tabolism, 2-diethylaminoethyl 2,2-diphenylvalerate HC1 (SKF 525-.4). Studies in which S K F 525-A, piperonyl butoxide, sesoxane, and tropital (piperonal - bis - ( 2 - [2 - 'n - butoxyethyoxy]ethyl) acetal) were employed as inhibitors of the in vivo metabolism of hesobarbital revealed no int,erfering peaks. Method 13 provides a rapid means of screening toxicological specimens for their barbiturate cont,ent. The small sample size needed permits ut,ilization of blood drawn from the finger t'ip or ear lobe. When blood containing phenobarbital, amobarbital, secobarbital, and pentobarbital was extracted according to ;\lethod B, all of t,hese compounds could be readily detected a t the 10 pg,/ml. level. Preliminary experiments indicate that the method can be used without alteration for the determination of drugs
other than barbiturates in blood. For example, Nethod A has been successfully employed for the determination of the in vivo half-life of meprobamate in rats. Experiments designed to extend the method to the analysis of other drugs are in progress. ACKNOWLEDGMENT
The author greatfully acknon-ledges the technical assistance of Victoria Blauth and Susan Shipherd. LITERATURE CITED
( I ) Bonnichsen, R., JIaehly, A. C., Frank, A,, J . Forensic Sci. 6 , 411 (1961). (2) Braddock, L. I., JIarec, S., J . Gas Chromatog. 3, 274 (1965). (3) Hrochmann-Hanssen, E., Svendsen, A. B., J . Pharm. Sei. 50, 804 (1961). (4)Brochmann-Hanssen, E., Svendsen, 9. B., J . Pharm. Sci. 51, 318 (1962). (5) Cieplinski, E. W., ANAL. CHEM.35, 206 11963).
(6) p b i n g , F., Scancl. J . Clin. Lab. Inwst. 7, suppl. 20, 114 (1955). ( 7 ) Gudzinowicz, B. J., Clark, S, J., J . Gas Chimnatog. 3, 147 (1965). (8) Gutenmann, W. H., Lisk, D. J., J . A g r . Food Chena. 12, 46 (1964). (9) Horning, E. C., Tan den Heuvel, W. J. A, Creech, B. G., M e t h o d s B i o c h e m . Anal. 11, 69 (1963). (10) Jain, N. C., Fontan, C. R., Kirk, P. L., AJicroch,ern. J . 8, 28 (1964). (11) Martin, H. F., Driscoll, J. L., ANAL. CHEY.38, 345 (1966). (12) Parker, K. D., Fontan, C. R., Kirk, P. L., Zbid., 35, 418 (1963). 3) Parker, K. D., Kirk, P. L., Zbid., 33, 13i8 (1961). 4) Reith, J. F., van der Heide, R. F., Zwaal, R. F. A., Pharna. Weekblad 100, 219 (1963).
RI. W.ANDERS
Department of Physiology Cornel1 University Ithaca, N. Y. 14850 WORKsupported by U. S. Public Health Service Grant No. GM-13527.
Determination of pK, Values of Some Substituted Nitroaliphatic Acids by Potentiometric Titration SIR: To evaluate the effect of several different functional groups on the acid strength and to obtain information on the feasibility of titrating mixtures of these acids, the pK, values of a number of aliphatic nitroacids have been determined by potentiometric titration in aqueous solution. The acids used were: 3,a-dinitro2 - hydroxybutanoic, 3.3 - dinitro2 - acetoxylbutanoic, 4,4,4 - trinitro- trinitro - 2 - hydroxybutanoic, 4,4,4 butanoic, 4,4,4 - trinitro - 2 - acetoxyl- trinitro - 2 - acetamidobutanoic, 4,4,4 butanoic, 2 - (2,2,2 - trinitroethyl) succinic, and 4,4-dinitropentanoicS Two of these compounds. 4,4-dinitropentanoic and 4,4,4-trinitrobutanoic acids, hare been studied by Long ( 5 ) . The methods used for the determination of pH and pK, hare been described by Albert and Serjeant (2) and by Bates (2), who give extensive references to the literature. EXPERIMENTAL
Materials and Reagents. T h e compounds used were synthesized by the Applied Chemistry Division of these laboratories. All compounds were purified by several recrystallizations from suitable solvents and were dried in a n Abderhalden drying apparatus for several hours a t 65' C. Those compounds which were thought to be unstable at room temperature were analyzed immediately and kept refrigerated. Purity of the compounds was established by elemental analysis and melting point determinations.
National Bureau of Standards grade potassium hydrogen phthalate and ACS reagent grade sodium tetraborate decahydrate (Matheson Coleman and Bell) were used for standardizing the p H meter. The 0.05M solutions of these have p H values of 4.00 and 9.18, respectively, a t 25' C. Additional buffers (pHydrion) were used for the ranges pH 2 0.05 and 8 f 0.05. The distilled water used for dilution and sample preparation was boiled for 5 minutes and stored in a polyethylene container equipped with a glass tube filled with Ascarite. Carbonate-free potassium hydroxide was prepared as described by Albert and Serjeant ( I ) . Apparatus. The titration vessel consisted of a 500-ml. flask i*-ith five necks t o accommodate the following: a IO-ml. microburet, a 5-inch glass electrode (Beckman S o . 41263). a 5-inch reference sleeve-junction type calomel electrode (Beckman 41240) filled with saturated potassium chloride solution, an intake tube for nitrogen, and an outlet tube filled with Ascarite. The nitrogen was passed through a wash-bottle containing a 50% aqueous solution of potassium hydroxide to remove carbon dioxide. This bottle was connected to another bottle containing water to remoye any remaining potassium hydroxide. A magnetic stirrer was used in the solution and the temperature was maintained at 25 & 0.05' C. by a suitable water bath. Procedure. Samples of the nitroacids were weighed in 100-ml. flasks and diluted to volume with distilled viater. The sample was then placed
*
in t h e five-neck flask, stirred for sei-era1 minutes under a n atmosphere of nitrogen, and titrated with 0.1N potassium hydroxide in 10 increments, each containing approximately 0.1 meq. of KOH. After each addition, the pH was read after 1 minute. I n some cases. more than 10 increments were needed to construct a titration curve. The compounds as 0.01X solutions were titrated with 0 . 1 s potassium hydroxide so that activitv effects would bk small. The following equations were used:
pK,"
=
pH
+ log [HA] - log [A-]
pK:
=
pK,"
+ 0.5 dc
(2)
(4)
where, [ ] = analytical concentration, { H + } = hydrogen ion activity, and I , = ionic strength in moles per liter. In cases where the p H fell below 4, corrections for the hydrogen ion concentration were made in accordance with Equation 3. These corrections were hydrogen ion activities calculated from the expression ( H ) = antilog (0-pH) and employed in Equation 3. The resulting constant is a mixed ionization constant between the concentration ionization constant and the thermodynamic ionization constant. This constant is converted to the therVOL. 38, NO. 13, DECEMBER 1966
e
1947
~~~
Table 1.
~
~
pK, Values for Nitroacids
Acid PK3 3,3-Dinitro-2hydroxybutanoic 2.37 =k 0.13 5.42 SC 0.07 3,3-Dinitro-2acetoxylbutanoic 1.85 i 0 . 0 8 4,4,4-Trinitrobutanoic 3.64 st 0.09 4,4,4-Trinitro-2hydroxybutanoic 3 05 =k 0.05 4,4,4-Trinitro-2acetoxylbut,anoic 2,02 =t0.04 4,4,4-Trinitro-22.35 i 0 . 1 2 acetamidobutanoic 3.43 i 0,15 2-(2,2,2-Trinitroethy1)succinic 3 , 0 3 h 0.13 4.42 i 0.09 4,4-Dinitropentanoic 3 , 9 8 i 0 , 0 8 1-Butanoic 4.82 2-Hydroxybutanoic 4.12 3-Hydroxybutanoic 4.29 4-Hydroxybutanoic 4,72 I-Pentanoic 4.81 Succinic 4.17 5.64
logarithm of this value. The scatter is the largest difference between any value in the set and the average pK,. RESULTS AND DISCUSSION
I
(3)
(4)
(4)
(4)
(3) (3) (3)
niodynamic constant by using Equation 4. For 0.01M solutions of the acids titrated, there was little difference in the pKs values. The method followed for titration and calculations is described by Albert and Serjeant ( 1 ) . Its applicability t o this problem was tested by determining the pK, of benzoic acid which was expected to be in the range of some of the acids under investigation. A series of runs was made for each compound. The maximum spread of pK, values was less than =kO.l5, with most of them less than =?=0.10. The pK, values reported, were obtained by averaging the antilogarithms of each value in a set and reporting the
The values obtained for the pK, of the eight acids are given in Table I. For the two acids whose pK, had been determined previously ( 6 ) ,a good agreement was found. For the 4,4,4-trinitrobutanoic acid they were 3.7 us. 3.64 and for the 4,4-dinitropentanoic they were 4.0 us. 3.98. For comparison, the pK, values of 1-butanoic, 1-pentanoic, succinic, and the three hydroxybutanoic acids as given in the literature ( 3 , 4) have been included in the table. ‘The effect of various functional groups on the pK, of 1-butanoic acid is shown in the following series. In order of increasing pK, (decreasing acid strength) they are: 3,3-dinitro2 - acetoxyl -
