Reference NBS

PUBLICATIONS AlllDb DBltOl

NBSIR 85-3295

A Comprehensive Method for the

Determination of Aquatic Butyltin

Species at Ultratrace Levels Using

Simultaneous Hydridization /Extraction

With GC-FPD

G. J. Olson

F. E. Brinckman

U.S. DEPARTMENT OF COMMERCE

National Bureau of Standards

Institute for Materials Science

Ceramics Division

Gaithersburg, MD 20899

and

C. L. Matthias

J. M. Bellama

Chemistry Department

University of Maryland

College Park, MD

December 1985

Final Report

Prepared for —'d Taylor Naval Ship Research and Development Center

OC polls, MO 21402

100

• 056

85-3295

1985

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NBSIR 86-3296 CiUOO t • 9 A COMPREHENSIVE METHOD FOR THE . U>

DETERMINATION OF AQUATIC BUTYLTIN ' SPECIES AT ULTRATRACE LEVELS USING / ? J

^

SIMULTANEOUS HYDRIDIZATION/EXTRACTION

WITH GC-FPD

G. J. Olson

F. E. Brinckman

U.S. DEPARTMENT OF COMMERCE

National Bureau of Standards

Institute for Materials Science

Ceramics Division

Gaithersburg, MD 20899

and

C. L Matthias

J. M. Bellama

Chemistry Department

University of Maryland

College Park, MD

December 1986

Final Report

Prepared for David Taylor Naval Ship Research and Development Center

Annapolis, MD 21402

U.S. DEPARTMENT OF COMMERCE, Malcolm Baldrige, Secretary

NATIONAL BUREAU OF STANDARDS. Ernest Ambler. Dirmctor
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A COMPREHENSIVE METHOD FOR THE DETERMINATION OF AQUATIC BUTYLTIN AND

BUTYLMETHYLTIN SPECIES AT ULTRA-TRACE LEVELS USING SIMULTANEOUS

HYDRIDIZATION/EXTRACTION WITH GC-FPD

1.0 Introduction

The increasingly diverse and pervasive use of butyltin compounds in

industrial, aquatic, and agricultural applications has led to concern

regarding the impact of these alkyltin compounds on the environment and

prompted recognition of the need for reliable environmental analytical

monitoring methods (1-3). Aquatic uses of organotin compounds, in

particular their incorporation as tributyltin biocides in controlledrelease paints on ships, represent a phenomenal growth area in its

infancy, yet pose the greatest immediate impact upon harbor and coastal

aquatic biota (4). Tributyltin species are very effective against common

marine fouling organisms such as barnacles. However, it is clear that

tributyltin is also highly toxic to various non-target aquatic organisms

at low concentrations. For example, tributyltin at low parts per billion

(ppb) levels is acutely toxic to amphipod larvae (5); lobster larvae and

zoeal shore crabs (6), sheepshead minnows (7), and mysid shrimp (8). At

sub-ppb levels, tributyltin causes sublethal effects in zoeal mud crabs

(9), mussel larvae (10) and copepods (11). Part of the presumed

redeeming quality of tributyltin in such environmental uses rests in its

degradation by Sn-C cleavage to comparatively innocuous di- and

monobutyltin and inorganic tin residues. Mono- and dibutyltins are less

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toxic than tributyltin to marine biota, consistent with the general trend

for RjjSn^ ; which is increasing toxicity with increasing molecular

size (12) from n=1 to n=3 . and marked decrease in toxicity for n=4 (4).

Inorganic tin is virtually non-toxic and may be an essential trace element

in animals and man (13).

The degradation of tributyltin in the marine environment is widely

assumed to follow a stepwise debutylation (4):

Bu^Sn"^ -» Bu2

Sn^'^ ^ BuSn^"^ -*• Sn(IV) (Eq. 1)

The butyl groups may be oxidized to CO2 by microbial activity (14).

However, no consistent, quantitative picture of the persistence of the

tributyltin species in its aquatic service environment has yet emerged.

Complicating the issue of persistence is the possibility of other

degradation pathways for tributyltin species including a number of

possible redistribution reactions catalyzed by environmental molecules

such as amines or sulfides (15) or other reactants (16). The possibility

of environmental methylation of butyltins has been raised by a recent

report of the presence of mixed butylmethyltin species in sediments (17),

presumably arising by biological methylation of anthropogenic butyltins.

This suggests additional pathways for tributyltin in the aquatic

environment. The toxicity of many of the products of these methylation

pathways is still unknown. A few of the possible Sn-C reactions include:

2Bu2Sn"^ * Bu2Sn^"^ + Bu^^Sn (Eq.2)

BU2

Sn^'^ + Bu^Sn"^ ^ BuSn^"^ + Bu^Sn (Eq.3)

Bu^Sn'*' + Me” ->• Bu^MeSn (Eq.4)

3
The source of methylcarbanion (Eq.4) presently is uncertain but may

be due to redistribution with biogenic or anthropogenic methyltin species

(18) or by intermediate oxidative methylation from algal metabolites

(19,20).

In sum, it has been assumed that toxic anthropogenic organotin

species in the environment lose their potency by degradation to less toxic

organotin species and, finally, to innocuous inorganic tin(IV). Analytical

methods reported heretofore, nevertheless, do not adequately test the

validity of the presumed degradation pathway. For example,

chromatographic procedures relying on charge separation fail to determine

neutral species such as tetrabutyltin or dibutyldimethyltin. Moreover,

laboratory derivitization using methyl Grignard reagents (21) obscure

butylmethyltin species, if present.

Clearly, any analytical method for adequate environmental monitoring

of tributyltin must be capable of determining at very low concentrations

(ng/L levels) the entire set of both butyltin and mixed butylmethyltin

species in order to assess the environmental fate of tributyltin. Because

of the increase in the world use of tributyltin as the active agent in

marine biocides, the need also exists for frequent routine monitoring of

areas of high ship traffic, such as marinas and harbors, and of sensitive

areas, such as oyster beds. The analytical method used for the large

number of samples that such monitoring would generate must be relatively

simple and fast, as well as highly sensitive.

Previous methods for the determination of butyltins have relied

primarily upon derivatizatlon to form hydrophobic, neutral organotin

species. Some investigators used reduction of equated alkyltin species

4
with sodium borohydride (NaBH]|) to the covalent hydrides

Bu„Sn(''-")*(3q) * H- - Bu„SnH(4.n) (^<5- 5)

followed by inert gas purge of the aqueous sample. The evolved volatile

alkyltln hydrides were trapped at liquid nitrogen temperatures, followed

by the gradual warming of the cold trap to allow the separation of

alkyltins by boiling point, and finally, tin was detected absorption

spectroscopy (22, 23, 24). Others have used extraction of aqueous samples

into organic solvents and alkylation of the extract with a Grignard

reagent to produce neutral tetraalkyl compounds which can then be

separated and identified using gas chromatography coupled with mass

spectrometric (GC-MS) (21) or flame photometric detection (GC-FPD) (25).

Liquid chromatography coupled with an atomic absorption detector has also

been employed for butyltin speciation (26). The recent work of Mueller

(27) employed a macroreticular resin for concentration and separation of

tributyltin from water followed by methylation and detection by GC-FPD and

GC-MS.

In the present work, we describe a relatively rapid, simple,

sensitive method for the complete determination of butyl- and mixed

butylmethyltin species in the water environment. The butyltin species are

reduced to their respective volatile hydrides with aqueous NaBH|| and

simultaneously extracted into CH2 CI2 for appropriate pre-concentration and

subsequent chromatographic analysis.

5
2.0

Erperimental

2.1 Materials

All glassware and Teflon separatory funnels were cleaned prior to use

by rinsing in methanol, washing with laboratory detergent and leaching

with warm 10$ nitric acid for at least 8 hr, followed by rinsing with

copious amounts of deionized water (18 MOcm). All other plasticware parts

were similarly cleaned except that the acid leaching time was shortened to

one hour. The Bu^SnCl(i|^j^) compounds and Bu^SnH used for preparation of

standard solutions and spikes (95*-98$ purity, Alfa Products, Danvers, MA)

were used as received without further purification. Di-n-propyltin

dichloride (M & T Chemicals, Inc., Rahway, NJ) was used as an internal

standard. Chromatographic grade dichloromethane was obtained from Burdick

and Jackson Laboratories, Inc. (Muskegon, MI). In addition, a dilute

(ppm), aqueous tributyltin research material was prepared chromatographically in our NBS laboratory (28). Organotin solutions were prepared at

concentrations of approximately 1000 mg/L as tin in spectrograde methanol.

Deionized water of 15-18 Mflcm resistivity obtained from a Milli-Q Reagent

Grade water system (Millipore Corp, Bedford, MA) was used to dilute these

stock solutions to the working range of approximately 550 ng/ml (0.5 ppm).

Fresh solutions of 4$ (w/v) sodium borohydride (Aldrich Chemical Co.,

Inc., Milwaukee, WI) were prepared daily in deionized water. No butyltin

species were detected in the reagent blanks.

2.2 GC-FPD System

A Hewlett-Packard (Avondale, PA) (HP) Model 5730 gas chromatograph

equipped with a HP flame photometric detector was used for this study.

Chromatographic separations were carried out on a 2-mm i.d. X 6 ft. glass

6
column packed with 1.5J OV-101 (liquid methyl silicone) on Chromosorb G

HP( 100-200 mesh size) (Varian, Sunnyvale, CA). A hydrogen-rich flame was

employed, supported by H

2 flowing at the measured rate of 110 mL/min, air

at 70 mL/min, and N

2 (zero grade) carrier gas at 20 mL/min. The FPD was

equipped with a 600-nm cut-on interference filter (band-pass 600-2000 nm)

(Ditric Opitics, Inc., Hudson, MA) to monitor the SnH molecular emission

(29, 30 ). The output signal from the FPD was recorded simultaneously on a

strip chart recorder and an integrator-plotter (HP Model 3390A). For all

runs reported herein, the column temperature was programmed at 23® C for

2 min. then heated to 170® C at 32® C/min. The detector temperature was

maintained at 200® C and injection port at 150® C.

2 . 3 GC-MS System

The GC-MS system is described in ref. 29 with the following

modifications for use with butyltins. The GC column was that described

above for the FPD system. For tetrabutyltin analysis, samples were

extracted into CH2 CI 2 without hydridization. The temperature program for

GC-MS tetrabutyltin was 50® C for 1 min. then to 170 ®C at 30 ®C/min.

The GC-MS system is interfaced with an on-line Computer Automation,

Inc. computer, and software from Teknivet Inc. (St. Louis, MO). This

system provided two modes for data acquisition; mass-spectrum mode and

selected ion monitoring. Major representative peaks were selected from

fragmentation patterns of the mass spectrum of tetrabutyltin (m/e - 119,

121, 177 , 179 , 233 , 235 ) for selected ion monitoring. Total ion current

mass spectra were obtained for tetrabutyltin and the mixed species.

2.4 Redistribution reactions

The mixed butylmethyltin species were prepared after the method of

Calingaert e^ ^. , (31) by refluxing at 80®C 5 mM of the appropriate

7
methyl- and butyltin chloride starting materials (Alfa Products) with

0.5 mM aluminum chloride catalyst (Fisher Scientific Co., Fairlawn, NJ) in

30 mL HPLC grade hexane (Fisher Scientific). The reactions were run for

5-6 hrs under N

2

. Aliquots of the reaction mixtures were hydridized as

CH2 CI2 dilutions in a two phase system with aqueous NaBHi^ and analyzed by

GC-FPD to determine the retention times of the various mixed butylmethyltin products. The reaction products were identified as butylmethyl-,

butyldimethyl, dibutyl- and dibutylmethyl-, dibutylmethyl- and

tributyltin, and confirmed by GC-MS.

2.5 Analysis Procedure for Butyltins

For a typical analysis of saline water with a butyltin concentration

in the sub-ug/L range (as tin), the following procedure was found to be

optimal. To 100 mL of sample in a 125 mL glass separatory funnel equipped

with a Teflon stopcock and Teflon-lined screw top (Wheaton Scientific,

Millville, NJ), were added 2.8 mL dichloromethane and 2.0 mL of 4% (w/v)

aqueous NaBHi|. In addition, a 1 0 uL spike of a 0.5 ppm aqueous solution

of di-n-propyltin dichloride was added to certain samples as an internal

standard. The funnel was capped and shaken by hand for 1 min, vented, and

then shaken (240 strokes/min) on a wrist action shaker (Burrel Corp.

,

Pittsburgh, PA) for 10 min. Following a 5 min settling period, the lower

organic layer was removed. An additional 1.4 mL of dichloromethane was

added and the extraction procedure repeated. The organic layers were

combined (approx. 2 mL) in polypropylene centrifuge tubes and evaporated

to 100-200 jiL or less under a gentle stream of air. Appropriate reagent

blanks were carried through the entire procedure. All quantitation was

achieved using the method of standard additions to the sample matrix. For

8
samples of concentration greater than 500 ng Sn/L, no evaporation

concentration step was required. While most of our work has been done

using the 100 mL sample size, samples of up to 800-1000 mL have been

analyzed using 1L Teflon screw-capped separatory funnels (Fisher

Scientific) and proportionately larger volumes of all reagents. Extracts

were concentrated to 50-100 pL in 15 mL glass centrifuge tubes (Wheaton).

2.6 EnvlronjDDental Samples

Environmental samples were collected aboard the research vessel

Ridgely Warfield or from docks or piers. Surface water samples were

collected in 4 liter glass bottles at 1 m depth. Surface microlayer

samples were collected by gently dipping a Teflon sheet (0.32 m ) to the

water surface and by rinsing of the adsorbed sample into a glass bottle

with about 25 mL of deionized water. Sampling station designations are

those of the Chesapeake Bay Institute. Samples from San Diego were

provided by Dr. P. F. Seligman and were stored on dry ice until analysis.

3.0 Results and Discussion

3.1 Solvent Choice and Hydridization

Dichloromethane is often the solvent of choice for extracting of

organic compounds from natural waters (32) and is effective in extracting

of organotins from tissues (8). Preliminary experiments showed that the

efficiency of dichlorome thane in extracting Bu^SnCli|^^ (n=1-4) from

deionized water (1 pg Sn/L) ranged from 60 % for tetrabutyltin to 95J for

tributyltin cation after a single 10 min extraction (unpublished data).

Hydride derivatization was required to insure sufficient volatility for

9
the GC analysis of butyltin species in the dichlorome thane extracts Fig.

1, top). The highest sample recoveries were obtained with simultaneous

extraction and hydridization as opposed to a conventional two-step

extraction and derivatization sequence (Fig. 1). The extraction

efficiency of the new procedure for tributyltin was determined by spiking

at 1 ppb with tributyltin chloride a 100 mL Chesapeake Bay water sample of

low intrinsic butyltin concentration and performing the simultaneous

hydridization/extraction. A calibration curve of peak area vs. ng of

tributyltin hydride was prepared with dilute dichloromethane solutions of

commercially available neat tributyltin hydride (98? purity, Alfa

Products, Inc.). The amount of tributyltin recovered in the extraction

process was determined using this curve. Extraction efficiency from the

environmental matrix was 1 1 (±10%) recovery of the spikes of tributyltin

cation. The possibility that the analytical work up might induce either

degradation or rearrangement reactions also has been of great concern. A

severe limitation for analysts has been the lack of any tributyltin

reference material for use in evaluating analytical schemes. A research

material of aqueous tributyltin has been prepared chromatographically at

NBS (28) and analyzed repeatedly at concentrations from the range of mg/L

to < 50 ng/L with no evidence of any degradation or rearrangement

products. Spikes of dibutyltin added to dilutions of the reference

material indicate that at a concentration of 1 \xg/L, detection of <3%

conversion to the dibutyltin degradation product would be readily achieved

(Fig. 2).

3.2 Detection Limits and Uncertainty

Calibration curves and the limits of detection at the 95% confidence

interval (33) for the analysis procedures for four major butyltin species

10
are shown In Figure 3. The detection limits of the method were determined

by analysis of spiked samples of 100 mL volume, with the dichloromethane

extract concentrated to 0.05 mL to give a preconcentration factor of

5 X 10^. The absolute detection limits for the GC-FPD instrument

(as ng Sn) were also determined, by hydridization of dichloromethane

solutions of known butyltin concentration (Fig. 3). Scale up to 800-

1000 mL samples resulted in approximately 5-6 fold improvement in

detection limits, with detection of 1-2 ng Sn/L as tributyltin and

tetrabutyltin and less than 1 ng Sn/L as dibutyltin being possible for

these larger samples. The resulting 5-6 fold lowering of detection limits

with an 8-10 fold increase in sample volume could in part be explained by

loss of the analyte to the separatory funnel walls, because of the large

surface area.

Additionally, the extraction efficiency may be reduced for these

large volumes because the large funnel could not be shaken as vigorously

as the 125 mL funnel with the automatic mechanical shaker.

The detection limit for environmental samples is dependent upon the

degree of extract concentration prior to analysis. Consequently, sample

loss on evaporation was also investigated. In these experiments, 1 mL of

the hydridized dicloromethane extract of Chesapeake Bay water spiked with

1 ug/L (n = 1-4) was evaporated under a gentle stream of gas

to 50 uL, then re-diluted to 1 mL with fresh dichloromethane. Comparison

between dry air and N

2 and ambient temperature (22°C) and 0®C

(ice-water bath) for use in evaporation were evaluated. Peak areas for

the tin species before and after the concentration step were compared.

Under all conditions, minimal losses (0-12$) on evaporation were seen for
di“, tri“ and tetrabutyltin, approximately 50$ losses were seen for

monobutyltin. For convenience, air and ambient temperature were used on

all subsequent analyses.

Reproducibility of the procedure was determined by carrying six 100

mL aliquots of the same sample through the entire analytical procedure.

Instrumental reproducibility was estimated by replicate (n=7) injections

of the same sample (Table I). Reproducibility of the analytical scheme

appears to be instrument limited.

The sources of potential errors in the analysis of complex

environmental samples are numerous. Errors in environmental analysis of

aqueous butyltins at parts-per-trillion concentrations result from sample

handling, storage and derivitization steps and can be attributed to the

physical and chemical properties of the butyltin analytic within the

matrix, as well as to instrumental limitations. For the analytical scheme

described here, the overall analytical error is the sum of individual

sources of error, i.e., sample losses to glass or Teflon, partition onto

suspended matter, efficiency of hydridization, efficiency of extraction

detector variability and syringe injection reproducibility. While this

list of errors is extensive, it is by no means exhaustive. It is possible

to minimize the effects of some of these potential errors by analyzing

samples immediately after collection, by the use of an internal standard

(dipropyltin dichloride) and quantitation using standard additions.

3*3 Application to Aquatic Environmental Samples

Because of the possibility of wide variations in pH and salinity of

natural waters, the analysis procedure was carried out at several pH and

salinity levels. Figure 4 shows that neither salinity nor pH

12
significantly affect simultaneous extraction and detection of butyltin

species in Chesapeake Bay waters. Butyltins spiked into sample waters

obtained from widely different locations in the Bay, representing

salinities ranging from 0 (deionized water) to 22.4 0/00, were recovered

with similiar efficiency. Adjustment of sample pH (mid-Bay site 858C,

pH - 7.7) to pH 6.3 with 1.0M HCl or to 9.4 with 1 .OM NaOH made little

difference in detection of butyltin species. These pH values exceed the

range expected for estuarine or marine samples. Evaluation of the

extractions from different pH or salinity samples as replicates shows

standard deviations of 3.2-12.3? for pH variation and 9.5-16.2? for

salinity variation calculated from peak areas of the butyltin species.

These values are comparable to the standard deviations for replicate

analyses of the same sample reported above (Table I) and for this reason

pH and salinity do not appear to contribute important errors in this

analysis.

Several environmental samples have been analyzed, including bulk

waters samples from Shelter Island (San Diego Harbor) and from Jones Falls

(Baltimore Harbor) (Fig. 5) and microlayer samples from Jones Falls.

Tributyltin concentrations of 68 ± 1 1 ng Sn per liter and dibutyltin

concentrations of 108 ± 19 ng Sn per liter were found in the San Diego

samples comparing closely with levels reported by Seligman £t (8, 22).

Tributyl- and dibutyltin species were also detected in surface waters from

commercial marinas at Annapolis, MD (Table II). Of interest in the Jones

Falls microlayer samples is a peak with retention time corresponding to

that of tetrabutyltin (Fig. 6). In addition, a peak with the same

retention time as tetrabutyltin is seen in washwater from samples

collected during dry-dock cleaning of organotin paints ship hulls.

13
Identification of this peak as tetrabutyltin was confirmed by GC-MS (Fig.

7). The peaks on the mass spectrum at m/e 205 correspond to a large

non-tin peak which is not completely resolved chromatographically from the

tetrabutyltin peak (Fig. 8). This is the first report of tetrabutyltin in

the environment, and suggests that the tributyl tin transformation

reactions shown in Eq. 2,3 may occur in harbor and marina waters. The

hydrophobic nature of the surface microlayer and its elevated levels of

microorganisms and metabolites is well documented (3^). The surface

microlayer may contain components capable of inducing redistribution

reactions of butyltins, e.g. by a mechanism similar to the Lewis base

induced redistribution mechanism for conversion of (CH

2

) 3 SnOH to (CH

2

) 4 Sn

described by Guard and co-workers (35). The microlayer is a preferred

sink for tetrabutyltin due to tetrabutyltin 's low water solubility and low

volatility.

The ability to detect ultratrace concentrations of environmentally

methylated anthropogenic butyltin species is paramount for fully

understanding the fate of tributyltin in the environment, and is achieved

with this analytical scheme. We have observed several small peaks in

environmental samples which suggest the presence of mixed methylbutyl

species in estuarine and harbor waters, but due to very low concentrations

(< 1 ng/L) of analyte and relatively poor sensitivity of the GC-MS system

we have not as yet confirmed the identity of these peaks. This work is

continuing.

The new technique was also used to detect biodegradation products of

tributyltin spiked into natural waters. Here, (Fig. 9) a sample of

Baltimore Harbor (Chesapeake Bay) water was amended with yeast extract

(0.05$ final concentration) to promote microbial activity and the MBS

14
research material tributyltln (100 iJg/L, final concentration). After 12

weeks incubation at 20 ®C, substantial degradation of tributyltin to

dibutyl and raonobutyltin species was noted. No mixed butylmethyltin

species were detected. A sterile control showed very little tributyltin

degradation. Further studies are now in progress to measure tributyltin

degradation rates in natural waters. These experiments employ low levels

of tributyltin (ca. 1 ug/L) and short incubation times (days).

4.0 ConclusIons

This approach to the analysis a butyltins in aquatic samples at

environmental action levels (ng/L) has been shown to be highly sensitive,

simple and non-disruptive to the molecular speciation of the sample.

Clearly, instant partitioning into the polar organic medium (CH2CI2

)

immediately after hydridic derivatization of aquated species

greatly favors analytical sensitivities and precision. Currently, we are

attempting to expand the scope and utility of the analytical method to

sediments in order to evaluate more fully the fate of tributyltin in the

estuarine environment.

5.0 Acknowledgements

This work was supported in part by the Office of Naval Research and

by the David Taylor Naval Ship Naval Research and Development Center,

Bethesda, MD. We thank Dr. Harold E. Guard for advice on gas

chromatography columns, Dr. Peter Seligman of the Naval Ocean Systems

15
Center for environmental samples, and William R. Blair for assistance with

the GC“MS.

Certain commercial products or equipment are mentioned in order to

adequately describe experimental procedures. In no case does such

identification imply endorsement by the National Bureau of Standards, nor

does it imply that the material is necessarily the best available for the

purpose. Contributions from the National Bureau of Standards are not

subject to copyright.

This work will be included in the dissertation of C.L.M. to be

submitted as a requirement for the Ph.D. degree from the University of

Maryland.

6.0 References

1. Federal Register, 47 , 54624 (1982). [Notice of addition of alkyltins

to EPA Priority Pollutants List under TSCA]

2. Thompson, J. A. J. , Sheffer, M. G. , Pierce, R. C., Chau, Y. K.

,

Cooney, J. J., Cullen, W. R., Maguire, R. J. "Organotin Compounds in

the Aquatic Environment: Scientific Criteria for Assessing Their

Effects on Environmental Quality," National Research Council of

Canada, NRCC Associate Committee on Scientific Criteria for

Environmental Quality, (1985).

3. "Investigation of the Effects and Uses of Biocides and Related

Compounds in North Carolina," (1983). Division of Environmental

Management, Water Quality Section, Department of Natural Resources

and Community Development, State of North Carolina.

4. Blunden, S. J., Hobbs, L. A., Smith, P. J. The Environmental Chemistry

of Organotin Compounds, in: Environmental Chemistry, H. J. M. Bowen

ed.. Royal Society of Chemistry, London, pp. 51-77 (1984).

5. Laughlin, R. B., Nordlund, K, Linden, 0. Long-term effects of

tributyltin compounds on the Baltic amphipod. Mar. Environ. Res.,

12, 243-271 (1984).

16
6. Laughlin, R. B., French, W. J. Comparative study of the acute

toxicity of a homologous series of trialkyltins to larval shore

crabs. Bull. Environ. Contam. Toxicol., 25 , 802-809 (1980).

7. Ward, G. S., Cramm, G. C., Parrish, P. R. , Trachman, H. , Slesinger,

A. Bioaccumulation and Chronic Toxicity of Bis( tr ibutyltin)oxide

(TBTO): Tests with a Saltwater Fish, in: Aquatic Toxicology and

Hazard Assessment: Fourth Conference, ASTM STP 737, D. R. Branson

and K. L. Dickson, eds., American Society for Testing and Materials,

Philadelphia, pp. 183-200 (1981).

8. Seligman, P. F. Fate and Effects of Organotin Antifouling Leachates

in the Marine Environment, Progress Report prepared for Energy

Research and Development Office, David W. Taylor Ship Research and

Development Center, 47 pp(1984).

9. Laughlin, R. B., French, W. , Guard, H. E. Acute and sublethal

toxicity of tributyltin oxide (TBTO) and its putative environmental

product, tributyltin sulfide (TBTS) to Zoeal mud crabs,

Phlthropanspeces harrisii . Water, Air Soil Pollut. 20 , 69-79 (1983).

10. Beaumont, A. R., Budd, M. D. High mortality of the larvae of the

common mussel at low concentrations of tributyltin. Mar. Poll. Bull.

_1_5, 402-405 (1984).

11. U'Ren, S. C. Acute toxicity of bis( tributyltin)oxide to a marine

copepod. Mar. Poll. Bull. 21. 303-306 (1983).

12. Laughlin, R. B. , French, W. , Johannesen, R. B. , Guard, H. E.

,

Brinckman, F. E. Predicting toxicity using computed molecular

topologies: the example of triorganotin compounds. Chemosphere, 1

3

, 575-584 (1984).

13* Cardarelli, N. R., Cardarelli, B. M. , Marioneaux, M. Tin as a vital

trace nutrient. J. Nutrition, Growth and Cancer j_, 1 81-1 94 (1 983).

14. Barug, D. Microbial degradation of bis( tributyltin)oxide.

Chemosphere J_0, 1 1 45-1 154 (1981 ).

15. Craig, P. J., Rapsomanikis, S. New route to tris(dimethyltin

sulfide) with tetramethyltin as co-products: wider implictlons. J.

Chem. Soc., Chem. Comraun. 114 (1982).

16. Manders, W. F., Olson, G. J., Brinckman, F. E., Bellama, J. M. A novel synthesis of methyltin triiodide with environmental

implications. J. Chem. Soc., Chem. Commun. 538-540 (1984).

17. Maguire, R. J. Butyltin compounds and inorganic tin in sediments in

Ontario. Environ. Sci. Technol. 2l» 291-294 (1984).

18. Thayer, J. S., Brinckman, F. E. The biological methylation of metals

and metalloids. Adv. Organomet. Chem. 20, 313-356 (1982).

17
19. Olson, G. J., Brinckman, F. E., Thayer, J. S. lodome thane as a potential metal mobilizing agent in nature. Environ. Sci. Technol.

j_8, 726-729 (1 984).

20. Rapsomanikis, S., Weber, J. H. Environmental implications of tin(II)

and methyltin( IV) ions in the presense of manganese dioxide.

Environ. Sci. Technol. j_9, 352-356 (1985).

21. Meinema, H. A., Burger-Wiersma, T. , Versluis-deHaan, E. F. , Severs,

E. C. Determination of trace amounts of butyltin compounds in

aqueous systems by gas chromatography/mass spectrometry. Environ.

Sci. Technol. J_2, 288-293 (1 978).

22. Braman, R. S., Tompkins, M. A. Separation and determination of

nanogram amounts of inorganic tin and methyltin compounds in the

environment. Anal. Chem. 12-19 (1979).

23. Hodge, V. F. , Seidel, S. L. , Goldberg, E. D. Determination of

tin(IV) and organotin compounds in natural waters, coastal sediments

and macro algae by atomic absorption spectrometry. Anal. Chem.

1256-1259 (1 979).

24. Valkirs, A. 0., Seligman, P. F. , Taylor, V, Stang, P. M. , Lieberman,

S. H. Personal communication.

25. Maguire, R. J., Huneault, H. J. Determination of butyltin species in

water by gas chromatography with flame photometric detection. J.

Chromatogr. 209 , 458-462 (1981).

26. Jewett, K. L. , Brinckman, F. E. Speciation of trace di- and

triorganotins in water by ion exchange HPLC-GFAA. J. Chromat. Sci.

19, 583“593 (1981).

27. Mueller, M. D., Butyltin detection at tree levels in water and

sediments using GC with flame-photometric detection and GC-MS.

Pres. Z. Anal. Chem. 317, 32-36 (1984).

28.

Blair, W. R., Olson, G. J., Brinckman, F. E. "An International

Butyltin Measurement Methods Intercomparison: Sample Preparation and

Results of Analysis." NBS Special Publication, in preparation.

29 Jackson, J. A., Blair, W. R., Brinckman, F. E. , Iverson, W. P. Gas

chromatographic speciation of raethylstannanes in the Chesapeake Bay

using purge and trap sampling with a tin-selective detector.

Environ. Sci. Technol. 16, 110-119 (1982).

30. Pearse, R. W. B. , Gaydon, A. G. The Identification of Molecular

Spectra, Chapman and Hall, London, pp. 327-8 (1976).

31. Calingaert, G. , Soroos, H., Shapiro, H. The redistr ibut ion reaction

VII alkyl compounds of mercury, tin, silicon and zinc. J. Am. Chem.

Soc. 62, 1107-1110 (1940).

18
32. Constable, D. J. C., Smith, R. S. , Tanaka, J. Comparison of solvent

reduction methods for concentration of polycyclic aromatic

hydrocarbon solutions. Environ. Sci. Technol. j_8» 975-978 (1984).

33. Parris, G. E. , Blair, W. R., Brinckman, F. E. Chemical and physical

considerations in the use of atomic absorption detectors coupled with

gas chromatographic systems for speciation of trace organometals

.

Anal. Chem. 378-386 (1 977).

34 . Lion, L. W. , Leckie, J. 0. The biogeochemistry of the air-sea

interface. Ann. Rev. Earth Plan. Sci. 9_, 449-486 (1 981 ).

35 . Guard, H. E. , Cobet, A. B., Coleman, W. M. III. Methylation of

trimethyltin compounds by estuarine sediments. Science 21

3

i 770-771

( 1981 ).

19
Table I. Reproducibility of Butyltins Analysis^

n BuSn Bu2Sn Bu^Sn BU||Sn

Replicate injections

of one sample extract^

7 12$ 5 % 12$ 9$

Replicate extractions

of same sample^

6 9 15 1

1

1

1

^Standard deviation

2 Sample of Chesapeake Bay water (site 858) spiked with 1.0 ug/L Bu SnCl^^-

(n = 1-4)

20
Table II. Concentration of butyltin species (ng/L Sn) in natural waters

Site BuSn^^ Bu-pSn^"^ Bu^Sn"^ BUijSn

Jones Falls Nd3 8 trace ND

p Jones Falls microlayer 1 65 91 1 872 395

Annapolis marina-l ND 10 14 ND

2 Annapolis marina microlayer ND 150 trace ND

Annapolis marina-2 ND 21 29 ND

Shelter Island ND 109 68 ND

San Diego Bay^ ND 4 2 ND

Dry Dock Sample ND ND 900 4200

^Collected at 1 m depth unless otherwise noted, 100 mL sample extracted

2 Concentration in wash water (approximtely 50 mL) collected from Teflon

sheet (0.32m^)

^Not detected

^Sample volume = 800 mL

21

EXTRACT/NO HYDRIDE

EXTRACT/HYDRIDE

HYDRIDE/EXTRACT

SIMULTANEOUS

Bu4Sn

8 12 MIN

Effect of extraction and hydridization sequence on detection of

butyltin species spiked (1.0 ug/L final aqueous concentration for

Bu2« 4 SnCl 4»fj, 4yg/L for BuSnClO into Chesapeake Bay water (site

85oc). "Extract/hydride” denotes that the extraction step was performed first followed by hydridization of the extract.
NBS RESEARCH MATERIAL

2, Analysis of chromatographically prepared trlbutyltin research

material. The bottom chromatogram shows the material spiked with

Bu^SnH 4»j^ at 3% (n-1-3) to 12$ (n-1 ) of the concentration of the

trlbutyltin level.
l-fel,

t-’oaK

12

DL«5

A

System GC-FPD

r2 ng Sn/L ng Sn

BuSnHs 0.940 22 0.18

BujSnH: 0.950 3 0.11

BujSnH 0.969 7 0.08

Bu4$n 0.985 7 0.28

200

( 400)

Sn, ng/L

3 . Calibration curves for Bu,_^Sn species spiked in Chesapeake Bay (site 701) water. Here 100 mL water was extracted and hydridized and the dichloromethane extract concentrated to 50 uL. Data points reoresent

means of duplicate 5 uL injections except for the lowest

concentrations which are means of 5 injections. In all cases

I"ter:;a! oonncenoe
PEAK

AREA

PEAK

AREA

4. Effect of pH (top) and salinity (bottom) on detection of butyltin

species spiked (1 yg/L) into Chesapeake Bay waters. Water from site

858c was used in pH experiments. Waters of differing salinities in

Chesapeake Bay were collected and analyzed in the salinity

experiments. Zero salinity denotes deionized water. Error bars

denote ± one standard deviation.
5. Butyltln species detected in samples form Baltimore Harbor (Jones

Falls) and San Diego Bay (Shelter Is.). The spikes in the Shelter

Is. chromatogram represent 50 ng/L Bu2 _ 4SnCl 4 _^ and 210 ng/L BuSnCl:,.

Dipropyltindichloride (internal standard) was added at 50 ng/L.

^
BLANK

I

1 1 r

0 4 8 12 MIN

6. Chromatograms of Baltimore Harbor microlayer samples diluted 1:10 and

1 :2 with deionized water prior to extraction.
Authentic Tetrabutyltin

Ship Wash Water Sample

7 . Maas spectrum of authentic tetrabutyltin (top) and tetrabutyltin

found in dry dock sample (bottom) containing tributyltin antifoul inn paint. Peak at m/e - 205 is large non-butyltin containing peak which is not fully resolved chromatographically (see Fig. 8).
Peak

Intensity

Selected Ion GC-MS

Wash Water Sample

Retention Time (min.)

8. Selected ion chromatogram demonstrating large unresolved peak with

m/e - 205 and retention time of - 6.8 min, with peak tailing into the

7.8 min tetrabutyltin peak(M/e - 235).
COLGATE CK

Bu2SnH2

BusSnH

0 8 12 MIN

Degradation of tributyltin spiked (0.1 aig/L, final concentration)

into Baltimore Harbor (Colgate CK) surface waters. Four 250 mL

bottles received 45 mL Colgate CK water and 5 mL of aqueous

tributyltin research material (1.0 mg/L) . Two of the bottles were sterilized (121® C, 15 min) prior to addition of tributyltin. One

sterile and one non sterile bottle received 1.0 mL sterile 25X yeast

extract (0.05? final concentration). After three months incubation

in the dark, the bottles were analyzed for butyltln concentrations.

APPENDIX A

PROTOCOL FOR THE ULTRA-TRACE SPECIATION OF BUTYLTINS IN SEAWATER BY GC-FPD

USING SIMULTANEOUS HYDRIDIZATION/EXTRACTION WITH THE TRACOR MODEL 570 DUAL

GC-FPD SYSTEM^

Equipment and Supplies

Gas chromatograph

with FPD detector and

Spectra-Physics integrators

Gases

nitrogen-zero grade for carrier

air-breathing quality

oxygen- tech, grade

hydrogen-tech, grade

Regulators for above gases, including

a regulator with a stainless steel

diaphragm for use with high purity

nitrogen such as the Matheson

3800 series

Glass columns to fit above'

6ft X 1/4 in O.D.

2mm I.D.

Optical interference filters

600-2000 nm cut on 610 nm (lOnm band pass)

Column packing: 1.5? 0V-1001 on Chromosorb G HP (100/120),

3 % OV-101 on Chromosorb G HP

Glass Wool (silane treated)

Examples of Suppliers and Notes

Tracor Instruments

6500 Tracor Lane

Bldg 27

Austin, TX

512-929-2051

Matheson

6655 Amberton Drive-unit 0

Baltimore, MD 21229

301-796-0517

Tracor or Supelco

Supelco Park

Bellefonte, PA 16823

Ditric Optical

Hudson, MA

One set of these will be provided

GC instrument, you may wish to

purchase spares

Varian Instrument Co.

650 E. Areqes Drive

Sunnyvale, CA 94084

Available from Supelco on Special

Order

Supelco

Certain commercial products or equipment are mentioned in order to

adequately describe experimental procedures. In no case does such identification imply endorsement by the National Bureau of Standards, nor does

it imply that the material is necessarily the best available for the

purpose.

1

APPENDIX A

PROTOCOL FOR THE ULTRA-TRACE SPECIATION OF BUTYLTINS IN SEAWATER BY GC-FPD

USING SIMULTANEOUS HYDRIDIZATION/EXTRACTION WITH THE TRACOR MODEL 570 DUAL

GC-FPD SYSTEM^

Equipment and Supplies

Gas chromatograph

with FPD detector and

Spectra-Physics Integrators

Gases

nitrogen-zero grade for carrier

air-breathing quality

oxygen- tech, grade

hydrogen- tech, grade

Regulators for above gases, including

a regulator with a stainless steel

diaphragm for use with high purity

nitrogen such as the Matheson

3800 series

Glass columns to fit above'

6ft X 1/4 in O.D.

2mm I.D.

Optical interference filters

600-2000 nm cut on

610 nm (1 Onm band pass)

Column packing: 1.555 OV-1001 on Chromosorb G HP (100/120),

355 OV-101 on Chromosorb G HP

Glass Wool (silane treated)

Examples of Suppliers and Notes

Tracer Instruments

6500 Tracer Lane

Bldg 27

Austin, TX

512-929-2051

Matheson

6655 Amberton Drive-unit 0

Baltimore, MD 21229

301-796-0517

Tracer or Supelco

Supelco Park

Bellefonte, PA I 6823

Ditric Optical

Hudson , MA

One set of these will be provided

GC instrument, you may wish to

purchase spares

Varian Instrument Co.

650 E. Areqes Drive

Sunnyvale, CA 94084

Available from Supelco on Special

Order

Supelco

^Certain commercial products or equipment are mentioned in order to

adequately describe experimental procedures. In no case does such identification imply endorsement by the National Bureau of Standards, nor does

it imply that the material is necessarily the best available for the

purpose.

1
GC septa Tracer or Supelco

Syringes for manual injection Precision Sampling Corp.

10 microliter size Available through Supelco

D ichlororaethane

spectro-grade

Sodium borohydride 9855 pellets (ea. pellet approx. 0.4g)

Organotin compounds for use in

quantitation:

monobutyltin chloride

dibutyltin dichloride

tributyltin chloride

tetrabutyltin

dipropyltin dichloride

Spectro-grade methanol for making

stock solutions

Separatory funnels with Teflon

screw-cap and Teflon stopcock

(250 ml size)

Pasteur pipets and bulb

Micropipets

Volumetric flasks, 50, 25, 1 0 ml

Bench top centrifuge

Tubes to fit above

(polypropylene)

Burrell wrist-action shaker

Sample evaporator

(Be sure air used for this

is free of oil and water)

Nitric Acid

Such as Omnisolve, available

through suppliers such as Fisher

Aldrich Chemical Co.

Milwaukee, WI 53201

Alfa Products

925 Andover St.

Danvers, MA 01923

Organometallics, Inc.

Route 1

1

East Hampstead, NH 03826

Omini-solve or similar

Wheaton Scientific

Special Order

ATTN: Maryalice Robinson

Probably available in your

Chem. Stock room

Such as Eppendorf, Pipetman,

Lange-Levy, etc. depending on the

preference of your chemists

Standard item in Chem. Stores

Eppendorf type

Cole Parmer

7425 N. Oak Park Dr.

Chicago, IL 60648

Burrell Corp.

Pittsburgh, PA

We use home-made device, but for

large sample through-put, you

might consider a commercial evaporator-concentrator such as in

Supelco 1985 catalog, p 179

Should be available in your

Chemistry storeroom
Laboratory detergent and brushes Should be available in your

Chemistry storeroom

Deionized water/Articf ical seawater For use in blanks and calibration

curves

Ring Stands and Clamps

Kimwipes, or similar wipers

Glass bottles with Teflon-lined

caps, for storage of stock

solutions (50 to 100 ml)

Aluminum foil
Analytical Procedure for the Tracer model 570

Gas Chromatograph-Flame Photometric System

CAUTION!

!

GC-FPD unit should be equipped with 600-2000 nm cut on filter for tin

detection in RIGHT detector (front photomultiplier) and 39^ nm filter for

sulfur detection in LEFT detector (rear photomultiplier). Be sure to

unplug power supply wires on both PM tubes or turn off main power to GC

before removing PM tube. to check or change filters or the PM tubes will be

permanently damaged. There is no separate power switch for the photomultiplier tubes on the Tracer G. C.

GC-FPD set-up and check

1 . Turn on nitrogen carrier gas and adjust regulator to deliver

50 psi.

2. Nitrogen flow rate = 20mL/min. This is approximately 17

divisions on the built-in flow meter on GC. Check with bubble flow meter

attached to flame exhaust tube before turning on flame gases. The

nitrogen flow is controlled by adjusting a knob labeled "SPLITTER GAS" on

the left side of the detector, under the flip-up cover on the top of the

G. C.

3. Turn on main power to GC-FPD (if unit is not on stand-by) and set

detector to 200°C and injection port to 150°C. Wait until detector is up

to operating temperature before trying to light flame.

1 v
4. Turn on hydrogen and air tanka, adjust regulators to 50 psi. For

easy lighting of flame, set air to 150 mL/min and hydrogen to 100 mL/min.

This ratio may require adjustment during flame lighting in order to ignite

detector. Flame is ignited by turning hydrogen flow OFF with knob on

detector, then using toggle switch to heat ignitor. While holding ignitor

switch ON, turn hydrogen flow ON with knob. Flame ignition can be

detected by holding a cold metal object (viz., a small wrench) to the

outlet of the exhaust tube and observing condensation. The flame will

probably not light the first time, and slight adjustment in gas ratios may

facilitate flame lighting. Repeated inability to light flame could be due

to deposits in exhaust tube. (See trouble shooting guide.)

5. For operation, set hydrogen at 170 mL/min and air to 75 mL/min on

the flow controllers.

6. Check that external attenuation = 100 (on GO front panel) and

internal attenuation = 1 (set with Spectra physics integrator).

7. Make sure that the oven fan in ON, and adjust CRT screen

brightness.

8. Call up desired temperature program from microprocessor memory.

When oven reaches required temperature the unit is ready to run.

9. The temperature program used for the 3? OV-101 column is -

(See Fig. 1

)

Initial over temperature: 40®C

Following injection hold 40°C for 2 min

then increase to 170°C at 40°C/min

hold for 1 min then to 180°C at 40°C/min

hold for 2 min then to 200°C at 40°C/min and hold for 3.5

min

V
Under these conditions the retention times are approximately;

Compound

Monobutyltin

Dipropyltin

Dibutyltin

Tributyltin

Tetrabutyltin

Note; This is control

memory

.

The temperature program use with the ^ . 5 % column is (See Fig. 2)

Initial temperature; 40®C

After injection hold 40®C for 2 min

then increase to 170®C at 40°C/min, hold for 2.5 min

then go to 190°C at 40°C/min

Under these conditions the retention times are approx:

R.T. (min)

2.90 ± 0.05

4.78 ± 0.05

5.91 ± 0.07

8.46 ± 0.1

11.87 ± 0.2

method 06 in G. C. microprocessor

R.T. (rain)

2.28 ± 0.05

4.34 ± 0.05

5.48 ± 0.07

7.71 ± 0.1

10.78 ± 0.2

Note; This is Control Method 02. The even number

control methods control the RIGHT detector and the odd

Compound

Monobutyltin

Dipropyltin

Dibutyltin

Tributyltin

Tetrabutyltin

VI
number methods control the LEFT. In its present

configuration, this particular instrument has only a

right detector, no left detector is installed.

Column temperatures over 200 ®C are not advised because the

butyltin hydrides tend to decompose above that temperature.

The maximum temperature for the column packings is 350°C.

The retention times noted are approximate and will vary

slightly from column to column due to differences in

tightness of column packing, etc. These retention times

should be checked using known compounds each time a new

column is installed and periodically over the life of a

column.

VI

1
Packing and Conditioning Columns

The analytical columns can easily be packed by plugging the column

outlet with glass wool, applying a slight vacuum and using a Pasteur pipet

bulb with the top cut off as a funnel to transfer the packing material to

the column. Gentle tapping of the column with a pencil or similar object

will insure uniform packing. When the column is filled to within about 1

inch of the inlet, remove the vacuum and plug the inlet with glass wool.

The column is now ready to be conditioned. This is accomplished by

installing the inlet end of the column into the injection port of the GC

with the carrier gas flowing (do not connect the column outlet to the

detector) and running a slow temperature gradient as recommended hy the

packing manufacturer. For the OV packing, the recommended procedure is to

purge the column at ambient temperature for at least 30 min with the

carrier gas flowing, then program the oven to heat from 50°C to the

highest anticipated operating temperature of the column at 2°C/min, then

hold this temperature overnight. The new column is then ready for use.

The packing material seems to have some active sites which bind the

organotin compounds irreversibly. To passivate these sites on a newly

packed column, make several injections (3“^ is usually sufficient) of

organotin analyte prior to the first injection used for quantitation.

These procedures are necessary only for newly packed columns. After

extensive use with environmental samples, the inlet glass wool and first

several centimeters of the column packing material may become discolored

(usually green) and inefficient. It is possible to gently remove the

discolored portion, then repack and replug with glass wool. This repaired

V i i i
column should be conditioned and passivated as a new column. Eventually

the packing material loses efficiency after extensive use (>200

injections). This is usually manifested in exaggerated peak tailing and

poor peak shape. The exhausted material can be blown out and the glass

column repacked.

For best resolution of monobutyltin trihydride analyte from the

solvent peak, use the 3$ OV-101. For a slightly faster G. C. run, but

poorer monobutyltin peak, the 1.5$ is satisfactory. (See Figs. 1 and 2).

The reduced loading packing (1.5$) has the advantage of less column bleed

and possibly fewer problems with heavy build up of material in detector

exhaust tube.

IX
Hydridization/Extraction Procedure

Reagent Preparation

1. Make stock solutions of mono-, di“, and tri-n*-butyltin chloride,

tetrabutyltin and di-n-propyltin dichloride in Spectro grade methanol at

concentrations of approximately 1000 ppm Sn in 50 mL volumetric flasks.

Transfer to acid washed borosilicate glass bottles and cover bottles with

aluminum foil. Maintained in the dark and tightly capped, these standards

have a shelf life of 3 months or more.

2. The working standards are prepared from the above methanol stock

solutions by dilution of 1:1000 in deionized water, to give a working

solution of approx. 1 ppm. For example, dilution of 50 microliters of

1000 ppm methanol stock into 50 mL of water yields a working solution of

1 ppm Sn. For working a the 1 ppb level this working solution can be used

directly (0.100 mL of this dilution into a 100 mL sample gives final Sn

concentration of 1 ppb). For work at the partS“per~trillion level, a

second dilution of 1:10 is advisable. Diluting 1 ppm solution 1.0 mL to

10 mL gives a working solution of 100 ppb. A 0.050 mL spike into a 200 mL

sample gives a final concentration of 25 parts-per-tr illion Sn.

Note: Prepare the aqueous working solutions daily.

3. Prepare 4? sodium borohydride solution in deionized water. Each

pellet weighs approximately 0.4 gram, so one pellet dissolved in 10 ml

water is a 4^ solution. Prepare daily.

X
Analysis Sequence

The following procedure is for 200 mL sample size, using a 250 mL

Wheaton screw top separatory funnel. For larger or smaller sample volumes

the reagents are scaled up or down accordingly.

1 . Place water sample in separatory funnel. Add dipropyltin

dicloride solution spike to give final concentration in the range that you

expect to find in the particular sample being analyzed. For most environmental waters, a final concentration of 25 or 50 ng Sn/L for the internal

spike is satisfactory. The same spike is added to all sample and calibration runs, and peak areas are normalized to the internal standard. Shake

funnel to disperse spike.

2. Add 6.0 mL high purity dichloromethane and 2.0 mL sodium

borohydride solution. Cap and shake by hand for about 30 sec. Open cap

and VENT, as considerable pressure builds up quickly. Recap funnel and

shake on mechanical shaker for ten minutes. Venting during mechanical

shaking is not necessary.

3. Remove funnel from shaker, open cap, allow 5 min. for phase

separation with caps loosened. Recovery is aided by hitting the funnels

with the heel of the hand to recover the layer of dichloromethane that is

often found floating on the sample’s surface. Remove the lower (dichloromethane) layer into centrifuge tubes. In sample with considerable algal

concentration or other particulate matter there may be an emulsion layer

composed of particate matter, water and solvent. This can be easily

broken by centrifugation for about 1 min. The upper water/particulate

layer can then be pipetted off.

XI
4. Add 3-0 tnL dichloromethane to the sample, hand shake and vent as

before. Shake mechanically for 10 min.

5. While second extraction is being done, the first extract can be

evaporated under a gentle stream of dry air. DO NOT EVAPORATE TO DRYNESS.

6. After the 10 min. shake, and following 5 min. equilibration, the

second solvent extract is added to the centrifuge tubes, combining the

extracts. For 100 mL samples both extracts will fit into the 1.5 mL

centrifuge tube. For larger sample volume (250 mL) two tubes, or more,

are required. For 1 L samples, 15 mL glass centrifuge tubes have been

used successfully.

7. The combined dichloromethane extracts are then concentrated by

evaporation under flowing air. For water samples of 1 ppb tributyltin

(TBT) concentration, the solvent extract need only be evaporated to about

1 mL, For samples in the 10“50 parts-per-trillion range, evaporation to

50^100 microliters is required. The final volume of the solvent need not

be measured exactly, as the use of the internal standard compensates for

small variations in volume, mechanical losses during work up, etc.

8. A five microliter portion of the sample extract is then injected

into the gas chromatograph.

9. The syringe should be flushed with dichloromethane immediately

after use, to prevent deterioration and contamination of the syringe.

Sample carry over in the syringe can be a problem, especially if going

from samples of high to low TBT concentration. Injection of 5 microliters

of dichloromethane as a syringe blank at the beginning of the day is

useful to insure that there is no syringe carry-over from previous uses.

10. Appropriate blanks should be analyzed daily, including a

sampling blank (i.e., a distilled water or artificial seawater sample that
was handled and stored in the same way as the environmental sample), and

one or more reagent blanks (i.e., distilled water or artificial sea water

which has not been exposed to the sampling procedure, but is carried

through the entire analytical work up). These blanks are absolutely

necessary to insure that no source of butyltin contamination has been

introduced into the analytical environment through incompletely cleaned

glassware or other sources. When working at these ultra-trace

concentrations contamination of the samples is very troublesome and

frequent analysis of blanks is the only way to be sure that the measured

concentrations represent what is found in the sample, not what may have

been added accidentally during or after sampling.

11. Calibration curves are generally done at the end of the day, to

insure that there is no carry-over of the butyltins used for calibration

into sample analyzed subsequently. A 3 or 4 point calibration curve is

usually sufficient, using the internal standard as well as all butyltin

species seen in the sample. Calibration curves can be run in deionized

water, artificial sea water or clean sea water. The choice of matrix

depends on the type of sample and availability. The use of the internal

standard allows for the use of a calibration curve, rather than doing a

standard addition technique for each sample, but it is possible that there

are interferents (positive and negative) that exist at specific sampling

sites. For this reason it is recommended that comparison of results

obtained by standard addition and calibration curve methods be done for

each site to be monitored, to insure that site specific interferents are

xi i i

not a problem.
Maintenance and Trouble-shooting Guide for GC-FPD System

A white solid material composed of column bleed and combusted analyte

tends to accumulate in the detector. This material decreases detector

sensitivity and can cause severe problems, such as inability to light

flame. Several procedures should be followed to minimize these

difficulties.

1 . The analytical column should be periodically "baked” at elevated

temperature (170-200®C) with carrier gas flowing for several hours

(overnight) to elute any highly retained materials. During this baking,

the detector should be burning or the column outlet should be removed from

the detector, to prevent build-up of refractory materials on the detector.

2. The exhaust tube which exits the detector should be removed daily

and rinsed with water to remove any accumulated material which could

impair gas flow through column and/or detector. Heat tape applied to the

exhaust tube, although not tried at MBS, may prevent condensates.

3. The detector should be removed periodically and cleaned with

acetone as described in the operator's manual for the detector. The

interval between cleanings is difficult to predict as it depends on the

number and type of samples being analyzed. A monthly or bi-monthly

cleaning schedule will probably be sufficient.

4. The o-rings of the detector must be replaced periodically as they

become brittle and crack with extended use. The operator’s manual

recommends replacement at intervals of six months, but more frequent

changes are probably advisable as preventive maintenance.

XIV
5. The windows (heat shields) inside the detector can become cloudy

and impair sensitivity. These windows should be inspected each time the

o-rings are changed and replaced if they begin to look cloudy.

6. If there is a catastrophic loss of sensitivity, the first thing

to check is whether the flame gas mixture is correct because response is

highly dependent on flame conditions. Next check pneumatic problems, in

particular a blockage in the flame exhaust pipe. If this does not solve

the problem, remove the detector and flush with acetone as described in

FPD manual. If this does not work, there is probably a leak caused by a

bad o-ring or a clouded heat shield. Replace o-rings (and heat shield, if

necessary). If none of these measures are successful, it may be that the

photomultiplier tube is going bad, as PM tubes show diminished response

with aging. In our experience, PM tubes have lasted several years and

none of the problems we have experienced have been caused by the PM tube.

7. Unusual peaks are sometimes seen since this method is tin

selective, not tin specific. These may be S, As, Pb or other compounds

which emit in the observed wavelengths when burned in a hydrogen rich

flame. These peaks often occur in the 5-7 minute retention time range,

but do not generally interfere with butyltin peaks. By using simultaneous

Tandem detection with two integrator/plotters, it is possible to determine

if the peak is caused by tin or by another element. One detector is

equipped with a 600-2000 nm filter selective for Sn and the other is

equipped with, for example, a 394 nm filter which is sulfur selective.

Comparison of ratios of authentic Sn compound peak areas on the tin and

sulfur filters will reveal if the unknown peak is Sn containing.

Measurements were made comparing the Sn/S peak area ratios for two sulfur

XV
compounds. For dimethyldisulf ide, the ratio is 0.029 ± 0.008 and for

2-mercaptoethanol, the ratio is 0.018 ± 0.002. A FID detector is also

present on the Tracer instrument, if simultaneous FID-FPD is desired.

XVI
Cleaning Glassware

Butyltin compounds absorb onto glass and Teflon surfaces and rigorous

cleaning procedures are required to prevent cross-contamination from

sample to sample. This is especially critical when going from high to low

concentration samples. After use the funnels should be shaken with

methanol for about 5 minutes, then washed with hot soapy water and a

brush. After the glassware is rinsed, it is completely disassembled and

leached with warm nitric acid overnight, followed by copious deionized

water rinses.

xvi i
complete

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BIBLIOGRAPHIC DATA

SMEO fS»« InlxrvcVort/

l. PUBLICATION OR 2. Parformini Orfan. Rapon No. 1. Publication Dau

REPORT NO.

NBSIR-85/3295

4. title and subtitle

A Comprehensive Method for the Determination of Aquatic Butyl tin Species at

Ultratrace Levels Using Simultaneous Hydridization/Extraction With GC-FPD

l- AUTMOR(S)

G.J. Olson, F.E. Brinckman, C. Matthias, O.M. Bell ama

i. PERPOBMING organization (tf Joint er eth0f then N BS, toe intuvctionM)

matiohal burbau op standards

DEPARTlieHT OP COMMERCE

VASHINCTOR, O.C. 2t2S4

7. Contract/Grant No.

I. Typa of Raport A Fanod Covarao

I. SPONSORING organization NAME AND COMPLETE ADDRESS (Street. City, Siote, ZIP)

16. SUPPLEMENTARY NOTES

( I Dooimant deecribas a computar program; SF-IBS, FIPS Software Summary, it attached.

11. ABSTRACT (A 200-word or loss foctval MurAnery of most significant information. If document Includoa a tignificoct

BIbllogrephy or lltarotvre ourroy, mention It horm)

A Bcthod for tht anAlyslB of aqaatlc butyltln axid ailxed aethylbotyltlxi Mpecles

na<Ti£ MlnultmneouB faydrldlxatlra vlth sodium borofaydrlde axid extraction Into

dlchloromethane Is described. The detection limits are 7 ng Sn/L for tetrabutyltln, 7 ng Sn/L for trl-n-butyltln, 3 ng Sn/L for dl-n-butyltln, and 16 ng

Sn/L for mono-n-butyltln. The presence of tetrabutyltln

In harbor vaters Is reported.

12- KEY WORDS (Six to twelve entries; alphabetical order; capitalize only proper names; and separate key words py semicolon g, alkyltln compoxznds; chemical speclatlon; Chesapeake Bay; clement selective detectors;

environment; extraction; gas chromatography; organometallic compounds; organotin bloclide!

13. AVAILABILITY

[Xl Unlimited

1 I For Official Distribution. Do Not Release sd NTIS

I I Order From Superintendent of Documents, U.S. Government Printinf Office, Washtnfton. D.C.

2D40Z

1 I Order From National Technical Information Service (NTIS), Sprinxf'eld. VA. 22161

M. NO. OF

PRINTED PAGES

50

IS. Pi

59. 9L

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