tion of o-. m - , and p-chlorophenylalanine and the dichlorinated phenylalanines was accomplished by treating the appropriate 0 chlorotoluene with sodium acetamidomalonate in absolute ethanol: the properties of the resulting substituted benzyl acetamidomalonates were in accord with those r e p ~ r t e d The . ~ ring-substituted phenylalanines were prepared by acid hydrolysis with boiling 6 A' HC1; their physical properties agreed with the reported values and amino acid analysis gave only a single peak (Table 11). Previous references to 2.3-. 2.5-. 2 . 6 . and 3.5-dichlorophenylalanine were not found. Chlorination of [.-Phenylalanine. The halogenations s u m m a rized in Table I were carried out as follows. A magnetically stirred solution of 1.65 g (10 mmol) of 1,-phenylalanine in 200 ml of 6 &VHCI was maintained a t room temperature while 0.71 g 110 mmol) of chlorine in 126 ml of the same solvent was added over a 10-min period. T h e reaction was maintained a t the stated temperature until the disappearance of the yellow color (15 mini and then stirred for an additional 15 rnin. Solvent was then removed in ~ ' a c u oand the crude product was subjected to amino acid analysis (Table I ) . Tlc on Eastman cellulose sheets S o . 6065 (methyl ethyl ketone-pyridine-water-HOAc. 70: 15:l5:2 1 gave Rr~uhei0.36, Rrmo 0.44, R r ~ m p, . a n d 2.38 0.57, Rfr2.5 a n d 3.4 0.64. S e p a r a t i o n of Chlorinated L-Phenylalanines. The separation of 2.0 g (10 mmol) of t h e isomeric mixture prepared above was a t tempted using Bio Rad AG 5OW cation exchange resin (100-1000 m e q u i d m e q u i v of amino acid) with cross linking from 2 to 8%. a solvent variance from 2 AVHC1 t o 6 AVHC1, and t h e same solvents containing an organic phase (methanol. 2-1070,i. All attempted variations failed to give a n adequate separation or yield of pure IV, The fractions were analyzed by tlc a n d amino acid analysis. Attempted fractional crystallization also failed. p-Nitrophenyl-[,-alanine Monohydrate (11). The Bergel and Stocklo procedure was followed (Table I111 except that the crude product obtained was recrystallized from water (3 mlUg) four times to give a 40% yield of pure nitro-L-phenylalanine monohy+7.9 f drate (11): m p 240-242" dec (lit.lo m p 238-241"); 0.2". ( c 1.77, 1.0 N HCli (lit.Io [ ( I ~ ~ +9.8 ~ I I It 0.2"): n m r [DzODC1, 3-1trimethylsily1)propanesulfonic acid as internal reference] 6 3.48 ( d , J = 7.0 H z , 2 H , -CH2-), 4.45 ( t , J = 7.0 Hz, 1 H, -CHCOzH), 7.55 ( d , J = 7.0 Hz, 2 H , aromatic protons, ortho to -CH*-), 8.08 ( d , J = 7.0 Hz, 2 H , aromatic protons ortho to -NOz). Anal. (CgHloX204, H 2 0 removed by heating in high vacuum) C. H . p-Aminophenyl-L-alanine Monohydrate (111). The method of Bergmann" was used and gave a quantitative yield of pure amino-1.-phenylalanine by filtering off the catalyst: m p 242-247" dec (lit,11 m p 235-242"); [ C ~ ] ~-42.0 ~ D i 0.5" (c 1.5. H 2 0 ) (lit." [CY]% -42"); amino acid analysis gave a single peak (Table 11): nmr ( D z 0 ) 6 3.25 ( d , J = 7.0 Hz, 2 H , -CHz-). 4.00 It. J = 7.0 Hz. 1 H. -CHC02H), i.30 is, 4 H, aromatic). p-Chlorophenyl-L-alanine(IV). 4 solution of 7.23 g (40 mmol) of p-amino-1,-phenylalanine in 24 ml of 1.0 lV HCI was mechanically stirred a t 0" while 2.00 g (40 mmol) of S a N O z in 6 ml of water was added over a 20-min period. After stirring for ,5 min longer. t h e resulting solution was added to 5.35 g (,54 mmol) of CuCl in 24 ml of concentrated HC1 at 0" over 20 min. T h e reaction was quite frothy ( a 500-mi flask is adequate for the reaction) with the bubbling continuing as the reaction was heated t o 60" for 30 min with vigorous stirring. The resulting mixture was dissolved in 350 ml of water and H2S was bubbled through the solution until the filtrate was clear. This clear aqueous solution was evaporated in L'UCUO, the residue was dissolved in HzO 1500 mll and neutralized ( p H 6.5) with 3 A' NaOH, and the solution was again evaporated to dryness. Chromatography of the residue on 120 g ot' silica with C H s O H - l i % NHa-CHC13 (10:1.5:13.5) as eluent gave 7.50 g (86%) of pure IY: m p 241-243" dec (lit,7 236-242"): [ L U ] ~ ~ I-3.9 , f 0.3" ( c 2.0. 1 S HC1) (lit.6 -3.5); [(1]'% -27.8 f 0.2" ( c 0.4, H z O ) ; n m r (D20-DCl) 6 3.35 (d, J = 6.5 Hz, 2 H , -CHz-), 4.40 (t, J = 6.5 Hz, 1 H , -CHCOzH), 7.32 ( d , J = 8.0 Hz, 2 H , aromatic protons ortho to -CH*-), 7.36 ( d , J = 8.0 Hz. aromatic protons ortho to -C1). Anal. (C9HloC1NOZ) C, H , N . Optical P u r i t y of IV. Hydrogenation of 0.603 g (3.02 mmol) oi IY in 100 ml of water in the presence of 0.20 g of 10% P d / C was carried out with hydrogen a t 40 psi for 2 hr. After filtering off the catalyst and washing it thoroughly with HzO, t h e resulting solution was adjusted to p H 5.50 with 1.0 A' X a O H and lyphilized. The rotation of the residue (0.678 g: theory, 0.675 g: 100.5%) was -33.90 f 0.05" (c 2.0, HzO), allowing for the presence of NaC1; L-phenylalanine has [ a I z 6-34.13 ~ f 0.03". A series of control experiments using L-phenylalanine and L-phenylalanine hydro-
chloride was performed to ensure t h a t there was no change of optical activity in the L-phenylalanine during t h e hydrogenation. neutralization, or lyphilization. T h e amino acid analysis showed only the phenylalanine peak and the total absence of IV in the hvdrogenated material.
References R . K . Koe and A . LVeissman. .J f'harmacii/. E x p . T h ~ r 131. . 499 (1966). .J. Matsumoto and 11. ,louver. (' K . Soc. Bioi.. 158* 2137 1 19641. S.S.Tenen, I's?chopharmac.oiopin. 10, 204 (1967,. E. E. Shillito, Brit. ;I Pharmacoi.. 38, 305 (1970). R. E. LThalen and It-.G. Luttge. Science, 169, 1000 (1970): A. Zitrin, F. A. Beach. .J. D.Barchas, and W. C. Dement. ihid.. 170, 868 (1970). ,I. H . Tong. c'. Petitclerc. A . D'Iorio. and X. I,. Benoiton. Can. J Hiochr,m., 19,877 11971). J. Burckhalter and V. Stephans. J . Amer. Chem. Soc., 73, j 6 , 3502 (1951). H. Faulstich. H . Smith. and S.Zabeley. . / i ( ~ t i i h Lipbig,\ Ann Chem.. 765 (1973). E. Erlenmeyer and A . Lipp. J u ~ t u sLic>bigsA n n . ('h
