Tryptophan Load Test Research Articles

Pyridoxine supplementation corrects vitamin B6 deficiency but does not improve inflammation in patients with rheumatoid arthritis

Patients with rheumatoid arthritis have subnormal vitamin B6 status, both quantitatively and functionally. Abnormal vitamin B6 status in rheumatoid arthritis has been associated with spontaneous tumor necrosis factor (TNF)-α production and markers of inflammation, including C-reactive protein and erythrocyte sedimentation rate. Impaired vitamin B6 status could be a result of inflammation, and these patients may have higher demand for vitamin B6. The aim of this study was to determine if daily supplementation with 50 mg of pyridoxine for 30 days can correct the static and/or the functional abnormalities of vitamin B6 status seen in patients with rheumatoid arthritis, and further investigate if pyridoxine supplementation has any effects on the pro-inflammatory cytokine TNF-α or IL-6 production of arthritis. This was a double-blinded, placebo-controlled study involving patients with rheumatoid arthritis with plasma pyridoxal 5'-phosphate below the 25th percentile of the Framingham Heart Cohort Study. Vitamin B6 status was assessed via plasma and erythrocyte pyridoxal 5'-phosphate concentrations, the erythrocyte aspartate aminotransferase activity coefficient (αEAST), net homocysteine increase in response to a methionine load test (ΔtHcy), and 24 h urinary xanthurenic acid (XA) excretion in response to a tryptophan load test. Urinary 4-pyridoxic acid (4-PA) was measured to examine the impact of pyridoxine treatment on vitamin B6 excretion in these patients. Pro-inflammatory cytokine (TNF-α and IL-6) production, C-reactive protein levels and the erythrocyte sedimentation rate before and after supplementation were also examined. Pyridoxine supplementation significantly improved plasma and erythrocyte pyridoxal 5'-phosphate concentrations, erythrocyte αEAST, urinary 4-PA, and XA excretion. These improvements were apparent regardless of baseline B6 levels. Pyridoxine supplementation also showed a trend (p < 0.09) towards a reduction in post-methionine load ΔtHcy. Supplementation did not affect pro-inflammatory cytokine production. Although pyridoxine supplementation did not suppress pro-inflammatory cytokine production in patients with rheumatoid arthritis, the suboptimal vitamin B6 status seen in rheumatoid arthritis can be corrected by 50 mg pyridoxine supplementation for 30 days. Data from the present study suggest that patients with rheumatoid arthritis may have higher requirements for vitamin B6 than those in a normal healthy population.

Plasma Pyridoxal 5′-Phosphate Concentration Is Correlated with Functional Vitamin B-6 Indices in Patients with Rheumatoid Arthritis and Marginal Vitamin B-6 Status

Many patients with rheumatoid arthritis (RA) have low plasma pyridoxal-phosphate (PLP) but a normal erythrocyte aspartate aminotransferase activity coefficient (alpha EAST), a measure of vitamin B-6 status in the erythrocytes, compared with healthy subjects. The goal of the present study was to examine the correlations of PLP levels in these two compartments (plasma and erythrocytes) with other established indices of vitamin B-6 status, and to determine which indicator better reflects functional status of vitamin B-6 in patients with RA. Multiple indices of vitamin B-6 status were measured in 33 patients with RA. Plasma PLP, urinary 4-pyridoxic acid (4-PA), net increase in plasma total homocysteine after a methionine load (DeltatHcy) and net increase in urinary xanthurenic acid after a tryptophan load (DeltaXA) were log-transformed to reach normality for statistical analyses. We found that log-plasma PLP levels were inversely correlated with both log-DeltatHcy (r = -0.368, P = 0.035) and log-DeltaXA (r = -0.333, P = 0.05). Plasma PLP was not correlated with alpha EAST or urinary 4-PA excretion. In contrast, erythrocyte PLP was inversely correlated with alpha EAST (r = -0.431, P = 0.012) and positively correlated with log-4-PA (r = 0.475, P = 0.005), but erythrocyte PLP was not correlated with the outcomes of a methionine or tryptophan load test. Erythrocyte PLP and log-4-PA, but not plasma PLP, were correlated with dietary intake of vitamin B-6 after adjusting for protein intake (r = 0.420, P = 0.015 and r = 0.333, P = 0.05, respectively). We suggest that in patients with RA, plasma PLP levels are a better diagnostic indicator of functional vitamin B-6 status than erythrocyte PLP levels.

Zum einfluss von weizenkleie‐ und grünmehlzulagen auf den vitamin‐B6‐ stoffwechsel von sauen mit suboptimaler vitamin‐B6‐versorgung

Ziel der vorliegenden Untersuchungen war, den Einfluß von “natürlichen Ballaststoffzulagen”; wie Weizenkleie und Grümehl bei suboptimaler Vitamin‐B6‐Versorgung auf Kriterien des Vitamin‐B6‐Stoffwechsels von ausgewachsenen Sauen zu prüfen. Dazu wurden zwei Versuche mit 2 Durchgängen an jeweils 12 ausgewachsenen Sauen (180 kg Lebendmasse) in 3 Gruppen durchgeführt. Die Zulagen betru‐gen im ersten Versuch je Tier und Tag 0 g, 225g und 675 g Weizenkleie und im zweiten Versuch 0 g, 575 g und 1150 g Luzernegrünmehl zu einem Vitamin‐B6‐armen Alleinfutter. Als Kriterien dienten die Vitamin‐B6‐Konzentration und ‐Ausscheidung in Kot und Harn, die Vitamin‐B0‐Bilanz (= Zufuhr ‐ Ausscheidung), die Vitamin‐B6‐Konzentration im Blut, hämatologische Parameter, die Aktivität der erythrozytären Aspartat‐Aminotransferase (EAST) und die Xanthurensäureausscheidung nach Trypto‐phanbelastung. Die Vitamin‐B6‐Konzentration im Kot lag bei 10–12 ug/g DM und wurde weder durch die Art noch durch die Höhe der Ballaststoffzulagen beeinflußt. Die gesamte Vitamin‐B6‐Ausscheidung dagegen wurde durch jede Zulagestufe gesteigert, wobei 60–70% des ausgeschiedenen Vitamin B6 im Kot erschienen. Die fäkale Vitamin‐B6‐Ausscheidung wurde durch die höhere Ballaststoffzulage linear gesteigert. Die bakteriell fermentierbare Substanz aus Weizenkleie und Grunmehl verursachte eine höhere bakterielle Vitamin‐B6‐Synthese als reine Cellulose.

Dietary Protein Quality Alters Ornithine Decarboxylase Activity but Not Vitamin B-6 Nutritional Status in Rats1

Weanling male rats were fed diets that varied in protein quality (casein or wheat gluten) and vitamin B-6 (0.0, 0.5, 1.0 and 1.5 mg pyridoxine HCl/kg diet) to test the hypotheses that low protein quality would depress vitamin B-6 nutritional status and that activity of ornithine decarboxylase (ODC) would be a sensitive functional indicator of vitamin B-6 nutritional status. The wheat gluten diet depressed body weight gain -17% at higher vitamin B-6 levels, as expected. However, vitamin B-6 nutritional status was not worse in gluten-fed compared with casein-fed groups, as evidenced by static measures (B-6 vitamer concentrations in plasma and tissues) and a functional indicator (tryptophan load test). The activity of ODC (holo- and total) in liver, kidney and small intestine did not vary significantly at the three higher levels of vitamin B-6 intake. In groups fed casein, total ODC activity in these tissues was two- to fivefold higher in rats fed diets containing 0.0 mg vitamin B-6/kg compared with higher B-6 levels, without corresponding differences in ODC mRNA abundance in liver and kidney. Concentrations of B-6 vitamers (except pyridoxal phosphate in plasma) increased linearly with dietary vitamin B-6 in plasma, liver, kidney and intestine. These data suggest that low quality protein fed as wheat gluten suppresses growth but not vitamin B-6 nutritional status, and that ODC activity is not a sensitive functional indicator of marginal vitamin B-6 status.

Insensitivity of the Tryptophan-Load Test to Marginal Vitamin B-6 Intake in Rats

The tryptophan-load test for vitamin B-6 nutritional status was administered to adult female Long-Evans rats fed graded levels of pyridoxine hydrochloride (PN.HCl) in two experiments, and its sensitivity to marginal vitamin B-6 intake was evaluated. In Experiment 1, rats were 4-h meal-fed an AIN-76A (20% casein) diet devoid of PN.HCl for 3 wk, then repleted (n = 12) for 6 wk with 4-h pair-fed meals of either 0.25, 0.5, 1.0 or 7.0 (control) mg PN.HCl/kg diet. In Experiment 2, rats (n = 16) were pair-fed for 10 wk either 0.0, 0.5, 1.0 or 7.0 (control) mg PN.HCl/kg diet, with 24-h access to food. Vitamin B-6 nutritional status was assessed at the end of each experiment. Except in rats fed 0 mg PN.HCl/kg diet, mean body weights were not significantly different among diet groups of either experiment. Plasma pyridoxal phosphate (PLP), pyridoxal and total vitamin B-6 concentrations, determined by HPLC, were very sensitive to gradations in dietary PN.HCl concentrations (P less than 0.05). Red blood cell endogenous and PLP-stimulated alanine and aspartate aminotransferase activity did not statistically differentiate all levels of dietary vitamin B-6, although the calculated activity coefficient for each enzyme (stimulated/endogenous activity) did. Urinary xanthurenic acid excretion following a tryptophan load [24.5 mumol (5 mg) L-tryptophan/100 g body weight, injected intraperitoneally] was significantly (P less than 0.05) elevated compared with controls only in the group fed 0 mg PN/HCl/kg diet. At the tryptophan dose used here, the tryptophan-load test was not useful in detecting marginal vitamin B-6 intake in rats.

Evaluation of Vitamin B-6 Status and Function of Rats Fed Excess Pyridoxine

We compared the vitamin B-6 status of 12-wk-old rats (n = 12) fed excess (1400 mg/kg diet) or the recommended level (7 mg/kg diet, control) of pyridoxine (PN) hydrochloride to test if excess vitamin B-6 would cause tissue depletion of pyridoxal phosphate (PLP), the active coenzyme form of vitamin B-6. Plasma PLP, tryptophan-load test results, food intake, and tissue and body weights were not different at wk 6. Red blood cell endogenous alanine aminotransferase activity and PLP concentration were elevated (P less than 0.01) in rats fed 1400 mg PN.HCl/kg diet. In contrast, PLP concentration in muscle was significantly lower (P = 0.01) in rats fed excess vitamin B-6 (9.7 +/- 0.8 nmol/g, mean +/- SEM) than in controls (14.9 +/- 1.4). PLP concentration in other tissues, including plasma, was not affected. In rats fed excess vitamin B-6, pyridoxal was increased in all tissues examined (P less than 0.05), and total vitamin B-6 was increased in plasma, red blood cells and kidneys (P less than 0.05). Total glycogen phosphorylase (a + b) activity in the gastrocnemius was not affected, but phosphorylase a activity was increased in rats fed excess vitamin B-6 (P = 0.025). Concentrations of dopamine and metabolites in the caudate nucleus of the basal ganglia were not affected. A transient, but significant, elevation in acoustic startle response, a central nervous system reflex, was observed in rats fed excess vitamin B-6. The depletion in muscle PLP could not hae been predicted by either plasma or red blood cell PLP concentration, although the latter did reflect vitamin B-6 intake.(ABSTRACT TRUNCATED AT 250 WORDS)

Separation of tryptophan metabolites by reversed-phase high-performance liquid chromatography with amperometric and fluorescence detection

An isocratic method for the separation of most of the tryptophan metabolites in the oxidative degradation pathway is described. The chromatographic analysis utilizes the combined selectivity and sensitivity of amperometric and fluorimetric detection. The effect of pH, ionic strength and operating potential on retention times and detector responses are evaluated. The use of dual electrochemical cells at two operating potentials together with a programmable fluorescence detector allows for improved selectivity for the detection of metabolites. The sensitivity achieved with this method allows for the detection of the metabolites in biological fluids at the picomole level. The method has been used to monitor serum samples obtained during a tryptophan load test.

Tryptophan loading test in premenstrual syndrome

SummaryAn oral tryptophan loading test (SOmg/kg) was performed in both women with premenstrual syndrome and controls. In controls at all sampling intervals, and the premenstrual syndrome subjects in the follicular and mid-luteal phases, tryptophan loading consistently increased whole blood serotonin levels. By contrast, during the late luteal phase, in subjects with premenstrual syndrome there was a decrease in blood serotonin.

Detrimental effects of repeated administration of the tryptophan-load test for vitamin B-6 status on deficient and pair-fed control rats

The tryptophan (trp)-load test is a commonly used test of vitamin B-6 status, in which urinary xanthurenic acid (XA) is measured following a dose of trp. Repeated trp-loads were administered to twelve week old, female Long-Evans rats and several physiologic variables were compared to those in non-loaded rats. Trp-loaded rats (n=7–8) were fed a purified diet devoid of pyridoxine HCl (DEF), or a control diet (AIN-76A) ad libitum (AL-CON) or pair-fed (PF-CON) to DEF for 17 weeks. Rats were injected i.p. with 5 mg trp/100 g body weight weekly for 11 weeks, then again at week 18. Another three groups (n=9–12) were fed similarly, but were never given a trp-load. XA excretion was elevated (P<0.01) in DEF on day 9, and continued to increase. XA excretion expressed as umoles XA/24 hr never differed significantly between AL- and PF-CON; however, at week 18, the percent of trp dose excreted as XA was significantly higher (P<0.03) in PF- than in AL-CON. Trp-loading reduced weight gain and feed efficiency calculated over 17 wk in DEF and PF-CON by 70–80% (P<0.05), but did not affect AL-CON. The magnitude of the effect was greater in DEF, but the effect was stronger statistically in PF-CON. Mean body weight of PF-CON was significantly lower in trp-loaded than in non-loaded rats after only two trp-loads; this effect in DEF was not significant until week 15. Tactile startle response was significantly depressed in DEF compared to controls in non-loaded rats (P<0.05), but not in trp-loaded rats due to significantly greater variability. Repeated trp-load tests in food-restricted rats may affect growth, feed efficiency and variability of response, and should therefore be used with caution in rat studies requiring assessment of B-6 status at frequent intervals.

Quantitative assessment of motor and sensory function in vitamin B-6 deficient rats

Ataxia, weakness and changes in peripheral sensitivity are consequences of vitamin B-6 deficiency. This experiment was conducted to quantify these effects, using tests commonly employed in toxicology studies. Female Long-Evans rats (mean weight=232g) were blocked by weight into 3 treatment groups (n=12): AL-CON, fed AIN-76A diet ad libitum; DEF, fed the same diet devoid of pyridoxine; and PF-CON, pair-fed to DEF. Motor function was evaluated with hindleg gait analysis, grip-strength, and whole-body pull-up. Sensory function was evaluated using tail-flick and landing foot-spread. Deficiency was detected with a tryptophan-load test on day 9 of a simultaneous experiment with an identical feeding protocol. On day 9, abnormalities were detected and quantified by gait analysis—angle and width of step were affected by diet (P<0.05) although there were no differences in body weights and food intakes at that time. The overall effect of diet on angle and width was highly significant (P<0.001). Food intake of DEF was depressed by week 2; at week 10, body weight and feed efficiencies were significantly different among groups. Except for a transient effect on tail-flick, no other test was affected by diet. Gait analysis may have potential as an index of vitamin B-6 status under these experimental conditions.

Attenuation of acoustic and tactile startle responses of vitamin B-6 deficient rats

Vitamin B-6 deficient rats exhibit changes in behavior, sensory function, and other nervous system abnormalities such as convulsive seizures and motor disturbances. Sensorimotor reactivity was evaluated quantitatively by measuring auditory and tactile startle responses in 12 week old female Long-Evans rats fed a diet devoid of added vitamin B-6 (DEF) or a control diet, either ad lib (AL-CON) or pair-fed to deficient rats (PF-CON). Deficiency was confirmed with a tryptophan-load test administered to a separate group of rats fed simultaneously according to the same protocol. At week 18, body weight and feed efficiency were different among groups ( p<0.001), and were lowest in DEF. Amplitude of response to both acoustic and tactile stimuli was depressed in DEF compared to both control groups, which generally did not differ in response. This effect was seen most dramatically in responses to the acoustic stimulus ( p=0.034), and especially to the first presentation ( p=0.017). Latency to maximum response was not affected by diet. Possible mechanisms for this nervous system abnormality, not previously reported in vitamin B-6 deficiency, are discussed.

Differences in vitamin B6 status indicator responses between young and middle-aged women fed constant diets with two levels of vitamin B6

The responses of adult women to two levels of vitamin B6, was conducted with five young and eight middle-aged subjects. A constant diet (2.3-2.4 mg vitamin B6 per day) was fed for four weeks followed by three weeks of the same diet supplemented with 8.0 mg pyridoxine. Plasma pyridoxal 5'-phosphate (PLP), plasma and urinary vitamin B6, and urinary 4-pyridoxic acid (4-PA) were determined. The older women had significantly lower plasma PLP, plasma and urinary vitamin B6 and slightly higher urinary 4-PA values on normal vitamin B6 intakes. With supplementation, only the difference in urinary total vitamin B6 remained significant. Tryptophan load tests revealed no significant between-group differences in xanthurenic acid or kynurenic acid excretion. These results demonstrate an age-related difference in vitamin B6 status indicators in women under controlled dietary intake of vitamin B6.

Vitamin metabolism and the effects of multivitamin supplementation in oral contraceptive users

The effect of combined oral contraceptives containing 150 ug of levonorgestrel and 30 ug of ethinyl estradiol on vitamin B-1, B-2, B-6, B-12, folates, vitamin A, carotenoids, vitamin E and tryptophan load test was studied in a group of 34 healthy non-lactating women. This was compared to an identical study conducted in another group of 19 subjects who used IUCDs for contraception, and also in two other groups of subjects who were given an additional multivitamin preparation on a daily basis (33 subjects) and on the days when OC was not taken (29 subjects). Assessments were made prior to, and during the third week of the 4th, 7th and the 13th cycles of OC treatment. Urinary xanthurenic acid excretion (XA), plasma vitamin A and the folate content of the sera and red blood cells were significantly increased by OC treatment, although this excessive XA excretion was adequately corrected with 10 mg of daily vitamin B-6 supplementation. The metabolism of the rest of the vitamins was not significantly altered by OC usage. This suggests that a routine use of multivitamin supplement may not be necessary.

Tryptophan metabolism in liver cirrhosis: influence of oral antibiotics on neuropsychiatric symptoms.

Oral tryptophan loading tests were performed on patients with cirrhosis of the liver before and after treatment with an oral antibiotic practically unabsorbed from the gastrointestinal tract. After antibiotic therapy, neuropsychiatric manifestations evoked by the oral administration of tryptophan were reduced in duration by about 50% and in severity as well despite a slightly enhanced elevation of serum total and free tryptophan levels, compared to pre-treatment loading tests. The urinary indican output also showed a significantly greater decrease in the post-treatment test (p less than 0.05) whereas no appreciable difference was observed in urinary 5-hydroxy-indoleacetic acid excretion between the pre- and post-treatment tests. The data suggest that neuropsychiatric manifestations seen following oral administration of tryptophan may be attributed to intestinal tryptophan metabolites rather than to this aromatic amino acid itself and/or its metabolites in the brain.

Glucose tolerance in subclinical pyridoxine deficiency in man

Oral glucose tolerance tests were carried out in 16 clinically normal subjects with pyridoxine deficiency (as adjudged by red cell transaminase activity and the 6h tryptophan load test), and in 16 nondeficient controls. The deficient subjects had fasting normoglycemia with hypoinsulinemia (p < 0.01), and a normal postglucose increment in insulin, but peak blood glucose and incremental glucose area were both significantly lower than in the controls (p < 0.01). Suppression of growth hormone during the glucose tolerance test was similar in both groups. These data are interpreted as indicating an enhanced sensitivity to the hypoglycemic action of insulin in pyridoxine-deficient individuals. Supplementation with pyridoxine in these subjects resulted in a tendency for insulin levels to rise (NS), and a significant increase in growth hormone levels. Impairment of growth hormone reserve may be the basis for the increased insulin sensitivity. Subclinical pyridoxine deficiency of the degree seen in this study is apparently associated with improved, rather than impaired, glucose tolerance.

Effects of oestradiol and vitamin B6 on tryptophan metabolism in the rat: implications for the interpretation of the tryptophan load test for vitamin B6 nutritional status.

1. The effects of the administration of oestradiol and vitamin B6on tryptophan metabolism in the rat have been assessed by measurement of the release of14CO2from [14C]tryptophan, in vivo, in order to determine whether, and to what extent, the abnormalities of tryptophan metabolism that are associated with oestrogen administration can be attributed to drug-induced vitamin B6deficiency or depletion. Two positional isomers of [14C]tryptophan have been used; [ring-2-14C]tryptophan as an index of the activity of tryptophan oxygenase (L-tryptophan: oxygen oxidoreductase (decyclizing),EC1.13.11.11) and [methylene-14C]trytophan as an index of the activity of kynureninase (L-kynurenine hydrolase,EC3.7.1.3).2. The administration of 500 μg oestradiol/kg body-weight led to a reduction in the release of14CO2from both positional isomers of tryptophan, suggesting that the activities of both tryptophan oxygenase and kynureninase are reduced following oestrogen treatment. The kinetics of the release of14CO2from [methylene-14C]tryptophan after the administration of oestradiol were compatible with competitive inhibition of kynureninase by oestradiol or a metabolite.3. The administration of 10 mg pyridoxine hydrochloride/kg body-weight also reduced the production of14CO2from both positional isomers of14C]tryptophan, suggesting some toxicity of such a high dose of the vitamin.4. In animals which had received the supplementary dose of vitamin B6, the administration of oestradiol led to further reduction in the production of14CO2from [ring-2-14C]tryptophan, suggesting a further reduction in the activity of tryptophan oxygenase, and an increase in the production of14CO2from [methylen-14C]tryptophan, but with a delay in the peak of production.5. These results confirm that there is no induction of tryptophan oxygenase by oestradiol, but rather reduced activity of the enzyme after the administration of a relatively high dose of the hormone. They also confirm that the inhibition of kynureninase by oestrogen metabolites that has been reported previously in partially-purified enzyme preparations also occurs in vivo.6. It is suggested that the abnormal results of the tryptophan load test that have been reported in women receiving oestrogens, and which have been interpreted as indicating some extent of drug-induced vitamin B6deficiency, can be accounted for by the inhibition of tryptophan metabolism by oestrogens or their metabolites. Therefore it seems likely that the practice of administering supplements of vitamin B6to women receiving oestrogens may not be appropriate, and indeed may exacerbate the changes in tryptophan metabolism that result from the administration of oestrogens. The tryptophan load test would appear to be unreliable as an index of vitamin B6nutritional status in women receiving oestrogens.

Serotonin metabolism and depression in oral contraceptive users

Serotonin and nicotinic acid ribonucleotide metabolic pathways of tryptophan metabolism were studied before and after tryptophan load test in thirty women using oral contraceptive steroids for a period of 2 to 5 years. Ten of them were suffering from depression. Another ten healthy women participated in this study as a control group. Twenty-four-hour urinary excretion of serotonin, 5-hydroxy indole acetic acid and total 5-hydroxyindoles were estimated as indices of serotonin pathway metabolites, while xanthrenate excretion was determined as an index of tryptophan oxygenase pathway. Plasma cortisol, urinary 17-oxosteroids and 17-hydroxycorticosteroids were determined to assess adrenal cortical function. Urinary creatinine output was assayed to check the adequacy of 24-hr urine collection. The changes induced by oral contraceptive steroids on tryptophan and corticosteroid metabolism were correlated with the associated depression. Changes in serotonin metabolism were demonstrated in the depression group before and after tryptophan load test, while in the non-depression group, these changes were only demonstrated after tryptophan load test. Results indicated that alterations in tryptophan metabolism are usually well compensated in the non-depression group but may accentuate or precipitate the development of depression in susceptible women.

Effects of age on serum tryptophan and urine indican in adults given a tryptophan load test.

An oral load of L-tryptophan (490 mumol/kg) was administered to 25 men and 25 women between the ages of 30 and 80 years. Blood samples were drawn before the load and at 2-h intervals for 6 h after the load. Urine samples were collected for 5 days. Fasting serum tryptophan levels averaged 79.5 mumol/l (+/- 12.9 s.d.). Peak serum tryptophan levels (911-1002 mumol/l) occurred 2 h after the load. Urinary indican excretion on the day of the load averaged 6.4 mumol/24h/kg of body weight (+/- 2.8 s.d.). The concentration of tryptophan in serum, the amount of indican excreted in urine, and the indican:creatinine ratio in urine depended on the age of the subjects. The findings are discussed in relation to previous reports on effects of age on tryptophan pharmacokinetics.

Tryptophan malabsorption in dementia. Improvement in certain cases after tryptophan therapy as indicated by mental behaviour and blood analysis.

Twenty-four senile dementia patients in the age range 65-85 years were subjected to a new sensitive tryptophan loading test: Dopa-Tryptophan loading ("Dop-Try loading") in order to determine whether Try malabsorption occurs in senile dementia and if so how frequent this is. For comparison the same loading test was performed on 23 subjects in the same age grouping but with normal metal function. The differences in Try absorption were statistically analyzed and this revealed that absorption was less effective in the senile dementia group. Four patients with Try malabsorption were treated with 3 G 1-Try + 50 Mg dl-5-hydroxytryptophan (5-HTP) per day orally for 20-30 days and four patients with normal absorption were treated in the same way. The Dop-Try loading test was repeated after this treatment. Two patients showed improved absorption and coincident mental improvement, while one showed decreased absorption and mental deterioration. One died during the course of the treatment. None of the four patients with normal absorption showed mental improvement after the treatment. This pilot study suggests that increase in Try absorption following the Try treatment is a necessary condition for improvement in the mental state. The study is being continued with larger numbers of patients.

Inhibition of kynureninase (L-kynurenine hydrolase, EC 3 . 7. 1 . 3) by oestrone sulphate: an alternative explanation for abnormal results of tryptophan load tests in women receiving oestrogenic steroids.

1. A partial purification of kynureninase (L-kynurenine hydrolase, EC 3 . 7. 1 . 3) from rat liver and a total resolution of the apoenzyme have been achieved. The hypothesis that conjugates of oestrogenic steroids compete with pyridoxal phosphate for the cofactor binding site of the enzyme, and so disturb tryptophan metabolism, leading to apparent vitamin B6 deficiency, has been tested. 2. Kynureninase from rat liver was partially purified, and the cofactor-free apoenzyme was prepared. Oestrone sulphate inhibited the enzyme uncompetitively with respect to pyridoxal phosphate, and competitively with respect to kynurenine, with a mean (+/- SE) inhibitor constant (Ki) of 82 +/- 6 microM. 3. The addition of a saturating concentration of pyridoxal phosphate to unfractionated liver homogenates led to an approximately fivefold increase in kynureninase activity, indicating the presence of a relatively large amount of apo-kynureninase in the tissue. 4. It is suggested that the abnormal results of tryptophan load tests in women receiving oestrogens are the result of inhibition of kynureninase by oestrogen conjugates, and that there is no evidence for oestrogen-induced vitamin B deficiency in such cases.