1.1 Background

1.1.1 History of procedure over the past several years the OSHA Laboratory has received requests to analyze samples for "trimethylbenzene". These have primarily been air samples collected on charcoal. Since several related aromatic compounds are routinely collected on charcoal, desorbed with carbon disulfide, and analyzed by gas chromatography, it was decided to try the same with all three trimethylbenzenes. The three isomers are 1,2,3-, 1,2,4-, and 1,3,5-trimethylbenzene.

1.1.2 Toxic Effects (This section is for information purposes and should not be taken as the basis for OSHA policy.) Both the 1,2,4- and the 1,3,5- isomers are (Ref 5.1) highly toxic by inhalation and by ingestion. Both are skin and eye irritants. Both isomers are central nervous system depressants and may cause respiratory disorders. The 1,2,4-isomer may also be narcotic. Other effects of exposure to these compounds include headache, tension, nervousness, inflammation and hemorrhaging of mucous membranes, convulsions and ultimately death. No specific information about the toxic effects of the 1,2,3-isomer was available.

1.1.3. Potential workplace exposure: (Ref. 5.2)

1,2,4-Trimethylbenzene is used as a chemical intermediate in the manufacture of trimellitic anhydride, pseudocumidine and various dyes and pharmaceuticals. 1,3,5-Trimethylbenzene is used as an intermediate in the production of anthraquinone vat dyes and ultraviolet oxidation stabilizers for plastics. 1,2,3-Trimethylbenzene is a precursor for a musk. 1,2,4-Trimethylbenzene is produced at about 22,500 tons/year in the United States. 'The other Trimethylbenzenes are produced at much lower rates.

1.1.4 Physical properties:  (Ref. 5.3)

Compound:	1,2,3 Trimethylbenzene	1,2,4 Trimethylbenzene	1,3,5 Trimethylbenzene
Molecular Weight:	120.186 (all isomers)
Density:	0.8944	0.8758	0.8652
Freezing Point:	-25.37°C	-43.8°C	-44.7°C
Boiling Point:	176.1°C	169.35°C	164.7°C
Odor:	Characteristic of aromatic hydrocarbons.
Color:	Each is a clear liquid.
Molecular Formula:	C8H12
Flash Point (Tag closed cup):	51°C	46°C	44°C
CAS Number:	526-73-8	96-63-6	108-67-8
IMIS Number:	T205	T306	T407
	(2505 for mixture of all three)
Structure:

1.2 Limit defining parameters

 1.2.1 The detection limit of the analytical procedure is 2 ng/injection for each isomer. This is the smallest amount of analyte which will produce a peak height 5 times the baseline noise.

1.2.2 The detection limit of the overall procedure is 1 µg/sample for each isomer, assuming a 1 mL sample volume, a 2 µL injection volume and nearly 100% retention and desorption efficiencies.

1.3 Advantages

1.3.1 The sampling procedure is convenient for both the industrial hygienist and the employee being sampled.

1.3.2 No reagents and only a small amount of apparatus are required for sampling.

1.3.3 Samples are convenient to ship.

1.3.4 The analytical method is reproducible, sensitive and does not require extensive handling of samples or standards.

1.3.5 It is possible to analyze at the same time other analytes which are collected and desorbed by the same procedure and which are separable from the trimethylbenzenes by gas chromatography, through the proper choice of column and GC parameters.

1.3.6 Samples can be reanalyzed if necessary.

1.4 Disadvantages

None were encountered during these studies.

2.1 Apparatus

2.1.1 A calibrated personal sampling pump, the flow of which can be determined within ±5% at the recommended flow.

2.1.2 Charcoal tubes: glass tube with both ends flame sealed, 7 cm long with a 6mm O.D., and a 4 mm I.D., containing 2 sections of 20/40 mesh charcoal separated by a 2-mm portion of urethane foam. The front section contains 100 mg of charcoal, the backup section contains 50 mg. A plug of silane treated glass wool is placed ahead of the front section and a 3-mm portion of urethane foam is placed behind the backup section

2.2 Sampling technique

2.2.1 The ends of the tube are opened immediately before sampling.

2.2.2 Connect the backup end of the tube to the sampling pump with flexible tubing.

2.2.3 Tubes should be placed in a vertical position to minimize channeling.

2.2.4 Air being sampled should not pass through any hose or tubing before entering the charcoal tube.

2.2.5 Place plastic caps on each end of the tube immediately after sampling. Wrap each sample lengthwise with OSHA Form-21.

2.2.6 With each batch of samples, submit at least one blank tube from the same lot used for samples. This tube should be subjected to exactly the same handling as the samples (break ends, seal, & transport) except that no air is drawn through it.

2.2.7 Send the samples (and corresponding paperwork) to the laboratory for analysis.

2.2.9. If any bulk samples are submitted for analysis they must be shipped in a separate container from the air samples and blanks.

2.3 Desorption and/or Extraction efficiency

Eighteen charcoal tubes and a blank were studied. Six tubes were injected with 2.8 µL each of a 1/1/1 mixture (v/v/v) of the isomers. The mixture was then diluted 9:1 (v/v) with carbon disulfide. Each of six tubes were injected with 7.0 µL of this dilution and six more tubes were injected with 1.4 µL. The tubes were refrigerated about 18 hours and then desorbed with 1 mL of 0.1% (v/v) n-hexylbenzene in carbon disulfide. The following results were obtained.

TABLE 1
	1,2,3-isomer 0.835 mg Spike	1,2,4-isomer 0.817 mg Spike	1,3,5-isomer 0.808 mg Spike
1.	92.340%	94.693%	96.797%
2.	95.651	98.666	101.239
3.	92.228	94.662	96.857
4.	95.944	98.477	100.566
5.	90.102	92.432	94.375
6.	90.986	93.591	95.868
Ave S.D.	92.875% 2.412	95.420% 2.579	97.617% 2.707
	1,2,3-ismoer 0.209 mg	1,2,4-isomer 0.204 mg	1,3,5-isomer 0.202 mg
1.	112.688%	117.076%	119.648%
2.	108.328	112.740	115.000
3.	108.436	110.928	113.664
4.	109.288	111.916	114.168
5.	114.040	116.872	119.832
6.	108.992	112.048	114.780
Ave S.D.	110.295% 2.441	113.597% 2.680	116.182% 2.796
	0.0417 mg	0.0409 mg	0.0404 mg
1.	111.440%	117.560%	116.160%
2.	102.680	110.400	106.900
3.	106.320	114.700	111.580
4.	107.060	111.300	113.420
5.	110.680	116.160	115.920
6.	112.360	119.760	119.860
Ave S.D.	108.423% 3.714	114.980% 3.619	113.973% 4.456

2.4 Retention efficiency

A retention efficiency study of six tubes, each injected with 2.8 µL of the 1/1/1 (v/v/v) isomer mixture, and a blank was then performed. Each tube was exposed to a 0.1 L/min flow of air of approximately 83% relative humidity at 23°C for 100 minutes, corresponding to an air volume of about 10 L. These data were obtained:

TABLE 2
	1,2,3-isomer 0.835 mg Spike	1,2,4-isomer 0.817 mg Spike	1,3,5-isomer 0.808 mg Spike
1.	91.460%	94.057%	96.082%
2.	92.463	94.843	96.980
3.	95.020	97.714	100.132
4.	92.840	95.257	97.404
5.	91.945	94.316	97.021
6	92.648	94.505	>96.544
Ave.	92.648%	95.115%	97.3605%

These averages were nearly identical to desorption efficiencies for the same loading of the corresponding isomers. Water vapor appears to have very little effect in dislodging trimethylbenzene from activated charcoal.

2.5. Storage

Two sets of six samples, each injected with 1.4 µL of the isomer mixture, and one blank, were prepared and stored under refrigeration for three days. One set was exposed to humid air as was done with the retention study samples while the other set was not. There was no loss on storage.

TABLE 3
No Air Drawn
	1,2,3-isomer 0.417 mg Spike	1,2,4-isomer 0.409 mg Spike	1,3,5-isomer 0.404 mg Spike
1.	90.762%	93.722%	95.996%
2.	91.360	94.332	96.504
3.	93.198	96.270	98.384
4.	91.878	95.216	97.476
5.	95.754	99.244	101.642
6.	90.672	92.150	95.668
Ave.	92.271%	95.156%	97.612
Humid Air Drawn
	1,2,3-isomer 0.417 mg	1,2,4-isomer 0.409 mg	1,3,5-isomer 0.404 mg
1.	91.726%	93.598	96.276
2.	92.750	96.595	99.209
3.	96.309	99.930	102.286
4.	99.050	102.286	105.060
5.	94.915	97.717	100.458
6.	93.965	96.911	99.544
Ave.	94.786%	97.896%	100.472%

2.6 Air volume and sampling rate studied

2.6.1 The air volume studied is 10 L.

2.6.2 The sampling rate studied is 0.1 liters per minute.

2.7 Suspected interferences should be listed on sample data sheets.

2.8 Safety precautions

2.8.1 Sampling equipment should be placed on an employee in a manner that does not interfere with work performance or safety.

2.8.2 Safety glasses should be worn at all times.

2.8.3 All safety procedures that apply to the workplace being sampled should be followed.

3.1 Apparatus

3.1.1 Gas chromatograph equipped with a flame ionization detector.

3.1.2 GC column capable of separating the solvent, an internal standard and three trimethylbenzenes from each other and from any interferences.

3.1.3 An electronic integrator or some other suitable means of measuring detector response.

3.1.4 Two-milliliter vials with Teflon-lines caps.

3.1.5.A syringe of 2 µL or other convenient size for sample injection.

3.1.6 A repipet for accurately dispensing a known volume of desorbing solution to all the samples. In this work a 1 mL Glenco dispenser was used.

3.1.7 A syringe of 10 µL or other convenient size to inject pure standards into volumetric flasks.

3.1.8 Volumetric flasks of 5 mL or other convenient size in which to prepare standards.

3.2 Reagents

3.2.1 Nitrogen, hydrogen and oxygen of GC grade purity.

3.2.2 Carbon disulfide, reagent grade.

3.2.3 An internal standard, which elutes after the analytes and their most likely contaminants. Reagent grade n-hexylbenzene seems to be the best choice.

3.2.4 Analytes, reagent grade. 1,2,4-and 1,3,4-trimethylbenzene are readily available at 99% purity each. However, there seems to be a problem in obtaining 1,2,3-trimethylbenzene above 90X purity. It appears to contain significant amounts of the 1,2,4-isomer as well as other aromatic compounds.

3.3 Standard preparation

3.3.1 Standards of the trimethylbenzenes are prepared by injecting a known volume of each isomer, usually 5 µL, into a volumetric flask, usually 5 mL, partly filled with the desorbing solution (carbon disulfide plus internal standard), then filling the flask to the mark and mixing the analytes with the desorbing solution by a few successive inversions of the flask.

3.3.3 By preparing standards of various concentrations and running them, a calibration curve may be prepared.

3.3.4 At least two initial standards which are theoretically equal and are within ±5% agreement must be prepared and run.

3.4 Sample preparation

3.4.1 The front and back sections of charcoal from each tube are transferred to separate vials.

3.4.2 To each vial is added 1mL of desorbant.

3.4.3 Each vial is sealed immediately and allowed to desorb for 30 minutes with occasional shaking.

3.5 Analysis

3.5.1 Gas chromatographic conditions

Injection size : 2.0 µL

Injector temperature: 200°C

Nitrogen flow rate: 25 mL min.

Detector temperature: 250°C

Hydrogen flow rate: 40 mL min.

Air : 300 mL/ min.

Detector: Flame ionization

Column: 101 × 1/811 FFAP/Chrom. WAW

Oven Program: 120°C/6 min, 5°C increase /min, 150°C/5 min

Attenuation: 7²

Instrument: Hewlett-Packard 5840

Elution times:

1,3,5-trimethylbenzene: 5.37 min.

1,2,4-trimethylbenzene: 6.30 min.

1,2,3-trimethylbenzene: 7.82 min.

n-hexylbenzene (ISTD): 12.70 min.

3.5.2 Chromatogram



3.5.3 Detector response is measured by electronic integration of peak areas or other suitable means.

3.5.4 An internal standard is used to compensate for small differences in injection sizes. The integrator is calibrated in ppm for a 10L air volume after correction for desorption efficiency.

3.5.5 Precision

A precision study, involving six injections each of standards at four, twice, once, one-half, and one-tenth of the target concentration was performed. 

TABLE 4
4X Target
	1,2,3-isomer 0.835 mg/mL	1,2,4-isomer 0.817 mg/mL	1,3,5-isomer 0.808 mg/mL
1.	101.743	100.917	100.548
2.	99.833	99.549	99.303
3.	99.833	99.334	99.092
4.	99.621	98.800	94.411
5.	98.127	97.906	97.605
6.	98.131	97.875	97.640
ave. S.D. C.V	99.513 1.336 1.343	99.064 1.146 1.157	98.766 1.124 1.138
2X Target
	1,2,3-isomer 0.417 mg/mL	1,2,4-isomer 0.409 mg/mL	1,3,5-isomer 0.404 mg/mL
1.	48.554	48.122	84.195
2.	48.772	48.892	48.728
3.	49.189	49.386	49.198
4.	48.952	48.638	48.766
5.	48.769	48.808	48.703
6.	48.905	49.046	48.962
ave. S.D. C.V.	48.845 0.221 0.454	48.815 0.424 0.868	48.759 0.334 0.684
1X Target
	1,2,3-isomer	1,2,4-isomer	1,3,5-isomer
1.	24.309	24.168	24.095
2.	24.356	24.269	24.179
3.	24.186	24.089	23.988
4.	24.169	24.113	24.016
5.	24.323	24.289	24.153
6.	24.280	24.217	24.052
ave. S.D. C.V.	24.270 0.076 0.314	24.191 0.082 0.338	24.080 0.076 0.315
0.1X Target
	1,2,3-isomer 0.0417 mg/mL	1,2,4-isomer 0.0409 mg/mL	1,3,5-isomer 0.0404mmg/mL
1.	3.298	3.407	3.257
2.	3.312	3.446	3.253
3.	3.293	3.422	3.247
4.	3.684	3.711	3.535
5.	3.679	3.867	3.604
6.	3.645	3.742	3.559
ave. S.D. C.V.	3.485 0.202 5.804	3.599 0.198 5.508	3.409 0.173 5.082

3.6 Interferences

3.6.1 Any compound which produces a peak which partially or completely overlaps a peak produced by an analyte or the internal standard is an analytical interference. Possible interferences should be listed on the sample data sheet. GC parameters should be adusted in whatever way necessary to eliminate any such interference.

3.6.2 Since the title compounds are most frequently derived from coal tar naphthas, other alkylated benzenes, styrenes and cycloalkylated benzenes which have similar boiling points and/or molecular formulae may be interferences. Many such compounds are theoretically possible, but relatively few are available in a high degree of purity.

3.6.3 Retention time data on a single column does not alone prove chemical identity. Samples over the target concentration must be confirmed by GC/Mass Spec or other conclusive means. 

3.7 Calculations

3.7.1 To calculate the ppm of analyte, based on a 10 L air volume. and a 1 mL desorption volume, in standards.
 

ppm =

1 µL/mL × 1 mL × p × 24.46 × 1000 L 10 L × 1 m³ × DE MW

24.46 = # liters/mole of any gas at 25°C and 760 mmHg.
MW =  molecular weight 
p = Density of analyte 
1 mL = Recommended desorption volume 
10 L = Assumed air volume 
DE = Desorption efficiency

3.7.2 Printed values for samples are calculated by:
 

actual ppm =

printout - printout blank (acutal air volume) / 10 L

3.8 Safety precautions

3.8.1 All solvents should be handled in a hood.

3.8.2 Skin contact with any solvent is to be avoided.

3.8.3 Safety glasses are to be worn at all times.

4.1 Further work should be done to eliminate interferences, since there are many possible interferences. Capillary columns could be investigated.

4.2 Additional work should be done with each isomer separately to determine whether any given isomer contains trace amounts of any other isomer. This can't be seen when all three are mixed.

5.1 Material Safety Data Sheets for 1,2,4- and 1,3,5-Trimethylbenzene from OCIS file.

5.2 Kirk-Othmer Encyclopedia of Chemical Technology (1982), exec.ed. Martin Grayson, vol. 18, pp. 881.5.

5.3 CRC Handbook of Chemistry and Physics, 53rd Edition, ed. by Robert C. Weast, pp. C-165-6.