cadmium iodide-starch reagent, prepared from commercially available reagents without further purification, is stable for a t least a month a t room temperature. K h e n purified starch is used, the reagent is stable for a t least 9 months (7'). Onthe other hand opinions are d i d e c l on the stability of the diphenylcarbazide reagent. Urone (10) has recently examined the stability of the reagent and finds that it depends greatly on the type and purity of the solvent used. -1queous solutions TI ere the least stable, although n lien purified solvents v-ere used the reagent Tyas said to be stable for years. Other n-orkers. however, prefer to use diphenylcarbazide 'olutions prepared either frebhly before use (3, 8) or TI hen the stable reagent of Ege and Silverman (4) becomes discolored [l month in the refrigerator (W)]. The latter workers also store the solid reagent in the refrigerator to retard deterioration. Furthermore, Allen (1) finds that the quality of the solid diphenylcarbazide varies with the supplier and suggests that the melting point of the crystalline material be checked before use. Luke (8) describes a fading of the colored product formed between diphenylcarbazide and chroniium(T1). The proposed iodometric procedure, like the diphenylcarbazide method, is nonspecific. I n presence of interfering compounds, such as oxidizing or reducing agents, a preliminary step
might' be required in the presect procedure (prior t'o the iodometric determination) similar to that given to the impurities present when chromium (111) is determined by oxidation tt> chromium(S'1) (prior to reaction v i t h diphenj-lcarbazide). K i t h such modifications the present niethcd n-ould therl become. in effect. an iodonietric determination of chromium. Thus. depeiil iing on the nature of the interferins comyounds. it niiglit he advisnlile T G ash the mmple to destroy wducing material.. -llternatively, inipiritic< might be precipitated or oxidized ii, alkaline niediuni with peroxide if 1' in? bromine ( 1 1 ) or in acid solution n-iti. bisniuthate ( 1 1 ) or permanganate (2: follon-ed by removal of escc;s osidur:l as is usual in these procedures (911. The colorimetric iodometiic ~ ) Y I ' , cedure presented in t81iis report isuggested not only for direct, proceduwinvolving chromiuni(V1) but al-o it 11' back-tit'rations such as the determimition of the organic content of industi' . wastes and sewage by n-et, combustiix n i t h a dilute solution of potassium dichromate in concentrated sulfurii: acid (6). One milliliter of 0.03lT(w./v.) dichromate in 97% sulfuric a c i i was used for t'he oxidation of 0 t i ? 410 fig. of carbohydrates or proteins i: 0.45 ml. of solution ( 5 ) . The escws dichromate was determined by t h starch-iodide reaction after dilution :I: the dichromate-sulfuric acid digest t o the concentrations described above.
Since the present paper was submitted, Vdovenko and Spivakovskaya ( 12 ) ha\-e reported an iodonietric deterniiiiation of chroniiuni in steel using potaGaiuni iodide. The latter reagent was used in the early stages of the present investigation on the iodonietric determination of chromium(VI), but n-as tliscontiniied n-hrn the more stable cndniiuni iodide-starch reagent became n T X ila ble , ACKNOWLEDGMENT
The author sincerely thanks George Pratt for valuable technical assistance. LITERATURE CITED
(1) Allen, T. L., ANAL. CHEW 30, 447
119581.
(2)' Cahnmann, H. J., Bisen, R., I b i d . , 24, 1341 (1952).
(3) Cline) R. W., Simmons, R. E., Rossmasler, \I-. R., I b i d . , 30, 1117 (1958). (4) Ege. J. F.. Silverman. L.. Z h i d , 19,
(5) Hallin-ell, G., Biochenz. J . 74, 4-57 iim).
(GCJohnson, 11. J , J . Bid. Chem. 181,
107 (1949).
( i )Lambert, J. L.,
.%SAL. CHEX 23, 1'247 (1951). (8) Luke, C. L., I h i d . , 30, 359 (1958). (9) Saltsman, B. E., I h i d . , 24, 1016
i l-9_5_ 2 )-
(10) Crone, P. F., Ihid., 27, 1354 (1955). (11) Urone, P. F., Anders, H. K., Ibid., 22, 1317 (1950). (12) Vdovenko, 11. E., Spivakovskaya, S . E.. Z a c o d s k a v a L a b . 25. 416-17 (1959); Chem. Ab&-. 53, 12941 (1959). RECEIVEDfor review Jdj- 21, 1959. Accepted April 25, 1960.
Micropyrolytic-Gas Chromatographic Technique for the Analysis of Organic Phosphates and Thiophosphates C. E. LEGATE and H. D. BURNHAM Wood River Research Laborafory, Shell Oil Co., Wood River, 111.
b A technique is described for the identification of organic radicals in organic phosphates and thiophosphates. The compound is pyrolyzed in the inlet system of a gas chromatograph, and the volatile pyrolysis products (generally olefins) are fractionated and collected individually for identification by mass or infrared spectrometry. The olefins are formed generally by breaking of a carbonoxygen bond and abstraction of hydrogen from a beta carbon atom with no skeletal isomerization. Thus, the structures of the olefins are directly related to the structure of the alkyl groups initially present. Only when hydrogen i s not available on a beta carbon atom-e.g., neopentyl radicals-are olefins formed by carbon-skeletal rearrangement. Examples are given of 1042
ANALYTICAL CHEMISTRY
the determination of the exact configuration of the alkyl radicals in several model organic phosphates and metal dialkyl thionothiophosphates. A gas chromatographic inlet system suitable for pyrolysis or for conventional vaporization is described.
Zinc dialkyl thionothiophosphates of the following general structure have
s t
s
0
0
R-0-p-S-Zn-S-P-0-R
t
I
R
R
been used for many years as antioxidants and load-carrying agents. They are present in commercial motor oils and in commercial additives. These
additives vary in thermal stability and performance characteristics and it has been reported (3) that these differences reflect variation in the nature of the alkyl groupings. The chemical reactivity of alkyl and alkyl aryl phosphates and thiophosphates makes characterization of the exact structure of individual members of a given class difficult. They are, for example, extremely resistant to hydrolysis, so that the alcohols from which they n-ere synthesized cannot readily be regenerated for identification. On the other hand, their poor thermal stability prevents obrervation of the parent peaks in the mass spectra which is so useful in the identification of the aryl phosphates. The mass spectra provide a clue, honever, which has led t o a solution of the problem.
conipound to be analyzctl
;1 1ieliu:ii
flow rate of 15 to 20 nil. per minute,
c c c c It c=c-c-c+c-c-e-c cis
c
)] c
A - - - - - +
[ r" ] +
2 i 5 " C . Pb --S-P(OH),
2
(hypothetical nonvolatile product)
+ trans
The structures of the olefins are directly related to those of the alkyl groups initially present in the dithiophosphate and thus can be used to determine the structure of this type of compound. The nonvolatile reaction product shown is presented only as a hypothetical possibility became this material was not recovered from the pyrolyzer for identification. JIicroreactor-gas chromatography has been applied to the determination of organic structures by several other investigators. Zlatkis, 01-6, and Kimball (5) analyzed mixtures of amino acids by oxidizing them in a microiractor and separating the resulting T olntile aldehydes on a chromatographic column in series n-ith the reactor. Lehrle and Robb ( 2 ) identified unknon n polymers by studying the thermal degradation products of polymers n-hich were pyrolyzed rapidly in the inlet system of a gas chroniatograph. Thonipson et al. (4) h a r e developed a micromethod for the catalytic removal of sulfur from complex sulfur compounds, for which there were no standards, to yield corresponding hydrocarbons nhich were ai-ailable as standard compounds.
range of possible pyrolysis products in preference to a more specific column of higher resolution but more limited applicability. Conventional GLC fraction-collecting techniques were employed. Procedure. It is often possible t o learn t h e general nature of t h e alkyl and/or aryl radicals present from t h e preliminary mass a n d infrared spectral analysis of t h e phosphate or thiophosphate. If t h e approximate decomposition temperature of t h e compound is also known, suitable experimental conditions for t h e micropyrolytic-gas chromatographic analysis can be readily selected. Column temperatures are chosen according to the boiling range of the volatile decomposition products expected, while pyrolyzer temperatures are chosen according to:the thermal stability of the
providing a residence time in the pyrolyzer of approximately 5 seconds. has been satisfactory. -4, sample of 5 p1. is usually sufficient for obtaining fractions large enough for &IS analysis. For infrared analysis of fractions, it may be necessary to use 25 to 50 pl. 01' make replicate runs t o identify minor pyrolysis products. If both heavy and light fragments are expected, as in the case of some alkyl aryl compounds, it may be necesqary to empioy two or more column temperatures. The heavy fragments are identified first and the column temperature is then lowered sufficiently t o resolve any lighter pyrolysis products eluted as a group at the higher column temperature. With the column a t 30" C., decomposition products of Cs to Cg alkyl substituents can he sufficiently well resolved to meet nioet analytical needs. Organic phosphates and thiophosphates are generally viscous liquids which can be handled by syringe after warming. Those nhich are solid or very viscous can be diluted with a small amount of some thermally stable solvent having a noninterfering retention volume-acetone, benzene, or toluene. If no structural or thermal stability information is available, a series of exploratory experiments is required, in n hich the temperatures of the pyrolysis chamber and analytical column are varied. Several precautions should be obseri ed to obtain the maximum information from t h w experiments. The sanlple 'ize should he hqJt as small as
I
PYROLYSIS CHAMBER AUALYTICAL
-
-
W P L MLET
ELIL M
EXPERIMENTAL
Apparatus. T h e only special feat u l e of t h e chromatograph used in t h e investigations is t h e modified inlet system shon 11 in Figure 1. This inlet system is suitable for either pyrolysis or conventional vaporization. It consists essentially of a 3 X l,'4 inch stainless steel t u b e packed n i t h quartz wool a n d heated TT i t h a t u b e furnace having a rating of 200 watts. T h e furnace, n-hich is controlled by a variable tiansformer, is capable of heating the pyrolysis chamber to 1000" C., although the maximum temperature employed thus far is 600" C. Samples are injected with a 50-pl. capacity syringe, through a self-sealing rubber cap attached to an air-cooled extension of the pyrolyzer. The analytical column is a 6 foot x 1/4 inch coiled copper tube packed v i t h
THERMOSTAT DOOR
Figure 1 . GC inlet system suitable for pyrolysis or conventional vaporization Stainless steel tube, ' / s X 1 inch, fltted with syringe rubber cap (Fisher Scientific Co., Catalog No 3-21 5 ) 2. Stainless steel tube, 3 X '/4 inch, packed with quartz wool 3. Stainless steel union, Swagelok, '/a-inch tube to '/4-inch tube 4. Copper tube, '/4 inch X suitable length 5. Stainless steel tube, '/a X 7 inches 6. Iron-constantan thermocouple 7. Alundum tube, 2l/2 X 1 inch in outside diameter X "/le inch in inside diameter, wound with 4 feet of 1 9 - g a g e Chromel-A wire which is then coated with Alundum cement 8. Alundum tube, 23/4 X l'/s inches in outside diameter X 1 '/2 inches in inside diameter 9. Transite disk, 17/s inches in diameter X inch thick 10. Transite platform, 3 l / 2 inches square, and mounting brackets 1 1 . Steel bands, "16 inch wide (supports for tube 7 and electrical connections for heater wire) Parts 1 , 2 , 5 , and 6 a r e silver-brazed together 1.
VOL. 32, NO. 8, JULY 1 9 6 0
1043
Table I.
Source and Purity of Model Compounds
Compound Source Lead salt of 0,O’-din-amyl thionothiophosphate Lnboratory synthesis ( 1 ) Zn salt of 0,O’-di-ndodecyl thionothiophosphate Laboratory synthesis Zn salt of 0.0‘-dineopentyl thionothiophosphate Laboratory synthesis Potassium d t of 0,O’-diisopropyl thionothiophosphate American Cyanamid Co. Lead salt of 0.0’-di(4-methvl-2pentyl jthionothiophosphate Kolker Chemical Co. Zn salt of 0,O’-dicyclohexyl thionothiophosphate Iiolkcr Chemical Co. 2-Ethyl-I-hexyl diphenyl phosphate llonsanto Chemical Co.
Table
(I.
Analysis Theorr Found %lI % P SCS 7 c 1 1 % P
5s
27 8
13 1
8 3 17 2 21 3
6 0
129
6 5
5 8 108
10 8 10 3 2 1 . 2
10 8
10 5 20.3
6 6
6 2
Commercial grade
258
7 7 1 6 0 2 3 8
11 9 11 2 23 2
8.9
7 7 1 4 7 8 4 14 7
Commercial grade
Pyrolysis of Model Phosphate and Thiophosphate Compounds
Temneratiirr, C. GC Pyrolyzer column O
Model Compound Pyrolyzed
Principal Volatile Products
Primary Alkyl Structure Pb salt of 0,O’-di-n-amyl thiono- 315 31.0 1-Pentene thiophosphate Zn salt of 0,O‘-di-n-dodecyl thiono- 290 150 0 1-Dodecene thiophoep hat e Zn salt of 0,O’-dineopentyl thiorio- 460 30 0 2-Methyl-1-butene (42-n t %)a t hiophosp hat e 2-Methyl-2-butene (56 1% t %) Ca and lighter olefins ( 2 wt. C;, Secondary Alkyl Structure K salt of 0.0’-diisopropyl thiono- 300 30 0 Propene thiophosphate Pb salt of O,O’-di(4-methyl-2-pentj-l) 275 30 2 cis- and (runsthionothiop hoephat e 4-Mcthyl-2-pentene (59 R t. Fc) 4-Methyl-1-pentene ( 3 5 v-t. yc) 2-llethyl-2-pentene (6 xt. Zn salt of 0,O‘-dicyclohexyl thiono450 30.0 Cyclohewne thiophosp ha t e Alkyl Aryl Structure 2-Ethyl-1-hexyl diphenyl phosphate 230 108.0 2-Ethyl-1-hexene (63 wt. %) Phenol (37 wt. %) a Basis total volatile pyrolysis products.
possible consistent with the sensitivity of the gas chromatograph and sample requirements of any auxilliary methods employed for identification of GLC fractions. Small samples not only prevent overloading of the GLC column but permit the use of short residence times in the pyrolyzer, which in turn allow very little time for side reactions to occur. The analytical column temperature should be kept relatively high during initial experiments with compounds containing unknown organic radicals to prevent the possibility that a heavy alkyl or aryl fragment might escape detection by remaining on the column. The temperature of the pyrolysis chamber should be high enough to produce flash pyrolysis of the sample.
1044
ANALYTICAL CHEMISTRY
The temperature a t which peaks are first detected on the chromatogram is the initial decomposition temperature of the compound under the chosen experimental conditions of residence time, atmosphere, and internal surface characteristics of the pyrolysis chamber. If the peaks are broad and poorly resolved, slow decomposition of the sample is indicated, because the decomposition products are reaching the analytical column in a dilute state rather than as the desired plug of vapor. The pyrolyzer temperature is increased until the peaks on the chromatogram are sharp and well resolved, indicating that flash pyrolysis of the sample is occurring. Detection of large amounts of carbon dioxide or sulfur dioxide indicates that the pyrolyzer temperature
is too high. Such complete degradation of the molecule prevents identification of the principal groups. Approximately 500 pl. of nietal dithiophosphates can be pyrolyzed in the inlet system before cleaning is necessary. If this limit is not exceeded, results of replicate analyses agree satisfactorily both qualitatively and quantitatirely. Cleaning of the inlet system requires about 1 hour. The unit can be detached from the chromatograph in a few minutes. -4fter removal of the quartz wool, the pyrolysiq tube iq reamed in a drill press to remove all deposits. packed with fresh quartz n 001, and reinstalled. PYROLYSIS
OF
MODEL
COMPOUNDS
The validity of the technique in identifying various types of alkyl and one type of aryl side chains in organic phosphates and thiophosphate? is shown by the results of the analysis of s e w n model compounds. Table I lists the source and purity of the compounds, while Table I1 summarizes the pyrolyses. As shown in Table 11. olefins are formed generally by cleavage of the C-0 bond, folloned by simple abstraction of hydrogen from a beta carbon atom with no skeletal isomerization. Thus the structures of the olefins are, generally, directly related to the structure of the alkyl groups initially present. Straight-chain primary alkyl structures form only one olefin during pyrolysis. Similarly, the cyclohexyl group forms only one olefin but requires a high temperature to produce a reasonable yield of cycloheuene. \There more than one olefinic product is possible, all iqomers are found. I n the 4-methyl-2-pentyl structure shown in Equation 1 t n o beta carbon atoms are available from which hydrogen can be abstracted. The three hexenes expected are found a5 well as a minor amount of 2-methyl-2pentene. The latter olefin is therniodynamically the most stable hexene and presumably arises from isomerization during the pyrolysis. Only when hydrogen is not available on a beta carbon atom are olefins formed by carbon skeletal rearrangement. Such is the case Jyith the dineopentyl thionothiophosphate. This compound is stable thermally because the formation of olefins requires the cleavage of a C-C bond as ne11 as a C-0 bond. The neopentyl groups were converted almost entirely to C5 olefins, the total amount of C4 and lighter olefins being only 2 weight % of the volatile pyrolysis products. The 2-methyl-1-butene and 2-methyl-2-butene which were formed (presumably by a carbonium ion path) represent the minimum amount of carbon-skeletal rearrangement which can lead to the production of olefinic produ
Ambiguities can arise in which the detection of a specific olefin or olefins alone cannot be considered as positive evideiice for tlie presence of one specific alkyl group. In these situations, auxiliary spectroscopic or thermal stability data can often be useful in making specific idcntifications. For example. a propyl structure represents a n amhigiious cnse because propene could be formed upoil pyrolysis of either a propyl or an isopropyl group. T o establish that the propyl group indicated is of the iso rather than the normal configuration it i i nwessary to obtain the infrared spectrum of the thiophosphate. A n absorption doublet near 7.25 microns is characteristic of structures in which trio methyl groups are attached to a common tertiary carbon (Smethyltype l~rnncliing) . DISCUSSION
I lie niicropyrolytic-gac chromatographic tcclinique offers a practical method for the characterization of org;anophosplioru;IrLi~ compounds n-hich are too unstable to permit production of parcnt ion; in the mass spectromcter. r 7
It has the advantages of speed, small sample size requirement, and high sensitivity coupled with the possibility that the technique can be made quantitative with proper calibrations. Approximately one-half hour is required for most pyrolyses and the subsequent separation of the pyrolysis products. The time required for identification of the GLC fractions necessarily varies according t o the complexity of the sample and the specificity of identification required. Limited attempts to scale up the method t o provide larger quantities of pyrolysis products have not been successful. It is believed that the success of the tcchnique is due largely to the short residence time in the pyrolyzer which allon-s very little time for side reactions to occur. The technique might be applied advantageously in studies of the thermal stability and the kinetics of decomposition of organophosphorus conipounds by observing the nature and quantitative distribution of pyrolysis products as a function of temperature. The method might also be applied in determining the structure of other com-
pounds k n o m to pyrolyze readilyesters, mercaptides, polymers, etc. ACKNOWLEDGMENT
The authors are indebted to L. W. Taylor and G. P. Pilz for the mass and infrared spectral measurcnwnts, respectively, and to S . K. iYc,lson for the synthesis of the model thiophosphates prepared in this laboratory. LITERATURE CITED
(1) Freuler, H. C. (to Union Oil Co of Cahfornia), S. Patent 2 , 3 6 4 , 2 8 3 4
u.
(Dec. 5, 1944). ( 2 ) Lehrle, R. S , Rohb, 6. C., n’atztre 183, 1671 (1959). ( 3 ) Scanley, C. S Larson, Read, SAE Preprint 107C, SAE Sational Fueland Lubricants RIeetinq, Tulsa, Okla., SOV. 5-6, 1958. (4) Thompson, C. J , Co!eman, H. J., Ward, C. C., Rall, H.T., A s ~ L .CHEV 32,424 (1960). (5) Zlatkis, A., Or6, J F , Kimball, -4. P., %d., 32, 162 (1960). ~
RECEIVED for review December 21, 1959. rlccepted March 30, 1960. Pittsburgh Conference on Analvtical Chemistrv and Applied Spectroscm&, Pittsburgh,” Pa., March 1960.
Report on Recommended Specifications for Microchemical Apparatus Volumetric Glassware.
Micropipets
Committee on Microchemical Apparatus, Division of Analytical Chemistry, American Chemical Society Chairman, Hoffmann-La Roche hc., Nutley, N. J. Arthur H. Thomas co., Philadelphia, Pa. V. A. ALUISE, Hercules Powder co., Wilmington, Del. E. W. D. HUFFMAN, Huffman Microanalytical laboratories, Wheatridge, colo. E. L. JOLLEY, Corning Glass Works, Corning, N. Y. J. A. KUCK, College of the City of New York, New York, N. Y., and American Cyanamid co., Stamford, Conn. J. J. MORAN, Kimble Glass co., Vineland, N. J. C. L. OGG, Eastern Utilization Research & Development Division, Agricultural Research Service, U. S. Department of Agriculture, Philadelphia, Pa. C. E. PETRI, U. S. Atomic Energy Commission, New Brunswick, N. J. A t STEYERMARK, H. K. ALBER,
I
x ACCORDANCE with the practice follo\red in all previous reports (9-4) of the Committee on Rlicrochemical Apl)aratus. these specifications are for pieces of appmttue that are eit’her the most widely used in their respective fieltls of application or are an improven i m t over such apparatus according to twth made hy members of this comniittw and cooperating chemists. I n thc 1:itter case. emphasis is placed on spwificatioiis which increase the general 11s~fi11nessof the particular apparatus, aiid simplify its fabrication. 111 this rpport, specifications are rec-
ommended for micropipets, Folin-type ( 5 ) (Figure I ) , micro washout pipets ( 6 ) (Figure a), and micro weighing pipets, density-type (pycnometer) (1) (Figure 3). These pipets are classified as micropipets, whereas pipets in a similar capacity range were classified as microliter pipets in the 1958 report of this committee. This should not be construed as a n attempt by the committee to differentiate among the various pipets on a technical basis, but rather on the basis of usage. This is done to prevent confusion which might result from the fact that the pipets
shown in Figures 1 and 2 are listed in trade catalogs as micropipets, designed for use in the fields of biological and clinical chemistry and the pipets shown in Figure 3 are listed as micropipets. density-type, designed for use as micro weighing pipets. The Folin-type pipets are specified in 0.1- and 0.2-ml. capacities. T o ensure complete delivery of the volume indicated b y the graduation mark on these pipets, it is general practice to wash out material adhering to the inner surface with small amounts of wash liquid drawn up from tlie tip. VOL. 32, NO. 8, JULY 1960
1045
