Anal. Chem. 1986, 58,448-453
448
calculated rates, and the relative standard deviations are consistent with deviations previously noted in this laboratory for the standard PSD (1)and other passive dosimeters (3). A statistical analysis of the entire data set suggests that differences between individual devices may contribute about 6% to the total relative standard deviation. Standard deviations are largest for those chemicals having low retention volumes, as anticipated from consideration of Figure 3. It will be noted from the data in Table I that some points were rejected in the mean and s t a n d p d deviation calculations. These rejections were made for points more than two standard deviations from the mean. With nominal sampling rates of 2.5 cm3/min, the curves shown in Figure 3 suggest that these reduced-rate PSD’s should be usable for 24-h sampling of chemicals having retention volumes as low as 15 L/g and for 8-h sampling of chemicals having retention volumes as low as 5 L/g. This is a marked improvement over the unmodified PSD (nominal sampling rate = 80 cm3/min), which was limited to 2-4-h sampling for many VOC’s of interest to the air monitoring community.
ACKNOWLEDGMENT We thank G. W. Keigley for his assistance and helpful
discussions during the course of this work.
Registry No. Acrylonitrile, 107-13-1;1,l-dichloroethylene, 75-35-4; dichloromethane, 75-09-2; chloroform, 67-66-3; 1,2-dichloroethane, 107-06-2;l,l,l-trichloroethane, 71-55-6; benzene, 71-43-2; trichloroethylene, 79-01-6; trans-1,3-dichloropropene, 10061-02-6;toluene, 108-88-3;tetrachloroethylene,127-18-4.
LITERATURE CITED (1) Lewis, R. G.;Mullk, J. D.; Coutant, R. W.; Wooten, G. W.; McMillin, C. R. Anal. Chem. 1985, 5 7 , 214-219. (2) Coutant, R. W.; Lewis, R. G.; Mullk, J. D. Anal. Chem. 1985, 5 7 , 2 19-223. (3) Coutant, R. W.; Scott, D. R. €nv/ron. Sci. Techno/. 1982, 76, 410-413.
RECEIVED for review June 17,1985. Accepted October 1,1985. Although the research described in this article was funded wholly by the US. Environmental Protection Agency through Contract No. 68-02-3487, it has not been subjected to Agency review. Therefore, it does not necessarily reflect the reviews of the Agency, and no official endorsement should be inferred. Mention of trade names or commercial products does not constitute endorsement or recommendation for use.
Modified Soxhlet Procedure for the Quantification of Resin and Rubber Content of Guayule Elizabeth J. Nurthen and Barry V. McCleary* Biological and Chemical Research Institute, New South Wales Department of Agriculture, Rydalmere, 2116,Australia Peter L. Milthorpe and J. Wayne Whitworth’ New South Wales Department of Agriculture, Condobolin, 2877, Australia
The standard Soxhlet procedure has been modified to allow the rapld and rellabie analysis of resin and rubber in guayule. After removal of most of the resln and rubber from samples by extraction with acetone and then hexane, respectlvely, the samples are hamogenized In acetone and reextracted in hexane. The blending step reduces the overall time of extraction and enables the quantltatlve recovery of resins and rubber. The procedure Is not labor intensive, and 12 samples can be readHy handled by a single operator in a day. An analysls format employing a Tecator Soxtec Is also descrlbed, which reduces extraction tlmes further.
Guayule (Parthenium argentatum Gray) is one of a few plant species that contains substantial quantities of rubber (1). However, unlike Hevea brasiliensis where the rubber is contained in ducts, in guayule it is deposited in single thinwalled cells mainly in the outer layers of stems (2). Thus, rubber extraction requires either mechanical disintegration of plant material followed by solvent treatment or blending under conditions that induce the rubber to coagulate (3,4). Present address, New Mexico State University,Las Cruces, NM. 0003-2700/88/0358-0448$01.50/0
A number of procedures have been used to quantify the rubber (cis-polyisoprene) content of guayule plant material. Trichome and leaf morphology were utilized as an indicator of high rubber bearing plants by Mehta et al. (5), whereas Bauer (6)employed microscopic techniques. Procedures based on the use of carbon-13 nuclear magnetic resonance (13C NMR) ( 7 , 8 ) , proton magnetic resonance (lHNMR) (9),or infrared spectrophotometry (10) have been developed, but since sophisticated equipment and specialized technical expertise are required, these procedures are not likely to be routinely adopted in analytical laboratories or field stations. Rather, solvent extraction procedures would appear to be more appropriate. Many such procedures have been described in the literature, ranging from the time-consuming Soxhlet extraction method of Spence and Caldwell (11) (or its modifications (12))to more rapid procedures where samples are blended in various highly flammable and explosive organic solvents. In one such method the fresh plant material is milled in liquid nitrogen and then extracted with acetone and hexane by blending in a Virtis “45“ homogenizer (13). Other “rapid” procedures have involved the use of a Tekmar Tissumizer (14) or a Polytron homogenizer (Brinkman-Willemshomogenizer) (15,16)with dried and milled plant material. In these procedures, individual samples need to be homogenized four or more times in highly inflammable organic solvents at or near 0 1986 American Chemlcal Society
ANALYTICAL CHEMISTRY, VOL. 58, NO. 2, FEBRUARY 1986
room temperature. This practice and the routine transfer of large quantities of flammable solvents between homogenizer cups and centrifuge containers is potentially dangerous through the possibility of solvent explosions. Also, the noise generated during blending is annoying and, although individual samples can be processed rapidly, the methods are laborious. Therefore, we decided to critically evaluate the Soxhlet extraction procedure to increase sample throughput without sacrificing accuracy or reliability. In this paper we describe two solvent extraction procedures that allow accurate and relatively rapid quantification of resins ( I , 3) and rubber in guayule. One of these procedures uses conventional Soxhlet apparatus while the other employs a Tecator Soxtec. In both procedures, preliminary extraction of the plant material with acetone and then hexane precedes a short homogenization in acetone and further extraction in hexane. The prime considerations in the development of these procedures have been accuracy, reliability, safety, and ease of use on a routine basis. The procedures are employed on a routine basis to analyze plant material from experimental sites throughout New South Wales, Australia. EXPERIMENTAL S E C T I O N Plant Material Preparation. Guayule plant material was collected from 17 trial sites throughout New South Wales. The maximum height and diameter of plants were measured. The plants were then either sampled individually or bulked in groups of up to four. Plant tops were cut 7.5-10.0 cm above ground level and the stumps left to reshoot. When whole plants were to be analyzed, the entire plant was dried and separated into tops and roots before chipping. Plants were dried in a forced-air oven at 50 "C for 48 h, leaves were removed by thrashing, and the weight of the remaining woody material was recorded. The material was then cut into small pieces with hand clippers and manually ground with a Corona grain grister (Landers and CIA.S.A., Columbia), which shears the plant material between a stationary and a moving rippled plate with subsequent extrusion of the material in "wormlike" agglomerates. This procedure was later automated by using a Rover compost shredder (with a 3.73 kW motor) to chop the material into small pieces. These were then mechanically ground using a John Bungle and Son No. 7 grain grinder (manufactured in Melbourne, Victoria). This grinder works on the same principle as the Corona grain grister. Samples (5 g) of ground material were analyzed for rubber and resin by the modified Soxhlet procedure or Tecator Soxtec apparatus (3 g). Moisture contents were determined by drying samples (5 g) at 80 "C for 24 h. If necessary, samples were weighed into extraction thimbles and stored in sealed containers with a desiccant at -20 "C before analysis. Samples may be lyophilized before weighing to reduce water contents to 1.0-1.5% of dry weight. Analytical Methods. ( a ) SoxhletlBlend Extraction Procedure. Samples of dried and milled plant material (5 g) were weighed onto Kimwipe tissues (Kimberly-Clark),which were then folded and placed inside Whatman cellulose extraction thimbles (28 X 80 mm). The thimbles were extracted with acetone to remove resins (150 mL) for 2 h in a standard Soxhlet apparatus. Acetone extracts were poured into dried and preweighed disposable polypropylene cups (Sieper co. Ltd., Strathfield, N.S.W., Catalog no. 484) and evaporated to dryness over a steam bath. The thimble compartment of each Soxhlet apparatus was then filled with hexane (150 mL), and the air-dried, sample-containing thimbles were soaked in this, overnight. Samples were then extracted for 2 h and the hexane extracts (containing rubber) evaporated to dryness in preweighed, disposable polypropylene cups, as described above. The solid residues of plant material were transferred to MF24/3 Quickfit tubes modified with I329132 joints. These tubes were modified to allow their direct attachment to a screwcap B29 solvent safe fitting on an IKA Ultraturrax 18K shaft (Figure l),providing a closed system for blending. Acetone (20 mL) was added to each tube, the tubes were immersed in crushed ice, and the contents were homogenized at half maximum speed for 20 s. The homogenates were quantitatively returned to the same Soxhlet thimbles from which they were removed, overlaid with the same Kimwipe tissue, and extracted with acetone
I
449
b Screwcap 829 solvent safe fitting
18K
Shaft
I I W
Figure 1. Tube and blender arrangement for the homogenization of
plant tissue In acetone. for 1 h. The acetone extracts were added to the same cups as the first acetone extracts and the solvent was removed on a steam bath. These extracts were then oven dried (6 h, 80 "C), allowed to equilibrate to room temperature in a desiccator, and weighed to give resin contents. The samples in the Soxhlet thimbles were air-dried and reextracted with hexane (1 h). These extracts were added to the first hexane extracts and dried. Combined rubber fractions, after removal of hexane on a steam bath were oven dried (6 h, 80 "C) and the sample cups cooled in a desiccator before weighing. ( b ) SoxteclBlend Extraction Procedure. Samples (3.0 g) of dried and milled guayule were weighed onto Kimwipe tissues and placed into Soxtec thimbles. The samples were extracted with acetone (60 mL) for 15 min in the boiling mode with the circulating oil at 140 "C. The thimbles were then raised to the rinse position and rinsed for a further 30 min with the circulating oil at 140 "C. If frothing occurred, the temperature of this oil was lowered to 110 "C. The acetone extract was transferred, with washing, into preweighed polypropylene disposable cups and the acetone evaporated over a steam bath. The material in the thimbles was then extracted in the boiling mode with hexane (60 mL) for 15 min at 140 "C (circulating oil temperature) and then in the rinse mode for 30 min at 140 "C (oil temperature). If frothing occurred, the temperature was reduced to 110 "C or more solvent was added. These hexane extracts were rinsed into preweighed polypropylene cups and evaporated to dryness over a steam bath. The extracted plant material was then transferred to homogenization tubes and, after blending, the homogenized plant material was returned to the original thimbles and rinsed with acetone by use of a wash bottle. The washings were collected in the same cups as the original acetone extracts. The original Kimwipe tissues were then placed in the thimbles over the samples and the samples again extracted with hexane (50 mL) in the boiling mode for 10 min at 140 OC (oil temperature) and in the rinsing mode for 30 min at the same temperature. The hexane extracts were added to the cups containing the first hexane extracts and the solvent was removed by evaporation on a steam bath. When all the solvent was removed, the cups were dried at 80 "C for 6 h, allowed to cool in a desiccaQr, and weighed. ( c ) Carbon-13 NMR Analysis of Partially Extracted Samples. Ground plant samples were extracted with acetone for 2 h in a Soxhlet apparatus. The residues were then air-dried and extracted with hexane for from 10 rnin to 8 h. Rubber in the hexane extracts
450
ANALYTICAL CHEMISTRY, VOL. 58, NO. 2, FEBRUARY 1986
Table I. Effect of Temperature and Time of Drying on the Determined Rubber Content (Soxhlet/Blend Procedure) and Moisture Content of GuayuleD
temp, “C
rubber content (moisture content) at the following drying times of whole stems or sections 16 h 24 h 48 h
-20 42 51 82
4.6b (12.4)c 5.2 (15.9) 4.8 (1.9)
4.9 (8.8) 5. I (7.4) 5.3 (1.7)
5.3 (1.6) 5.5 (5.1) 5.2 (2.8) 5.3 (2.0)
=Cultivar593, 20 months old. *Determined rubber content (% w/w). cMoisture content of dried Dlant material. was measured gravimetrically and the plant residues were exhaustively air-dried and analyzed by NMR. The resins and rubber left in the samples were then extracted by the Soxhlet/blend procedure. Original plant samples and solvent extracted samples were also analyzed by NMR. NMR spectra were obtained at approximately 25 “C with a Varian XL-200 spectrometer operating at 50.3 MHz for carbon-13. Samples (1 g) were each packed to the same level in 10-mm NMR tubes. Demineralized water (1mL) was added to each sample. This addition sharpened the NMR signals. The spectrometer was tuned using a D,O sample, which was then removed and the spectrometer run unlocked. A separate sample of extracted guayule rubber was used as the reference for quantitative measurements. The spectra were obtained under conditions of gated decoupling in order to suppress nuclear Overhauser enhancement and with a delay of at least 3T1 between pulses.
RESULTS AND DISCUSSION Several factors complicate the quantitative extraction of rubber from guayule: the rubber occurs as discrete globules in individual cells (2);plant material cannot be finely ground due to the high content of resins and rubber; rubber is polymeric and undergoes hydrophobic and hydrogen bonding interactions with other rubber molecules; drying of plant material may cause oxidative degradation (I),resulting in incomplete rubber extraction; rubber occurs in nature as a normal distribution of molecular weights, thus solvents that extract rubber within a lower molecular weight range may not extract rubber within higher ranges. The problem of obtaining a representative sample of plant material for analysis must also be resolved as the rubber content of individual branches varies. The effect of temperature and time of drying on the determined rubber content of a sample of 20 month old guayule is shown in Table I. Values obtained for freeze-dried material and plants dried a t temperatures up to 82 O C were similar. In freeze-dried material, oxidative cross-linking or depolymerization is not likely to occur, shown by the presence of nondenatured chlorophyll in the acetone extract. Further, since the amount of rubber extracted from all the plant samples is similar, i t can be concluded that under the drying conditions described here, oxidation is not a significant problem if whole stems or sections were handled. However, quite different results were obtained if milled plant material was dried at 80 “C for extended periods. Under these conditions, the amount of extractable rubber decreased dramatically (10-20% in 48 h), being approximately matched by an increase in the amount of the acetone-soluble material. This indicates that the rubber was being depolymerized under these conditions and that this depolymerized rubber can be
Table 11. Effect of Drying and Milling Methods on t h e Determined Level of Resin and Rubber in Two Guayule Cultivars (Soxhlet/Blend Procedure) cultivar (27 months old)
drying and milling methods component
a
b
C
d
593
resin rubber
7.0 6.7
7.0 7.5
7.5 7.8
7.6 7.6
11600
resin rubber
11.1 9.1
11.0 9.3
11.9 10.5
12.0 9.9
“Dried in a forced-air oven at 52 “C for 48 h and ground in a Van Gelder mill. bFreeze-driedfor 48 h and ground in a Van Gelder mill. ‘Dried in a forced-air oven at 52 “C for 48 h and then ground in corn grister. dFreeze-driedfor 48 h and then ground in a corn grister. s16r Resin f r a c t i o n -.
16r Rubber f r o c t i o n
I
a a
6
L-
a I
Time of i n i t i a l a c e t o n e
extraction
hr
Flgure 2. Effect of time of extraction of guayule with acetone on
recovery of resln and subsequent recovery of rubber: first extraction (0);second extraction (0); total (A). The sample analyzed had a moisture content of 7.3% and was from Hansom bulk material. extracted by acetone into the “resins” fraction. This fraction also includes volatile essential oils and nonvolatile terpenoids and long chain fatty acids. Guayule plant material with high levels of rubber and resin is difficult to mill. We found that a standard Van Gelder rotary cutter type mill effectively chopped and granulated the plant tissue but left a layer of rubbery material inside the cutter so that chopped samples were not representative of the original plant material. However, effective milling was achieved with a hand-operated Corona grain grister, which has a shearing-type action, with the sample being ground between a moving T d a stationary rippled plate. The ground material appeared as “rubberlike” worms and represented the true composition of the original material. The effect of drying and milling conditions on the levels of resin and rubber determined in two samples of guayule is shown in Table 11. We consistently obtained higher levels of rubber and resin in material processed through a grain grister than for material through a Van Gelder mill, probably due, in part, to losses of resin and rubber in the Van Gelder mill. The classical Soxhlet extraction procedure of Spence and Caldwell (11) involves extraction over protracted periods of time, making the method tedious and of little value in supporting an agronomic program where large numbers of samples need to be analyzed. We have critically evaluated the times required for resin and rubber extraction and found that most of the resin is removed within 2 h. Extended extraction with acetone removed some rubberlike material (Figure 2), but the problem was not as serious as suggested by Black et al. (15). In fact, increasing the time of extraction from 2 to 8 h caused only a decrease of 0.2% in the measured rubber content. The increase in the amount of acetone solubles with extraction times up to 8 h, rather than being due to depolymerization and extraction of rubber (as suggested by other authors (15)), is more likely to be due to a slow extraction of sugars and
ANALYTICAL CHEMISTRY, VOL. 58, NO. 2, FEBRUARY 1986
L
Extroction
time
451
, hr
Figure 3. Effect of time of extraction with hexane on the determined amount of rubber in guayule. The acetone extracted material was either extracted directly with hexane (0)or presoaked for 16 hr before extraction (0).The amount of rubber extracted by the Soxhiet/blend procedure is shown by the arrow. The sample analyzed was 27 month
old cultivar 11600. I "
Table 111. Effectiveness of the Soxhlet/Blend Procedure in the Extraction of Rubber from Guayule" rubber time of extraction extracted, with hexane, min % ( w / w ) ~ 0 10 20
40 60 180 480
d
rubber content by NMR, %
rubber unextracted; % (w/w)*
(W/W)b
10.4
10.9
2.8 4.1
n.d!
6.3 7.7 10.0 10.5 10.9
n.d.
6.2
3.7 1.6 0.9
3.2
5.9
7.5
1.3 0.4 g
e
"The sample analyzed was from Hansons bulk material. bmg/ 100 mg of acetone extracted plant material. The acetone extract represented 10.8% of the original plant sample. CThiswas determined by the double extraction (Soxhlet/blend) procedure after NMR analysis. Double extraction, the Soxhlet blend extraction procedure as presented in this paper. eNil. Not determined.
/
#Less than 0.4%. amino acids, which are sparingly soluble in acetone. The effect of time of extraction with hexane on the determined amount of rubber in guayule is shown in Figure 3. Also shown is the beneficial effect of allowing the samples to soak in hexane (overnight) before extraction. The inclusion of this presoak step allowed the extractable rubber to be effectively removed within 3 h. However, NMR analysis of the plant material remaining after extraction with hexane for 8 h (Table 111) showed that all the rubber had not been extracted. Consequently, the plant material was then homogenized in acetone with an Ultraturrax blender and further extracted with acetone and hexane. The additional rubber recovered by this operation represented from 5 to 12% of the total rubber extracted, and NMR analysis of the residue indicated that it was devoid of rubber (Figure 4). The recovered rubber was quite pure as shown by the carbon-13 NMR spectra which was like that for pure rubber (1). These results show that the inclusion of an homogenization step after the bulk of the rubber and resin has been extracted greatly improves the value of the Soxhlet extraction procedure for rubber and resin analyses in guayule. The accuracy and
160
120
80 ppm
LO
0
Figure 4. The 50.3-MHz carbon-13 spectrum of acetone extracted guayule piant material (a) before extraction with hexane, (b) after extraction with hexane for 2 hr, and (c) after homogenizatlon and reextraction with acetone and hexane. Sample details are given in Table 111. Peaks A to E are characteristic of rubber.
reliability of the procedure are shown in Table IV where the results obtained on analyzing four selected samples in triplicate on three different days over a 2-week period, is shown. The Soxhlet/blend procedure for rubber analysis was compared to several other published procedures. Table V shows a good agreement between the current method and the Polytron blender method of Black et al. (15). With the Polytron blender method, the most effective solvent was cyclohexane but hexane was more effective in the Soxhlet/blend procedure. The liquid nitrogen/Virtis blender method of Garrot et al. (13)consistently produced rubber analyses which were only 60-80% of those obtained by the Soxhlet/blend procedure. Also, we had difficulty in handling more than three or four samples because a resinous/rubbery mass coated the inside of the mill, which began to heat up even though we continued pouring liquid nitrogen through it. A further complication with this method is that the samples have to be analyzed immediately after they are prepared. Ice crystal formation and water vapor movement in frozen samples made it impossible to obtain a representative sample for analysis after the material had been stored frozen. We found the "blendor method" of Tysdal, as described by Hammond and Polhamus (17),to be of little value as an analytical procedure for rubber in guayule. After the milled fresh material in an ethanol-water-salt mixture was blended, the slurry was poured into water and some "rubbery" material could be obtained (with difficulty). This rubber still contained a high proportion of resins and woody material and represented less than 30% of the rubber in the sample. The remaining rubber was distributed between the woody material remaining on the filter screen and the ethanol-water-salt solution that passed through the filter (the rubber was present as very fine particles). Recently, a fully self-contained Soxhlet-type apparatus, the Tecator Soxtec, became commercially available. This appa-
ANALYTICAL CHEMISTRY, VOL. 58, NO. 2, FEBRUARY 1986
452
Table IV. Determination
2 Rubber and h i i n Content of Guayule Using the Soxhlet/Blend Proceduren day 1
cultivar
day 2
day 3
mean of 3 days % resin
% rubber
% resin
% rubber
% resin
% rubber
10.82 5.11 7.41
10.03 6.73 5.78
10.96 5.07 7.26
9.96 6.75 5.98
10.55 5.01 6.96
9.91 6.72 5.42
10.78 5.06 7.21
9.97 6.73 5.73
3.94
6.22
3.80
6.49
3.83
6.35
3.86
6.35
0.041
0.067
0.041
0.067
0.041
0.067
0.069
0.085
11634 4265 XF Hansom bulk 11591 std error of meanb
% resin'
% rubber
Statistics Summary std dev of single observation rubber resin same day different day
0.071 0.132
coeff of variation, % rubber resin
0.116 0.174
1.1
1.6
2.0 nValues are means of three determinations. bBased on a pooled estimate of the variance for each variety mean.
2.4
Table V. Comparison of the Polytron Blender and the Soxhlet/Blend Extraction Procedures for the Analysis of Rubber in Guayule sample
polytron blender cyclohexane" hexane
A B C
6.50' 12.17
2.70
5.51 11.10 1.88
Soxhlet/ blend cyclohexane hexaneb 6.01 11.61
6.25 13.26 2.17
" The solvent recommended in the standard procedure (15). The recommended solvent for the Soxhlet/blend procedure. 'Determined rubber contents (%, w/wh
20
30
t
0 LO
20
30
LO
Table VI. Rubber and Resin Values for Guayule Extracted with Soxhlet/Blend and Soxtec/Blend Procedure cultivar" Texas wild Texas 174 12331 N565 N576 N396 N593 Texas 33 N565 11609 593 correlation ( r )
resin content, % rubber content, % Soxhlet Soxtec Soxhlet Soxtec 7.44 6.94 6.97 9.44 8.53 8.93 8.78 5.88 8.90 9.81 8.69
7.63 6.98 6.71 9.13 7.86 8.64 8.77 5.70 8.87 9.64 8.32
9.14 6.75 9.12 11.15 8.83 9.39 9.85 9.11 9.42 12.38 7.75
0.98
9.21 6.72 8.76 10.77 8.26 9.26 9.91 8.73 9.04 12.26 7.40 0.99
All samdes were from 34 month old plants. ratus offers the advantage that the samples can be "cooked" in the solvent before the desired components are extracted. This should facilitate more rapid extraction. The unit offers the added advantage of being more robust than the standard Soxhlet apparatus and six samples can be extracted simultaneously. Results for 11 samples analyzed by the Soxhlet/blend procedure or using the Soxtec/blend procedure (see Experimental Section) are shown in Table VI. There is good agreement between the two methods. This equipment is now in routine use in our laboratory, enabling the analysis of 12 samples by a single operator in a normal working day. The procedures described in this paper have been used to monitor the changes in the resin and rubber contents of guayule plant material from 17 experimental sites throughout New South Wales. Typical data for changes in rubber and resin content as well as yield for three cultivars grown on a sandy clay-loam soil are shown in Figure 5. A detailed account
L -20 - - 30 - - + LO- o L o Time
from
planting
,
months
Flgure 5. Changes in the resin and rubber contents and yields per hectare for three guayule cultlvars grown on a sandy clay loam soil in N.S.W. The cultivars are N563 (0),11619 (O),and N565 (A).
of resin and rubber production by a range of guayule cultivars at the various sites throughout New South Wales will be presented separately.
ACKNOWLEDGMENT The authors thank M. Batley, Senior Lecturer, Macquarie University, for performing and interpreting carbon-13 NMR studies. We also thank C. Edge, M. Daniels, and M. Hill for assistance in the preparation of this manuscript. Statistical analyses were performed by C. J. Kaldor. Registry No. Acetone, 67-64-1; hexane, 110-54-3.
LITERATURE CITED (1) "Guayule: An Alternative Source of Natural Rubber"; National Academy of Science: Washington, DC, 1977. (2) Backhaus, R. A,; Walsh, S.Bot. Gar. (Chicago) 1983, 744, 391-400. (3) Campos-Lopez, E.; Neavez-Camacho, E.; Maldonado-Garcia, R. I n "Guayule: Reencuentro en el Deslerto"; Campos-Lopez, E., McGlnnies, W., Eds.; CIQA: Saltlllo, Coahulla, Mexico, 1978; Pub 371. (4) Naqvl, H. H. In "El Guayulero"; Naqvi, H. H., Ed.; Guayule Society Publications: University of California Riverside, CA, 1984; Vol. 6, pp 6-13. (5) Mehta, I. J.; Dhillon, S. P.; Hanson, G. P. A m . J . Bot. 1979, 66, 796-804. (6) Bauer. T. "Guayule Natural Rubber Development Project: 1st Year Report"; California Department of Food Agriculture, Division of Plant Industry: Sacramento, CA, 1979; p 24. (7) Vislntainer, J.; Beebe, D. H.; Myers, J. W.; Hirst, R. C. Anal. Chem. 1981, 53, 1570-1572. (8) Hayman, E.; Yokoyama, H.; Schuster, R. J . Agric. Food Chem. 1982, 30, 339-401.
Anal. Chem. 1986, 58,453-455 (9) Tonnet, M. L.; Downes, R. W., J . Scl. Food Agrlc. 1983, 3 4 , 169-1 74. (IO) Banlgan, T. F.; Verbiscar, A. J.; Oda, T. A. Rubber Chem. Techno/. 1982, 55, 407-415. (11) Spence, D.; Caldwell, M. L. Ind. Eng. Chem., Anal. Ed. 1834, 5,
371-375. (12) Holmes, R. L.; Robbins, H. W. Anal. Chem. 1847, 19, 313-317. (13) Garrot, D. J.; Johnson, D. L.; Rubls, D. D.; DIII, G. M. Anal. Chem. 1981, 53, 543-544. (14) Banigan, T. F.; Verbiscar, A. J.; Weber, C. W. J . Agrlc. Food Chem. 1882, 30. 427-431. (15) Black, L. T.; Hamerstrand, G. E.; Nakayama, F. S.; Rasnlk, B. A. Rub-
453
ber Chem. Technol. 1983, 56, 367-371. (16) Perry, D. A,; Naqvi, H. H.;Hanson, G. P. I n "Proceedings of the 3rd International Guayule Conference, Pasadena, CA, 27 April-I May, 1983";Gregg, E. C.; Tipton, J. L.; Huang, H. T., Eds.; Guayule Rubber Society Publication: University of California, Rlverside, CA, 1983;pp 333-340. (17) Hammond, E. L.; Polhamus, L. G. Tech. Bull.-U.S. Dep. Agric. 196PqNo. 1327.
RECEIVED for review June 25, 1985. Accepted September 9, 1985.
Specific Thermometric Determination of Trace Vanadium in Steel Based on Its Catalytic Effect on the Oxidation of Gallic Acid by Bromate Rafael Forteza a n d Victor CerdB* Department of Analytical Chemistry, Faculty of Sciences, University of the Illes Balears, Spain
The oxldatlon of gallic acid by bromate uslng vanadium as a catalyst was thermometrically monltored. The reactlon rate was obtained graphically from the temperature-time curve. The reaction rate was proporllonal to the V(V) concentratlon In the 2.5-25 pg/L range. The relative standard devlatlon for 15 pg/L vanadlum was 2.0 % Slnce other Ions as hlgh as 6 mg/L did not Interfere at these V levels, thls method can be speclflcally applied to a great number of problems. Thus the proposed method was used for the determlnatlon of vanadium In steel, wlth hlgh accuracy and preclslon.
tribute to the temperature changes of the samples. Furthermore, aging of reagents is not necessary in the thermometric method, since reproducibility is unaffected by this factor.
EXPERIMENTAL SECTION Apparatus. The temperature monitoring system consisted of a rapid response thermistor of the thermometer type (with a nominal resistance value of 100 kR at 25 "C), a Wheatstone bridge fed with 8.93 V from a stabilized source, and a recorder. Sensitivity was 0.01 "C/mV with a full-scale deflection on the recorder of 5 mV (25 cm). This temperature monitoring system is a modification of the apparatus of Lumbiarres et al. (16). The solution was stirred with a 500 rpm synchronous motor. A new design of Several catalytic photometric methods for vanadium dethe adiabatic cell has been introduced with respect to the previous termination have been proposed (1-8). Among the most papers (11-15) in order to improve the thermal stability and thus sensitive and selective is the oxidation of gallic acid, which obtain a greater sensitivity (Figure 1). A syringe was used in order to inject the samples (method 1) is catalyzed by vanadium traces. Jarabin and Szarvas (9) or bromate (method 2) into the thermometric cell. describe a qualitative test for trace vanadium based upon its catalytic action on the oxidation of gallic acid by (NH4)2S208, Reagents. A stock standard solution of 1mg/mL V(V) was prepared by dissolving 2.297 g of NH4V03(reagent grade) in a yielding a brown color. Fishman and Skougstad ( I ) applied 1:100 HNO, solution, which was then transferred into a 1000-mL this reaction to determine trace amounts of V(V) in water (0.2 The volumetric flask and diluted to volume with 1:lOO "OB. pg/L), a method that has been adopted in the field of water latter was diluted with deionized water and renewed daily to analysis. However, this photometric method has some limiobtain the working standard solutions. tations, mainly in its reproducibility. Potassium bromate has Prideaux buffer solution (pH 3.4, I = 0.5 mol/L) (17) was been used as an oxidizing agent, allowing up to 18.2 pg/L (2) prepared by dissolving 12.37 g of boric acid, 23.06 g of concentrated or 0.1 pg/L (8) of vanadium to be determined. In all instances phosphoric acid, 12.01 g of glacial acetic acid, 8.96 g of sodium hydroxide, and 19.98 g of potassium chloride (all of reagent grade) some ions interferred seriously in the photometric methods, in deionized water and diluting to 1 L. including Fe(III), Ti(IV), Mo(VI), and W(V1). Gallic acid solution (near saturation) was 0.05 mol/L in buffer Fishman and Skougstad's method was improved by Weiguo solution, potassium bromate 0.35 mol/L, and sodium bromate (10)in order to increase reproducibility: aging some reagents 1.40 mol/L. for 24 h or more a t 30 "C in a water bath, equilibrating the All solutions were stored in the thermostated room (22 & 1 "C) other solutions for about 15 min in the same water bath, etc. where the experiments were carried out. In previous papers we have studied several methods for the Procedure. Method 1. Ten milliliters of gallic acid 0.05 mol/L determination of inorganic ions based on their catalytic effect in buffer solution, 10 mL of potassium bromate 0.35 mol/L, and monitored with a very simple thermometric apparatus (11-15). deionized water were placed in the cell compartment for a final Among the main advantages we have pointed out is that volume of 50 mL. The recorder was turned on (chart speed, 2.5 cm/min), and when the base line had become horizontal, a volume colored compounds or precipitates do not interfere in the of sample was added with a syringe (maximum 1 mL) to give a thermometric measurements. final vanadium concentration between 2.5 and 25 pg/L. From By the method proposed in the present paper, based on the the resulting curve, the initial rate of reaction was calculated by thermometric monitoring of the catalytic action of vanadium use of the slope (tan a,see Figure 2). in the gallic acid-bromate reaction, we have obtained the same Method 2. In order not to disturb the thermal stability of the sensitivity levels as in the spectrophotometric methods. The cell the volume of sample to be injected must be low with respect main advantage of this method is its specificity, since the to overall volume. Thus, an alternative method to improve colored reactions between the oxidized reagent and the insensitivity was tested: 10 mL of gallic acid 0.05 mol/L in buffer terfering ions in the spectrophotometric method do not consolution, enough sample to give a final vanadium concentration
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0003-2700/86/0358-0453$01.50/00 1986 Amerlcan Chemlcal Society
