II. Assessment by tryptophan metabolites, vitamin B6, and pyridoxic acid levels in urine1 2

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1 The vitamin B6 requirement in oral contraceptive users. Assessment by tryptophan metabolites, vitamin B6, and pyridoxic acid levels in urine1 2 lizabeth A. Donald,3 B.Sc., M.S., Ph.D. and Teresa t Boss#{233},4B.Sc., M.Sc. ABSTRACT Data were obtained from seven college-age women who used estrogen-containing oral contraceptives and who volunteered to participate in a metabolic study to investigate the requirement for vitamin Be. They consumed a basal diet containing.36 mg/day of the vitamin for 42 days. During the first 1 days (predepletion period) the diet was supplemented with pyridoxine hydrochloride to bring the total intake to 2.6 mg/day. After 32 days of depletion the diet was supplemented with pyridoxine hydrochloride for a daily intake of.96, 1.56, and 5.6 mg for 8, 9, and 7 days, respectively. Complete 24-hr urine collections were made and composited. Urine was analyzed for total vitamin B, and 4-pyridoxic acid; and for kynurenine, 3-hydroxykynurenine, kynurenic and xanthurenic acid levels both, before and after 2 g loading doses of L-tryptophan. Results were compared with data obtained from non-oral contraceptive users who consumed a similar diet. Tryptophan metabolites significantly increased during the depletion phase and rapidly decreased with pyridoxine hydrochloride supplementation. A vitamin B, intake of 1.56 mg/day was sufficient to restore all metabolites to original levels in oral contraceptive users. Urinary vitamin B. and 4-pyridoxic acid levels decreased during depletion and were returned to original levels with 1.56 mg/day. An intake of 5.6 mg caused large amounts of each to be lost in the urine. The data obtained suggest that.96 mg was not adequate to meet the needs of oral contraceptive users as predepletion levels of most parameters studied had not yet been reached. An intake of 1.56 mg restored levels in almost all subjects and 5.6 mg caused large amounts of vitamin B. and 4.. pyridoxic acid to be excreted. An intake of 1.5 mg is similar to the previously suggested intake for nonusers. Am. J. Clin. Nuir. 32: l4.-132, Women using oral contraceptives (OC) containing synthetic estrogen and progestogen combinations excrete abnormally high levels of tryptophan metabolites after a load dose of tryptophan. Several studies (1-8) have indicated that such abnormalities are correctable by intakes of 2 to 4 mg of pyridoxine hydrochloride (PN-HC1). Other investigators have found that other parameters used to measure the vitamin B6 requirement do not indicate the need for excessive intakes of vitamin B6 by OC users. Lumeng et al. (9) found low plasma pyridoxal levels in only four of 11 subjects who demonstrated abnormally high xanthurenic acid (XA) levels in the urine. Two recent studies (1, 11) assessed vitamin B6 needs in OC users using a multiparameter approach. Findings indicated that.8 mg of pyridoxine was not adequate to correct tryptophan metabolism in OC users but 2. mg restored all but XA excretion to initial levels. Vitamin B6 intakes between.8 and 2. mg also restored urinary 4-pyridoxic acid (4-PA) as well as several parameters in the blood. The authors concluded that OC users had no need for high intakes of vitamin B6, and the use of OC would have limited clinical effect on vitamin B6 nutnture. n the present study a multiparameter approach was also used to assess the requirement for vitamin B6 by OC users. The effect of load-doses of tryptophan on the excretion of metabolites was observed; as well as the From the Faculty of Home conomics, University of Alberta, dmonton, Alberta, Canada T6G 2M8. 2 Supported in part by a grant from The Medical Research Council, Ottawa, Canada. Professor. 4Sessional Lecturer. Downloaded from ajcn.nutrition.org by guest on February 12, The American Journal of Clinical Nutrition 32: MAY 1979, pp Printed in U.S.A.

2 VTAMN B, RQURMNT N OC USRS 125 amount of vitamin B6 and 4-PA excreted in urine. Requirement as assessed by pyridoxal level and aspartate and alanine amino transferase activity in erythrocytes has been reported elsewhere (12). Subjects ight college-age women using OC and eight nonusers served as experimental and control subjects, respectively. Subjects were judged to be healthy and informed consent was obtained from each. Details of subjects have been previously reported (12). Oral contraceptives Subjects used a variety of contraceptives for 1 to 25 months. Seven of the eight used a combination type and one used a sequential preparation. See a previous report for details (12). Diet The diet was similar to one used in a previous study of the vitamin B6 needs in non-oc users (13). The basal diet was supplemented where needed to meet the 1968 Recommended Dietary Allowances (14). The diet contained.36 mg of vitamin B6. nergy needs of subjects were met by supplying, ad libitum, high energy foods such as butter, candy, etc. Details have been reported previously (12). xperimental design During the 1-day predepletion phase both control and experimental subjects consumed the basal diet supplemented with 1.7 mg of PN-HC1. The total daily intake was 2.6 mg which met the adult Recommended Dietary Allowances (14) for vitamin B6. Only experimental subjects underwent the depletion phase. They consumed the basal diet for 32 days. The diet was then supplemented with PN-HC1 for total daily intakes of vitamin B6 of.96, 1.56, and 5.6 mg for 8,9, and 7 days, respectively. Metabolic studies Complete 24-hr urine collections were made throughout the study. Collections were composited and frozen at -2C until analyzed. Completeness of collection was checked by determining creatinine levels (15). Data for one subject in the experimental group have been omitted from calculations of metabolites in urine because oflow creatinine values. A load-dose of 2. g of L-tryptOphan was mixed with chocolate and given with breakfast on days 2, 11, 18, 25, 32, 39, 43, 5, 59, and 66 to the experimental subjects. Urine collected on these days (postload) was analyzed for XA and kynurenic acids (1(A), kynurenine (Kyn), and 3-hydroxykynurenine (3- OH-Kyn). Urine collected on days previous to the ones listed above (preload) were analyzed for the same tryptophan metabolites. Control subjects were given a load dose of tryptophan on day 2 and day 1. Analyses for the same tryptophan metabolites were made on these days, as well as for preload values on days 1 and 9. An elution columnchromatographic method, followed by fluormetric or spectrophometric determinations of individual metabolites was used (16). Urine collected on the pretryptophan load days was analyzed for vitamin B6 according to the Atkin s method as described by Storyick et al. (17) using Saccharomyces carlsbergensis as the test organism. Urine collected on the post tryptophan load days was analyzed for 4-PA using the method described byfujitaetal.(l8). Results Tryptophan metabolites The amount of Kyn (32.3,LM/24 hr) excreted during the 1th day of the predepletion phase by subjects using OC was significantly higher (P <.1) than amounts excreted during day 9 (1.2 tm/24 hr) by subjects not using OC when both groups had been given a 2. g load-dose of tryptophan (Table 1, Fig. 1). The levels excreted by control subjects given the load-dose were similar to levels found in experimental subjects not given tryptophan. The level of Kyn in the urine collected from subjects using OC significantly increased (P <. 1), both pre- and postload, during the depletion phase. These pre- and postload levels were significantly different (P <.1) from each other at the beginning as well as at the end of the phase; 6.7 versus 91.2 LM/24 hr and 4. versus 149.,tM/24 hr, respectively. An intake of.96 mg of vitamin Downloaded from ajcn.nutrition.org by guest on February 12, 213

3 126 DONALD AND BOSS TABL 1 Mean ± S; urinary excretion of tryptophan metabolites in subjects using and not using OC (tm/24 hr) Oc uscrsh Nonusers Pretryplophan B, in diet (mg) Daysondiet Kyn 7. ±.5 KA 1.7 ±1.3 XA 11.9 ± ± ± ± ±.6 9. ± ± ± ± ± (6) ± ± ± ± ± ±1. 4. ± ±.6 1. ± ± ± ± ± ± ± ± ± ±.5 d 12.2 ± ±1.3 d 1.3 ± ±1.1 Posteryptophan Daysondiet Kyn 47.(6) ± ± ± ± (4) ± ± ± ± ±2.6 3-OH Kyn 5.1(6) (6) 126.6(4) 141.8(6) ±.6 ±4.6 ±17.3 ±29.7 ±39.2 ±42.7 ±1 1.1 ±1. ±.4 KA (6) ±25.8 ±8.6 ±9.7 ±12.4 ±19.5 ±1.6 ±12.8 ±8.3 ±6.6 ±8.8 XA ±21.8 ±28.1 ±4.7 ±61.5 ±11.4 ±38.1 ±43.9 ±32.2 ±12.1 ±3.1 a Values for 3-OH Kyn users too low to read accurately. b Seven subjects unless noted in parentheses. subjects. d Values too low to read accurately. 6, 6. VTAMN B6 NTAK.MG/DAY 15.8 ±2.8.6 ± ± ± ±.4 5. ± ± ±2.3 ight Downloaded from ajcn.nutrition.org by guest on February 12, DAYS ON DPLTON-RPL11ON DT FG. 1. Mean urinary excretion of tryptophan metabolites after a loading dose of 2. mg of tryptophan in OC users. XA, a u u u a ; Kyn, i ; and 3-OH kyn, B6 for 8 days caused a further, but insignifi- plied either 1.56 or 5.6 mg of vitamin B6. cant increase in the excretion of Kyn, both These levels of intake returned excretion valpre- and postload. xcretion levels dropped ues to the range found during the predepledramatically (P <.1) when the diet sup- tion phase.

4 VTAMN B, RQURMNT N OC USRS 127 During the predepletion phase all subjects, both users and nonusers of OC, excreted similar levels of 3-OH Kyn after administration of the tryptophan load-dose (Table 1, Fig. 1). Nonusers excreted 5..LM/24 hr and OC users excreted 9.8.tM/24 hr. The level of 3-OH Kyn excreted before the tryptophan load was too low to measure. Consumption of the basal diet during depletion resulted in increased urinary levels of 3-OH Kyn (P <.5) in users. Consumption of.96 of vitamin B6 decreased excretion levels (P <.5). Further supplementation of the diet with PN-HC1 to either the 1.56 mg or 5.6 mg level ofmtake restored 3-OH Kyn to levels similar to those found during the predepletion phase. Preload KA levels in control and experimental subjects were similar and remained almost constant throughout the entire study regardless of vitamin B6 intake (Table 1). Administration ofthe tryptophan load caused a significantly elevated level of KA (P <.1) in both groups of subjects. Postload KA excretion by OC users at the end of the depletion phase was significantly higher (P <.1) than during the predepletion phases. KA excretion decreased (P <.1) and reached the predepletion level of approximately 5 LM/24 hr after consumption of.96 mg of vitamin B6 for 8 days followed by 1.56 mg for 9 days. Preload XA levels in experimental and control subjects were similar in the predepletion phase (Table 1, Fig. 1). Administration of tryptophan produced increases in excretion in both groups. While the nonusers exhibited an increase from approximately 9 to 3 tm/ 24 hr (P <.5), the OC users showed a more significant increase from 11 to 13 M/24 hr (P <.1). The consumption of the low vitamin B6 diet caused a marked increase (P <. 1) in the excretion of XA in experimental subjects and the various levels of supplementation resulted in a rapid decrease in excretion to levels less than originally observed. f no tryptophan load was given, the excretion of the metabolite remained almost constant regardless of vitamin intake (Table 1). Figure 2 was obtained by calculating the net yield of tryptophan metabolites using the method of Leklem et al. (11). This illustrates the marked increase in excretion of metabolites when subjects using OC were fed low levels of vitamin B6 and the marked decrease when the diet was supplemented with the vitamin. When the vitamin B6 intake was 2.6 mg/day the net yield of metabolites was greater in subjects using OC than in those not using OC. Total vitamin B6 excretion The excretion of vitamin B6 in urine is shown in Table 2 and Figure 3. Results obtained for the present diet were compared with results obtained in subjects after a similar diet but not using OC. These results have been previously reported (13). When 2.6 mg ofvitamin B6 was consumed during the predepletion phase the amount of vitamin B6 excreted in the urine was similar for both experimental and control subjects (Table 2). When the intake of the experimental subjects decreased to.36 mg/day the excretion of the vitamin also decreased (P <.1). Supplementation with PN-HC1 caused a significant increase (P <.1) in excretion when the diet was supplemented to either the 1.56 or 5.6 mg levels. Comparison of these results with results obtained in an earlier study (13) with nonusers indicated that the changes observed during depletion and repletion phases were similar (Fig. 3). n both studies an intake of approximately 1.5 mg of vitamin B6 per day restored excretion levels to the range observed before depletion. Consumption of 5.6 mg of vitamin B6 for 1 week resulted in a very large portion of the vitamin being lost in the urine. 4-PA excretion When both control and experimental subjects consumed 2.6 mg of vitamin B6 per day (Table 2) the amount of 4-PA excreted at the end of predepletion phase by the control subjects was higher (P <.1) than the amount excreted by the experimental subjects. The removal of PN-HC1 from the diet caused a significant decrease (P <.1) in the excretion of 4-PA and restoration of the vitamin to the diet caused a significant increase (P <.1) at both the 1.56 and 5.6 mg level of intake (Table 2, Fig. 4). The rate of decrease and increase in these subjects using OC was similar to changes found in a previous study (13) of nonusers Downloaded from ajcn.nutrition.org by guest on February 12, 213

5 DONALD AND BOSS. C,, 4, 4 -a 6, / / P FG. 2. Yield of metabolites excreted in OC,..s... vitamin B6 intake,mg,doy days on di.t (Fig. 4). n both studies, an intake of approximately 1.5 mg of vitamin B6 was enough to restore levels of 4-PA excreted to those found before the depletion phases began. The ingestion of 5.6 mg of PN-HC1 for 1 week caused a very large amount of the metabolite to be excreted (Fig. 4). Discussion Subjects using OC excreted almost twice the amount of tryptophan metabolites compared with nonusers (Fig. 2). These results agree with the findings of others (11). strogen-containing OC alter cortisol metabolism of the adrenal cortex, thus, increasing levels of both protein-bound and free glucocorticoids (19-24). Tryptophan oxygenase, the first enzyme in the tryptophan-niacin pathway, is glucocorticoid inducible (25, 26), thus, estrogen could accelerate the rate of the trypurine. Subjects using OC, - s; subjects not using tophan to niacin pathway. Rose and Mc- Guinty (27) found that cortisol administration to humans increased excretion of XA, KA, 3-OH Kyn, and 3-hydroxy-anthranilic acid after a load dose of 2 g of tryptophan. The net result of tryptophan oxygenase induction would be an increased need for the coenzyme PL-P. Depletion of body stores of vitamin B6 caused an increase in the urinary excretion of Kyn, 3-OH Kyn, KA, and XA after a dose of 2. g of tryptophan. Consumption of.96 mg of vitamin B6 for 8 days caused a decrease in tryptophan metabolites but was not sufficient to decrease excretion to original levels. A vitamin B6 intake of 1.56 mg/day for 9 days was sufficient. With the exception of Kyn, an intake of 5.6 mg of vitamin B6 per day caused further decreases to mean levels that were less than initially found. At an intake of 1.56 mg of vitamin B6 for Downloaded from ajcn.nutrition.org by guest on February 12, 213

6 VTAMN B6 RQURMNT N OC USRS days, the excretion of 3-OH Kyn by all subjects had reached levels equal to or less than the predepletion level. Five of seven subjects had reached this level of excretion for Kyn and KA; and six of seven subjects for XA. By day 66, after a vitamin B6 intake of5.6 mg for 7 days, all subjects had reached predepletion levels for 3-OH Kyn and XA and six of seven subjects for KA. Kyn levels for some unexplained reason, markedly increased with intakes of 5.6 mg/day. Thus, for this metabolite, no subjects were at predepletion levels at completion ofthe study, even though five of seven subjects had achieved predepletion levels at the previous level of intake of 1.56 mg. Mean predepletion levels of urinary tryptophan metabolites were reached in the majority of subjects with a vitamin B6 intake of 1.56 mg/day; and in almost all subjects with an intake of 5.6 mg/day. Using these parameters, the amount of vitamin B6 required by subjects using OC is somewhere between 1.5 and 5. mg/day. These results agree with the findings of Lekiem et al. (11) who reported that while excretion levels of tryptophan metabolites were not returned to predepletion levels by.8 mg of PN-HC1 2. mg was sufficient to do so. Lekiem et a!. (1 1) found that, after a load dose of tryptophan, 3-OH Kyn was excreted in the largest amount by OC users, with lesser amounts of XA and Kyn excreted in the urine. n the present study larger amounts of XA were excreted, followed by smaller amounts of Kyn and 3-OH Kyn. Leklem also calculated that at the peak of deficiency 51% of the administered tryptophan dose was cxcreted as tryptophan metabolites, whereas, in the present study only 8% of the dose was excreted in this way. These differences could be due to the more severe degree of depletion reached in the Leklem study. At the level of vitamin B6 intake used in the present diet, changes in the excretion of tryptophan metabolites were not observed in OC users when they either stopped or started using OC nor did the kind of OC used influence results. Total vitamin B6 excreted in urine from users decreased during the depletion phase and began to increase with an intake of.96 mg of the vitamin per day. Consumption of 1.56 mg/day restored mean excretion levels to those observed during the predepletion phase (Fig. 3). Predepletion levels were reached in six of seven subjects. Addition of even larger amounts of PN-HC1 to the diet caused large amounts of the vitamin to be TABL 2 Mean ± S; urin ary excretion of vitamin B and 4-PA by subjects using () and not using (11) OC OC users Nonusers B. in diet (mg) Days on diet Downloaded from ajcn.nutrition.org by guest on February 12, 213 Vitamin B (im/24 hr) ±.4 ±.4 ±.2 ±.3 ±.3 ±. 1 ±. ±.2 ±.4 ±.4 ±.4 Days on diet PA (7) (pm/24 hr) ±.13 ±.14 ±.12 ±.1 ±.2 ±.8 ±.8 ±.12 ±.42 ±.69 ±.16 ±.41 Nonusers B in diet (mg) Days on diet Vitamin B (flm/24 hr) ±.5 ±.3 ±.2 ±.1 ±.1 ±.1 ±. ±.1 ±.1 ±. ±.2 ±. ±.1 ±.3 4-PA (,um/24 hr) ±.3 ±.2 ±.2 ±.1 ±.1 ±.1 ±.1 ±.1 ±.1 ±.1 ±.1 ±.1 ±.1 ±.1 Reference 13.

7 13 DONALD AND BOSS lost in the urine but even then only six of seven subjects had reached predepletion levels (Fig. 3). An intake of approximately 1.5 mg of vitamin B6 would seem to be enough to meet the vitamin needs of most OC users. This recommendation agrees with the suggested intake found in a previous study with nonusers (Fig. 3) (13). The changes in urinary excretion of 4-PA with vitamin B6 depletion-repletion were similar to changes found with levels of vitamin B6 in urine. Repletion with an intake of.96 mg, followed by 1.56 mg/day increased mean excretion to predepletion levels and consumption of 5.6 mg caused the excretion of very high amounts of this metabolite (Fig. 4). Cl C.3 Four of seven, and seven of seven subjects had excretory levels restored at intakes of 1.56 and 5.6 mg, respectively. Using 4-PA excretion as the parameter, an intake of about 1.5 mg/day would likely be, and certainly less than 5 mg would be sufficient to meet the needs of OC users. These levels are similar to suggested intakes for nonusers (Fig. 4) (13). Brown et al. (1) reported that an intake of.85 mg of pyridoxine was not adequate for either OC users or nonusers but an intake of 2. mg/day caused a larger amount of the metabolite to be excreted than during the predepletion phase. They found that with PN-HC1 supplementation, from V4 to 1/2 of the ingested vitamin was excreted as 4-PA with Downloaded from ajcn.nutrition.org by guest on February 12, B6 6 non-users FG. 3. Mean ± S, vitamin B, excretion in urine. Subjects using OC, u; subjects not using OC,...

8 VTAMN B RQURMNT N OC USRS 131 -C (N 4, V V B6 * > a- users non - users FG. 4. Mean ± S,4-PAexcretion in urine. SubjectsusingOC, -.: subjects not using OC the higher levels occurring later in each repletion phase. n the present study, 34, 32, and 29% was excreted for each of the three depletion phases compared with 23% before depletion began. Based on the various parameters measured, an intake of 1.5 mg of vitamin B6 was sufficient to meet the needs of most of the OC users, and an intake of 5. mg caused large amounts of vitamin B6 and 4-PA to be excreted in the urine. These levels of intake were similar to the suggested intake of 1.5 mg/day for non-oc users (13). n 1975, Leklem et al. (11) reviewed the work reported regarding the vitamin B6 requirements of women using OC. They concluded that evidence did suggest that the use 1.54 of estrogen-containing OC may produce a vitamin B6 deficiency in certain subjects. However, controlled dietary research had not shown clear-cut differences in vitamin B6 requirements by OC users and nonusers when a variety of parameters were used to assess need. The authors suggested that the limited number of subjects studied may have prevented small differences from being statistically significant. The results of the present study (12) support the conclusions of Lekiem et al. (11) that the majority of OC users do not have a higher requirement for vitamin B6 than nonusers. The authors acknowledge the technical assistance of Barbara Lowell-Davis, Carol Williamson, and Lois La- 6 Downloaded from ajcn.nutrition.org by guest on February 12, 213

9 132 DONALD AND BOSS belle. The cooperation of the subjects is greatly appreciated, for without them, this study could not have been done. References. ROS, D. P. xcretion of xanthurenic acid in the urine of women taking progestogen-oestrogen preparations. Nature 21: 196, Rosr., D. P. The influence of oestrogens on tryptophan metabolism in man. Clin. Sci. 31: 265, PRC, J. M., M. J. THORNTON AND L. M. MULLR. Tryptophan metabolism in women using steroid hormones for ovulation control. Am. J. Clin. Nutr. 2: 452, BROWN, R. R., D. P. ROS, J. M. PRC AND H. WOLF. Tryptophan metabolsim as affected by anovulatory agents. Ann. N.Y. Acad. Sci. 166: 44, ALY, H..,. A. DONALD AND M. H. W. SMPsON. Oral contraceptives and vitamin B6 metabolism. Am. J. Clin. Nutr. 24: 297, ROS, D. P., R. STRONG, P. W. Ar4u.is AND P.. HARDNG. xperimental vitamin B, deficiency and the effect ofoestrogen-containing oral contraceptives on tryptophan metabolism and vitamin B, requirements. Cliii. Sci. 42: 465, LUHBY, A. L., M. BRN, M. GORDON, P. DAVS, M. MURPHY AND H. SPGL. Vitamin B, metabolism in users of oral contraceptive agents.. Abnormal urinary xanthurenic acid excretion and its correction by pyridoxine. Am. J. Chin. Nutr. 24: 684, ROS, D. P., AND P. W. ADAMS. Oral contraceptives and tryptophan metabolism: effects of oestrogen in low dose combined with a progestogen and of a lowdose progestogen (megestrol acetate) given alone. J. Chin. Pathol. 25: 252, LUMNG, L., R.. CLARY AND T. K. L. ffect of oral contraceptives on the plasma concentration pf pyridoxal phosphate. Am. J. Cliii. Nutr. 27: 326, BROWN, R. R., D. P. ROS, J.. LKLM, H. LNK- SWLR AND R. ANAND. Urinary 4-pyridoxic acid, plasma pyridoxal phosphate, and erythrocyte amino transferase levels in oral contraceptive users receiving controlled intake of vitamin B,. Am. J. Chin. Nutr. 28: 1, LKLM, J.., R. R. BROWN, D. P. Ros, H. LNK- SWLR AND R. A. ARND. Metabolism of tryptophan and niacin in oral contraceptive users receiving controlled intakes of vitamin B,. Am. J. Clin. Nutr. 28: 146, Boss, T. R., AND. A. DONALD. The vitamin B, requirement in oral contraceptive users.. Assessment by pyridoxal level and transferase activity in erythrocytes. Am. J. Chin. Nutr. 32: 115, DONALD,. A., L. D. MCBAN, M. H. W. SMPsoN, M. F. SUN AND H.. ALY. Vitamin B, requirement of young adult women. Am. J. Clin. Nutr. 24: 128, NATONAL RSARCH COUNCL. Recommended Dietary Allowances. (rev. ed. 7). Washington, D.C.: National Academy ofsciences, 1968, publ. no OSR, B. L. Hawk s Physiological Chemistry (14th ed). New York: McGraw Hill Co PRC, J. M., R. R. BROWN AND N. YSS. Testing the functional capacity of the tryptophan-niacin pathway in man by analysis of urinary metabohites. n: Advances in Metabolic Disorders, edited by R. Levine and R. Luft. New York: Academic Press, 1965, vol. 2, p STORVCK, C. A.,. BNSN, M. DWARDS AND M. WOODRNG. Chemical and microbiological determination of vitamin B,. Meth. Bioch. Anal. 12: 226, FUJTA, A., D. FUJTA AND K. FRJNO. Flurometric determination of vitamin B,.. Fractional determunation of pyridoxal and 4-pyridoxic acid. 3. Vitmthol. 1: 279, SANDBRG, A. V. AND W. R. SLAUNWHT, JR. Transcortin: a corticoid binding protein of plasma.. Levels in various conditions and the effects of estrogens. J. Chin. nvest. 38: 129, KLLR, N.,. U. RCHARDSON AND F.. YATS. Protein binding and the biological activity of corticosteroids: in vivo induction of hepatic and pancreatic alanine aminotransferase by corticosteroids in normal and estrogen-treated rats. ndocrinology 84: 49, BRGGS, M., AND M. BROGS. Plasma hormone concentrations in women using steroid contraceptives. J. Obstet. Gynecol. Brit. Common. 79: 946, LUCS,. J., AND R. LUCS. Oral contraceptives and endocrine changes. Bull. World Health Organ. 46: 443, SANDBRG, A. A., H.. ROSNTHAL AND W. R. SLAUNWHT, JR. Certain metabolic effects of estrogens. n: Metabolic ffects of Gonadal Hormones and Contraceptive Steroids, edited by H. A. Sathanick, D. M. Kipnes, and R. L. Van der Wide. New York: Plenum Press, LNDHOHM, J., AND N. SCHULTZ-M#{246}LLR. Plasma and urinary cortisol in pregnancy and during estrogen-gestagen treatment. Scand. J. Chin. Lab. nvest. 31: 119, BRADMAN,. P., AND D. P. ROS. ffects of sex hormones on three glucocorticoid-inducible enzymes concerned with amino acid metabolism in rat liver. ndocrinology 89: 125, ALTMAN, K., AND. GRNGARD. Correlation of kynuremne excretion with liver tryptophan pyrrolase levels in disease and after hydrocortisone induction. J. Clin. nvest. 45: 1527, ROS, D. P., AND F. M. MCGUNTY. The influence of adrenocorticol hormones and vitamins upon tryptophan metabolism in man. Cliii. Sci. 35: 1, Downloaded from ajcn.nutrition.org by guest on February 12, 213