter (ng/L) because of the 

current (2001) limitations of the data base. 

The number of significant figures reported are 

listed as follows: 

 If the mercury concentration is less 

than the long-term method 

detection level (LT–MDL), then the

Analytical Method 7 
result is reported as less than (<)

the method reporting level (MRL), 

in micrograms per liter. Samples 

between the LT–MDL and MRL

will be “E” coded to indicate an

estimated concentration. 

 If the mercury concentration is 

greater than or equal to the MDL,

but less than 0.100 g/L, then the

result is reported to the nearest 

0.001 g/L.

 If the mercury concentration is 

greater than 0.100 g/L, then the

result is reported to the nearest 

0.01 g/L.

DISCUSSION OF RESULTS 

The CV–AFS mercury method was 

developed to eliminate the use of the acid 

dichromate preservative and to provide a 

method that is capable of measuring ambient 

mercury concentrations in natural water. A 

U.S. Department of Transportation regulation 

on the shipment of natural-water samples 

preserved with acid dichromate requires 

packaging and shipment protocols that are 

expensive. In addition, it is expensive to 

dispose of samples that contain high

concentrations of chromium because it is a 

priority pollutant. Several U.S. 

Environmental Protection Agency (USEPA) 

methods [for example, Method 1631 (U.S. 

Environmental Protection Agency, 1999], use 

hydrochloric acid (HCl) as a preservative. A 

water sample preserved with HCl passes all 

required corrosion tests and can be shipped 

without expensive packaging.

A filtered or unfiltered natural-water

sample is stored in a 250-mL borosilicateglass bottle with a fluoropolymer-lined cap 

after adding 2 mL of 6N HCl to about 200 mL

water (water level at the shoulder of the 

bottle). More acid must be added to samples 

that have high alkalinity to ensure that the pH 

is less than 2. If more than 2 mL of HCl is 

used, the total volume of acid should be listed 

on the Analytical Services Request form so 

that adjustments for dilution can be applied. 

It is important that samples are preserved with

HCl specifically packaged and tested for such 

use. Polypropylene vials that contain HCl 

preservative can be obtained from the USGS 

Ocala Water Quality and Research Laboratory

(part number 06910). The HCl preserves the 

mercury concentration in water samples for at

least 30 days; the relative standard deviation 

of the results was less than 10 percent for a

solution that had 10 ng/L mercury and that 

was analyzed two to three times a week for 1 

month. No significant difference was found 

between the HCl and the previously used 

(prior to April 2001) acid dichromate 

preservatives. The average concentration for 

a sample that had 10 ng/L mercury preserved 

with HCl and that was analyzed repetitively

for 90 days was not significantly different at 

the 95-percent confidence level (p=0.20) than 

the average concentration for a similar

solution preserved with acid dichromate. 

Results obtained for solutions that had 5, 10, 

25, 50, and 100 ng/L mercury (fig. 1) showed 

that the variation, as a function of mercury

concentration, was similar for both 

preservation agents. 

Mercury species in natural water can

take various forms, including elemental 

mercury, mercury complexes, plus various 

alkyl- and phenyl-mercury compounds. 

Oxidation of such species to mercury (II) by

using BrCl makes it possible to determine

concentrations of organic plus inorganic 

mercury and also serves as a preservative to

increase sample-holding times. Experiments 

have indicated that samples are stable for up 

to 3 months after the addition of BrCl. The 

percent-relative standard deviation for a 

sample that had 10 ng/L mercury that was 

analyzed more than 24 times for 3 months 

Determination of Organic Plus Inorganic Mercury in Filtered and Unfiltered 

Natural Water with Cold Vapor-Atomic Fluorescence Spectrometry

8
5 10 25 50 100

0

20

40

60

80

100

120

140

HCl

E

X

P

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RIM

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N

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L M

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C

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Y CONC

E

N

T

RATION,

IN NANO

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LIT

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MERCURY STANDARD REFERENCE MATERIAL

0

20

40

60

80

100

120

140

Acid dichromate

Percentile

 90th

 75th

 50th

 25th

 10th

 = 1st and 99th percentile 

 = Mean 

 EXPLANATION

Figure 1. Results for a series of solutions based on a National Institute of Standards and Technology

standard reference material preserved with acid dichromate or hydrochloric acid (HCl). Standard 

reference material concentration is in nanograms per liter.

Discussion of Results 9 
was about 15 percent, an acceptable level of 

variability considering the concentration level 

and daily deviations in instrument 

performance and calibration standards. 

The bias and variability of the method 

was tested by using standard reference 

materials from several different sources. The 

method performance is validated through the 

repetitive analysis of reagent-water, groundwater, and surface-water matrices spiked at

three different mercury concentrations. 

Comparison of the analytical performance 

between the new CV–AFS method and the 

previously used CV–AAS (prior to April 

2001) is not feasible because the method 

reporting level for the CV–AAS is only about 

100 ng/L. This reporting level is about 20 

times greater than the MDL for the new CV–

AFS method, and 100 ng/L corresponds to the 

upper calibration limit. Onsite samples 

collected before April 2001 were preserved 

with acid dichromate, thus making direct 

comparisons problematic. Furthermore, 

during a typical year, more than 90 percent of 

the results for samples analyzed by CV–AAS

for dissolved and whole-water recoverable 

mercury were less than or equal to 100 ng/L. 

The accuracy of CV–AFS was tested by

using two USGS standard reference water 

samples (SRWS). Calibration standards were 

prepared from NIST SRM-3133 in acid 

dichromate because all available reference 

water samples were preserved in the same

preservative. The SRWSs were diluted by a 

factor of 10 because the most probable mean 

mercury concentration for all available

SRWSs exceeded the calibration range. 

Additional mercury solutions preserved in 

acid dichromate were prepared at 10 and 50 

ng/L from Baker Instra-Analyzed 1,000-

g/mL standard solution to provide another 

reference check. These four reference 

solutions were analyzed repetitively for 2 

months after calibrating the instrument with 

NIST standards preserved with acid 

dichromate. Results for the diluted SRWSs 

compared favorably with expected 

concentrations (fig. 2). The experimental 

results of 38±2 (n=21) and 71±3 ng/L (n=19) 

correspond to the theoretical means of 420 

and 700 ng/L, respectively, after being

adjusted for dilution. The results for the 10- 

and 50-ng/L solutions prepared from the 

Baker Instra-Analyzed standard were 

acceptable at 10±2 and 55±3 ng/L, 

respectively.

Either nitrogen or argon gas has been 

used in analytical methods for mercury to 

purge elemental mercury from the reactor

cell. Experimental results (fig. 3) show that 

there is little difference between the two gases 

when mercury concentrations exceed 10 ng/L. 

The use of argon gas significantly increases 

the precision at 10 ng/L, because the 

sensitivity of the measurement is increased by

about a factor of 2 over nitrogen gas. Thus, 

the use of argon gas is necessary for 

measuring ambient mercury concentrations. 

The accuracy and variability of using argon 

was established by analyzing reference 

standards prepared from two different sources 

repetitively for 1 month. Mercury solutions 

preserved in HCl were prepared from Baker 

Instra-Analyzed 1,000-g/mL (25 and 75 

ng/L) and Spex CertiPrep 10-g/mL (10 and 

50 ng/L) standard solutions. Experimental 

concentrations were determined for these

solutions relative to NIST calibration 

standards. Linear regression analysis (fig. 4) 

of results shows a significant correlation 

between the experimental (Baker and Spex) 

solutions and the theoretical concentrations 

(slope=0.925, correlation coefficient=0.9992). 

The variability ranged from 5 to 15 percent 

and was a function of concentration. In

addition, the stability of the preservation and 

instrumentation is supported by the data that 

were accumulated for a month. 

Determination of Organic Plus Inorganic Mercury in Filtered and Unfiltered 

Natural Water with Cold Vapor-Atomic Fluorescence Spectrometry

10
Hg24 Hg26 10 50

0

20

40

60

80

100

1:10 Dilution

Mean=420 ng/L

Baker

Baker

1:10 Dilution

Mean=700 ng/L

EXPERIM

E

N

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L M

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C

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Y C

O

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CEN

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IN NAN

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GRAM

S

P

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E

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MERCURY STANDARD REFERENCE MATERIAL

 EXPLANATION

Percentile

 90th

 75th

 50th

 25th

 10th

 = 1st and 99th percentile 

 = Mean 

Figure 2. Results for U.S. Geological Survey standard reference water samples Hg24 and Hg26 and 

two dilutions of a Baker Instra-Analyzed standard preserved in acid dichromate. Mercury

concentrations for standard reference materials are in nanograms per liter (ng/L).

Discussion of Results 11 
0

20

40

60

80

100

120

140

Nitrogen

5 10 25 50 100

0

20

40

60

80

100

120

140

Argon

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ERI

M

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TAL M

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C

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C

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RATION,

IN NA

N

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RAM

S

P

ER LIT

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R

THEORETICAL MERCURY CONCENTRATION,

IN NANOGRAMS PER LITER

Percentile

 90th

 75th

 50th

 25th

 10th

 = 1st and 99th percentile 

 = Mean 

 EXPLANATION

Figure 3. The effects of using argon instead of nitrogen as the purge gas in cold vapor–atomic

fluorescence spectrometry. 

Determination of Organic Plus Inorganic Mercury in Filtered and Unfiltered 

Natural Water with Cold Vapor-Atomic Fluorescence Spectrometry

12
0 20 40 60 8

0

20

40

60

80

Slope: 0.925

Y-intercept: 3.3

Correlation coefficient: 0.9992 

EXPERIM

E

N

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CEN

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IN NANO

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THEORETICAL MERCURY CONCENTRATION,

IN NANOGRAMS PER LITER

0

Figure 4. Linear regression analysis of results for sample solutions based on two reference 

standards (experimental) and National Institute of Standards and Technology standard reference 

material (theoretical). Error bars correspond to the standard deviation. 

Discussion of Results 13 
The percentage recovery of results for 

various concentrations of mercury spiked into 

reagent water, filtered and unfiltered surface 

water, and filtered and unfiltered ground 

water was used to validate the method. Data 

were acquired during 7 nonconsecutive days. 

No more than two sets of data were acquired 

in 1 day. The range of average percent

recovery for all filtered and unfiltered 

matrices at 20, 45, and 75 ng/L was 89 to108, 

96 to 103, and 94 to 98 percent, respectively

(table 3). The variability in the recovery of

20 ng/L, a concentration about four times the 

short-term MDL, was about twice that of 

higher spike concentrations. In addition, the 

variability in recoveries at 20 ng/L for the

unfiltered matrices was about twice those for 

corresponding filtered matrices. There was 

little difference, however, in variability for

filtered and unfiltered matrices for spike

concentrations of 45 and 75 ng/L. Sorption to 

particulate matter might contribute to the

increase in variability for the 20-ng/Lmercury spike in unfiltered matrices.

Table 3. Average percent spike recoveries for mercury in reagent-water, filtered and unfiltered surfacewater, and filtered and unfiltered ground-water matrices

[Average is based on 9 to 11 replicates acquired on 7 nonconsecutive days; ng/L, nanograms per liter; %, percent; 

±, plus or minus; filtered, surface or ground water processed through a 0.45-micrometer membrane]

Mercury spike concentration, ng/L 

Matrix 20 45 75

Average 

recovery, %

Average 

recovery, %

Average 

recovery, %

Reagent water - - - - - - - - - - - - - - - - 94±11 100±6 98±8 

Surface water, filtered - - - - - - - - - - - 89±8 100±9 96±6 

Surface water, unfiltered - - - - - - - - - 108±20 101±10 95±8 

Ground water, filtered - - - - - - - - - - - 91±13 96±6 95±9

Ground water, unfiltered - - - - - - - - - 102±20 103±10 94±7 

CONCLUSIONS

The new cold vapor–atomic

fluorescence spectrometric (CV–AFS)

method for the determination of inorganic 

plus organic mercury species in filtered and 

unfiltered natural water has several 

advantages when compared to the previously

used (prior to April 2001) cold vapor–atomic

absorption spectrometric method. Primary

advantages are improved method detection 

limit and elimination of a corrosive

preservative. The CV–AFS method has a 

short-term method detection limit of 5 ng/L

without using gold amalgamation for mercury 

preconcentration. Without mercury

preconcentration, analysis throughput is 

increased while maintaining the capability of

determining mercury at concentrations 

typically regulated by water-quality standards. 

Mercury samples are preserved with 

hydrochloric acid (HCl) instead of acid 

dichromate. Although acid dichromate has 

been used extensively to preserve mercury

samples, shipping and disposal regulations 

have made continued use too expensive. 

Experiments suggest that there is no 

significant difference, however, between the

Determination of Organic Plus Inorganic Mercury in Filtered and Unfiltered 

Natural Water with Cold Vapor-Atomic Fluorescence Spectrometry

14
average mercury concentration in a water 

sample preserved with HCl and one preserved 

with acid dichromate. Ambient mercury

concentrations in samples preserved with HCl 

in borosilicate-glass bottles with fluoropolymer-lined caps have been shown to be 

stable for at least 30 days. Oxidation of 

inorganic and organic mercury species with 

bromine monochloride makes it possible to 

determine total recoverable mercury and 

extends the sample-holding time to 3 months. 

The bias and variability of the method 

were established using standard reference 

materials and spike recovery results. 

Comparison of the analytical performance 

between CV–AFS and the previously used 

method were not feasible because of the 

sizable disparity in method detection limits. 

The concentration of the upper calibration 

standard used by the new method corresponds 

to the detection limit of the old method. 

Results for USGS standard reference water 

samples were acceptable with respect to the 

most probable concentration. Other solutions 

that have varying mercury concentrations 

were prepared from certified standards from

two different sources. Linear regression of 

the results for these solutions were

significantly correlated (correlation 

coefficient=0.9992) with standard solutions 

traceable to National Institute of Standards 

and Technology materials. The analytical 

variability is expected to extend from 15 to 5 

percent with increasing concentration 

bounded by the calibration range. The 

average percent recovery of 20, 45, and 75 

ng/L of mercury in reagent-water, surfacewater, and ground-water matrices was 

determined during 7 nonconsecutive days. 

The average percent recoveries for all filtered 

and unfiltered matrices ranged from 89 to108 

(20 ng/L), 96 to 103 (45 ng/L), and 94 to 98 

percent (75 ng/L). The variability for the

recovery of the 20-ng/L spike, a concentration 

about four times the method detection limit, 

was about twice that of higher spike 

concentrations. In addition, the variability in 

percentage recoveries for unfiltered matrices

was twice that for matching filtered matrices, 

possibly because of sorption to particulate 

matter.

Analytical results from the new CV–

AFS method will most likely impact 

interpretation of long-term trends observed in 

water-quality studies involving mercury. The 

method detection limit of 5 ng/L is about a 

factor of 20 times lower than the previously

used method. More than 90 percent of the 

mercury concentrations reported with the 

previous analytical method during a typical 

year were less than or equal to 100 ng/L prior 

to April 2001. Implementation of the CV–

AFS method will likely provide more 

definitive results because its analytical range 

extends well below 100 ng/L. 

REFERENCES CITED 

American Society for Testing and Materials, 

2000, Annual Book of ASTM 

Standards, Section 11, Water: 

Philadelphia, American Society for 

Testing and Materials, v. 11.01, 

D1193, p. 10. 

Bloom, N.S., 1995, Trace metals and ultraclean sample handling: Environmental 

Laboratory, v. 7, p. 20. 

Fishman, M.J., 1993, Methods of analysis by

the U.S. Geological Survey National 

Water Quality Laboratory—

Determination of inorganic and 

organic constituents in water and 

fluvial sediments: U.S. Geological 

Survey Open-File Report 93-125, 

217 p. 

Fishman, M.J., and Friedman, L.C., eds., 1989, 

Methods for determination of inorganic 

substances in water and fluvial 

sediments: U.S. Geological Survey

Techniques of Water-Resources 

Investigations, Book 5, Chap. A1, p. 

289–291.

References Cited 15 
Horowitz, A.J., Demas, C.R., Fitzgerald, 

K.K., Miller, T.L., and Rickert, D.A., 

1994, U.S. Geological Survey

protocol for the collection and 

processing of surface-water samples 

for the subsequent determination of 

inorganic constituents in natural 

water: U.S. Geological Survey

Open-File Report 94-539, 57 p. 

Olson, M.L., and DeWild, J.F., 1999, Lowlevel collection techniques and 

species-specific analytical method for 

mercury in water, sediment, and 

biota: U.S. Geological Survey

Water-Resources Investigations 

Report 99-4018b, 11 p.

U.S. Environmental Protection Agency, 1996, 

Method 245.7—Determination of 

ultra-trace level (ng Hg/L) total

mercury in water by cold vapor 

atomic fluorescence spectrometry: 

U.S. Environmental Protection 

Agency, Revision 1.1, National 

Exposure Research Laboratory,

Research Triangle Park, Office of 

Research and Development, 29 p. 

_______1999, Method 1631—Revision B:

Mercury in water by oxidation, purge

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fluorescence spectrometry: EPA 

821-R-99-005, May 1999, 33 p. 

_______2000, Guideline establishing test 

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pollutants (Part 136, Appendix B. 

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U.S. Code of Federal Regulations, 

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Wendt, Karin, 1997, QuikChem Method 10-

138-37-1-A, Determination of mercury

by cold vapor generation and atomic 

fluorescence detection: Milwaukee, 

Wisconsin, LaChat Instruments, 20 p. 

Wilde, F.D., Radtke, D.B., Gibs, 

Jacob, and Iwatsubo, R.T., eds., 

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Zellweger Analytics Inc., 1996, QuikChem 

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_______1997, QuikChem 8000 automated ion 

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April 22, 1997, 64 p. 

Determination of Organic Plus Inorganic Mercury in Filtered and Unfiltered 

Natural Water with Cold Vapor-Atomic Fluorescence Spectrometry

16