 |
GLYPHOSATE
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| Method Number: |
PV2067 |
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| Control Number: |
T-PV2067-01-8911-CH |
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| Matrix: |
Air |
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| Target Concentration: |
1 mg/m3 (arbitrary level). There is no OSHA permissible
exposure limit (PEL)
or ACGIH threshold limit value (TLV) for glyphosate. |
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| Procedure: |
Samples are collected by drawing known volumes of air through glass
fiber
filters. Samples are desorbed with 0.025 M borate buffer, derivatized and
analyzed by high performance liquid chromatography (HPLC) using an
ultraviolet detector (UV). |
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Recommended air volume and sampling rate:
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100 L at 1.0 L/min |
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Detection limit of the overall procedure (based on the recommended air
volume):
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1 µg/m3 |
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| Status of method: |
Stopgap method. This method has been only partially evaluated and is
presented for information and trial use. |
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| Date: November, 1989 |
Chemist: Duane Lee
|
Carcinogen And Pesticide Branch
OSHA Analytical Laboratory
Salt Lake City, Utah
1. General Discussion
1.1 Background
1.1.1 History of procedure
The OSHA Analytical Laboratory received samples on glass fiber filters
and OVS-2 tubes
requesting the analysis of Roundup which is the isopropylamine salt of
glyphosate. A
NIOSH procedure was tried but it did not yield a satisfactory separation.
(Ref. 5.1) From
a literature search there were procedures for the analysis of glyphosate in
soil and water
samples. (Refs. 5.2 to 5.4) These procedures were modified for the
analysis of air samples.
This report describes the preliminary validation of a sampling and
analytical method using
glass fiber filters. The OVS-2 tubes were examined but felt to be
unnecessary since
glyphosate is a solid with a melting point over 200ºC.
1.1.2 Toxic effects (This section is for information only and should
not be taken as the basis of
OSHA policy.)
The acute oral LD50 for rats is 4300 mg/kg for
glyphosate. (Ref. 5.6)
1.1.3 Potential workplace exposure
Glyphosate is used as a non-selective, postemergence herbicide.
(Ref. 5.6) No information
could be found on the number of workers exposed to glyphosate.
1.1.4. Physical properties (Refs. 5.5 to 5.7)
| Molecular weight: |
169.07 |
| Molecular formula: |
C3H8NO5P |
| CAS #: |
1071-83-6 |
| Melting point: |
230ºC (decomposition) |
| Solubility: |
soluble in water at 25ºC 12 g/L,
insoluble in most organic solvents |
| Chemical name: |
glycine, N-(phosphonomethyl)- |
| Other names: |
Mon 0573; N-(phosphonomethyl)glycine;
phosphonomethyliminoacetic
acid |
| Description: |
white solid |
| Structure: |
 |
| UV scan: |
 |
1.2 Limit defining parameters
The detection limit of the analytical procedure
is 0.84 ng per injection. This is the amount of
analyte which will give a peak whose height is
approximately five times the baseline noise.
2. Sampling Procedure
2.1 Apparatus
2.1.1 A personal sampling pump that can be calibrated to within ± 5%
of the recommended flow
rate with the sampling device in line.
2.1.2 Gelman type A/E 37-mm glass fiber filters. The filters were
assembled in two-piece 37-mm
polystyrene cassettes with backup pads. The cassettes are sealed with
shrink bands and
the ends are plugged with plastic plugs.
2.2 Reagents
No sampling reagents are required.
2.3 Sampling technique
2.3.1 Immediately before sampling, remove the plastic plugs from the
filter cassettes.
2.3.2 Attach the cassette to the sampling pump with flexible
tubing.
2.3.3 Attach the cassette vertically in the employee's breathing zone
in such a manner that it
does not impede work performance.
2.3.4 After sampling for the appropriate time, remove the cassette and
seal with plastic plugs.
2.3.5 Wrap each sample end-to-end with an OSHA seal (Form 21).
2.3.6 Submit at least one blank for each set of samples. Handle the
blank in the same manner
as the samples, except no air is drawn through it.
2.3.7 Record the air volume (in liters of air) for each sample, and
list any possible interferences.
2.3.8 Submit bulk samples for analysis in a separate container.
2.4 Extraction efficiency
Six treated glass fiber filters were each liquid spiked with 20 µL of a
5.22 mg/mL glyphosate
standard. These were allowed to dry and placed in a drawer overnight. The
next day each filter was
extracted with 3.0 mL of 0.025 M borate buffer, shaken for 30 min and then
analyzed as per section
3.5. The results are listed in the table below.
Table 2.5
Retention Efficiency |
amount
spiked, µg |
amount
found, µg |
%
recovered |
104.4
104.4
104.4
104.4
104.4
104.4 |
100.44
103.46
105.75
109.59
140.94
103.1
average |
96.2
99.1
101.3
105.0
100.5
98.8
100.2 |
2.5 Retention efficiency
Six glass fiber filters were liquid spiked with 20 µL of a 5.22 mg/mL
standard and humid air (80%
relative humidity) was drawn through each filter at 1 L/min for 100 minutes.
The filters were extracted
with 3 mL of 0.025 M borate buffer, shaken for 30 min and then analyzed as
per section 3.5. The
results are listed in the table below.
Table 2.4
Extraction Efficiency |
amount
spiked, µg |
amount
found, µg |
%
recovered |
104.4
104.4
104.4
104.4
104.4
104.4 |
103.91
87.77
107.5
99.68
107.39
103.34
average |
99.5
84.1
103.0
95.5
102.9
99.0
97.3 |
2.6 Sample storage
Twelve glass fiber filters were liquid spiked with 20 µL of a 5.22
mg/mL standard and humid air (80%
relative humidity) was drawn through each filter at 1 L/min for 100 minutes.
Six of the samples were
stored at ambient temperature in a drawer, and six were stored in a freezer.
After four days of
storage, three samples from each group were extracted with 3 mL of 0.025 M
borate buffer, shaken
for 30 min and then analyzed as per section 3.5. The remaining samples were
desorbed and
analyzed after six days of storage. The results are given in the tables
below.
Table 2.6.1
Ambient Storage |
days
stored |
amount
spiked, µg |
amount
found, µg |
%
recovered |
4
4
4
6
6
6 |
104.4
104.4
104.4
104.4
104.4
104.4 |
97.49
100.91
100.38
94.18
94.71
95.67
average of 4
average of 6 |
93.4
96.7
96.1
90.2
90.7
91.6
99.2
91.6 |
Table 2.6.2
Freezer Storage |
days
stored |
amount
spiked, µg |
amount
found, µg |
%
recovered |
4
4
4
6
6
6 |
104.4
104.4
104.4
104.4
104.4
104.4 |
103.16
102.89
104.65
96.87
92.40
97.76
average of 4
average of 6
|
98.8
98.6
100.2
92.8
88.5
93.6
99.2
91.6 |
2.7 Recommended air volume and sampling rate
2.7.1 The recommended air volume is 100 L.
2.7.2 The recommended flow rate is 1.0 L/min.
2.8 Interferences (sampling)
It is not known if any compounds will interfere with the collection of
glyphosate.
2.9 Safety precautions (sampling)
2.9.1 Attach the sampling equipment in such a manner that it will not
interfere with work
performance or employee safety.
2.9.2 Follow all safety practices that apply to the work area being
sampled.
3. Analytical Procedure
3.1 Apparatus
3.1.1 A balance capable of weighing to the nearest tenth of a
milligram. A Mettler HL52 balance
was used in this evaluation.
3.1.2 Mechanical shaker.
3.1.3 A high performance liquid chromatograph (HPLC) equipped with an
ultraviolet (UV) detector.
A Hewlett-Packard (HP) 1090M with a diode array detector was used in this
evaluation.
3.1.4 An HPLC column capable of separating glyphosate from any
interferences. A 25 cm × 4.6
mm i.d. Zorbax NH2 column was used in this evaluation.
3.1.5 An electronic integrator, or some other suitable method for
measuring detector response.
The Hewlett-Packard 3357 Laboratory Data System and the Hewlett-Packard
1090M system
were used in this evaluation.
3.1.6 Volumetric flasks and pipets.
3.1.7 Vials, 4-mL with Teflon-lined caps.
3.1.8 Vials, 2-mL suitable for use on HPLC autosamplers.
3.2 Reagents
3.2.1 Acetonitrile, HPLC grade from Burdick and Jackson.
3.2.2 Glyphosate, Environmental Protection Agency (EPA #3801, 97.3%
purity).
3.2.3 Borate, sodium borate
(Na2B4O7·10H2O) from
Mallinckrodt. The borate buffer was 0.025 M
sodium borate with a pH = 9.
3.2.4 HPLC grade water, Milli-Q filtered water, Millipore Inc.
3.2.5 Acetone, high purity solvent from Burdick and Jackson.
3.2.6 9-Fluorenylmethyl chloroformate (FMOCCL), reagent grade obtained
from Aldrich. This was
made 0.002 M in acetone and used as the derivatizing reagent.
3.2.7 Potassium hydroxide, reagent grade from Baker. This was 7 N in
water and used to adjust
the pH of the mobile phase.
3.2.8 Potassium phosphate monobasic (KH2PO4),
reagent grade from Mallinckrodt.
3.3 Standard preparation
Prepare stock glyphosate standards by weighing 10 to 15 mg of
glyphosate. Transfer the glyphosate
to separate 10-mL volumetric flasks, and add borate buffer to the mark.
Make working range
standards of 0.03 to 80 µg/mL by pipet dilutions of the stock standards with
borate buffer. This range
corresponds to 0.09 to 240 µg per sample when an extraction volume of 3 mL
is used. Store stock
and dilute standards in a freezer.
3.4 Sample preparation
3.4.1 Transfer the glass fiber filter of each cassette to a 4-mL vial.
3.4.2 Add 3.0 mL of borate buffer to each vial and seal with a
Teflon-lined cap.
3.4.3 Shake the vials for 30 minutes on a mechanical shaker.
3.5 Derivatization of samples and standards
3.5.1 Transfer 1 mL of each sample and standard to 4-mL vials.
3.5.2 Add 1.0 mL of 0.002 M FMOCCL to each vial.
3.5.3 Cap the vials and then shake them for 10 to 15 seconds to ensure
mixing and allow them
to sit at room temperature for 30 min.
3.5.4 Transfer, if necessary, a portion of each sample and standard to
separate 2-mL vials for the
HP autosampler.
3.6 Analysis
3.6.1 Instrument conditions
| Column: |
25 cm × 4.6 mm i.d. Zorbax NH2 |
| Mobile phase: |
50% acetonitrile 50% water 0.05 M
KH2PO4 pH adjusted to 6.0 with
7 N KOH |
| Flow rate: |
1.0 mL/min |
| Column temperature: |
40ºC |
| Injection volume: |
25.0 µL |
| Retention time: |
9.6 min |
| Detectors: |
UV 206 nm |
| Fluorescence excitation = |
206 nm |
|
emission =
|
320 nm filter |
3.6.2 Chromatogram
3.7 Interferences (analytical)
3.7.1 Any collected compound having a
similar retention time and responds to
a UV and a fluorescence detector is
an interference.
3.7.2 Any compound that reacts with
FMOCCL is an interference.
3.7.3 HPLC conditions may be varied to
circumvent an interference.
3.7.4 Retention time alone is not proof of chemical identity. Analysis
by an alternate HPLC
column, ratioing between fluorescence and UV detectors and confirmation by
mass
spectrometry are additional means of identification.
3.8 Calculations
3.8.1 Construct a calibration curve by
plotting detector response versus
concentration (µg/mL) of glyphosate.
3.8.2 Determine the µg/mL of glyphosate in
each sample and blank from the
calibration curve.
3.8.3 Blank correct the samples by
subtracting the µg/mL in the blank
from each sample.
3.8.4 Use the following formula to determine the air concentration.
3.9 Safety precautions (analytical)
3.9.1 Avoid skin contact and exposure to glyphosate in air.
3.9.2 Avoid skin contact with all solvents.
3.9.3 Wear safety glasses at all times.
4. Recommendation for Further Study
The method should be fully validated.
5. References
5.1 Mosely, C. L. and Anderson, K.; Hazards Evaluations and Technical
Assistance Branch, NIOSH,
U.S. Department of Health and Human Services; Cincinnati, OH; Report No.
HETA-83-341-1557.
5.2 Miles, C. J.; Moye, H. A. J. Agric. Food Chem.
1988, 36(3), 486-491.
5.3 Gauch, R.; Leuenberger, U.; Mueller, U. Z. Lebensm.- Unters.
-Forsch. 1989, 188(1), 36-38.
5.4 Miles, C. J.; Wallace, L. R.; Moye, H. A. J. Assoc. Off.
Anal. 1986, 69(3), 458-461.
5.5 Registry of Toxic Effects of Chemical Substances 1985-86
Edition;
DHHS(NIOSH) Publication No.
87-114, U.S. Department of Health and Human Services: Cincinnati, OH, 1987;
p 2551.
5.6 Farm Chemicals Handbook; Berg, Gordon L. Ed.; Meister:
Willoughby, OH, 1989; p C147.
5.7 Merck Index, 10th ed.; Windholz, Martha Ed.; Merck:
Rahway, NJ, 1983; p 648.
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