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The content of indigo precursors in Isatis tinctoria leaves--a comparative study of selected accessions and post-harvest treatments.

We recently clarified the nature of indigo precursors in woad (Isatis tinctoria L.), by identifying the major indoxyl glycoside as isatan A (indoxyl-3-O-(6'-O-malonyl-beta-D-ribohexo-3-ulopyranoside)), and by correcting the structure of the related isatan B (indoxyl-3-O-beta-D-ribohexo-3-ulopyranoside). A quantitative densitometric assay for isatans A and B, and indican, was established and validated. HPTLC separation on silica gel was followed by densitometric analysis of indigoid pigments formed after treatment with dilute acid or base. The seasonal variation of indoxyl glycosides in woad leaves was investigated with first-year plants (rosette stage) of five defined I. tinctoria L. and one I. indigotica L. accessions. Isatan A content reached up to 7.6% of dry weight in I. tinctoria, and up to 21.8% in I. indigotica. The influence of various post-harvest treatments was studied. High concentrations of isatans A and B were found in freeze-dried leaf samples, whereas the content of indican was lowest. Conventional drying at ambient or 40 degrees C led to complete disappearance of isatans A and B. The concentration of indican, in contrast, was 3- to 5-fold higher in leaf samples submitted to drying at ambient and 40 degrees C, respectively.

Chromatography, High Pressure Liquid↗

Quantitative analysis of indigo and indigo precursors in leaves of Isatis spp. and Polygonum tinctorium.

Analysis of extracts from two woad species (Isatis tinctoria and Isatis indigotica) and Polygonum tinctorium revealed that only one indigo precursor (indican) was present in Polygonum, but two precursors were found in Isatis spp. This was done using high performance liquid chromatography (HPLC), coupled to an evaporative light scattering detector (ELSD). In Isatis spp., the indigo precursors indican and a fraction representing isatan B were identified. The proportion of indican and isatan B was different between the two Isatis spp. tested. For the first time, it was possible to quantify the precursors in woad plant species, and the results were found to be in good agreement with those made from total indigo quantification using two different spectrophotometric methods or a derivatization technique.

Chromatography, High Pressure Liquid↗

Hepatic detoxification and hepatic function in chronic active hepatitis with and without cirrhosis.

The detoxification capacity of the liver in chronic active hepatitis (CAH) without liver cirrhosis (LC) is not sufficiently known. Therefore, we examined, in 156 patients with morphologically proven CAH of different stages, plasma ammonia, free phenols, indican, glucuronic acid and urea synthesis rate as parameters for liver detoxification. We found a significant increase of ammonia, phenols, and indican and a significant decrease of glucuronic acid and urea synthesis rate parallel to the stage of CAH without LC. In 34 CAH patients with complete recovery, a retrospective 10-year follow-up was possible. Parallel to the normalization of liver morphology and general liver tests, detoxification parameters also normalized. However, the detoxification disorders in CAH without LC are mild in nature and do not produce hepatic encephalopathy. Probably, they are caused by a reduced synthesis of the urea-cycle enzymes and of glucuronyltransferase in the liver.

Ammonia↗

The urinary excretion of bacterial amino-acid metabolites by rats fed saccharin in the diet.

Administration of a diet containing 7.5% saccharin to adult male rats for 40 days caused a three- to four-fold increase in the daily excretion of indican and rho-cresol. Indican is formed from indole which is a microbial metabolite of tryptophan, whilst rho-cresol is formed by the gut flora from tyrosine. The excretion of phenol, which is also a microbial metabolite of tyrosine, was abolished by saccharin administration for 40 days. Analysis of urines collected at 13, 18 and 24 months during a two-generation cancer bioassay showed that these changes occur throughout the life of saccharin-treated rats. These data indicate that saccharin changes the metabolism of amino acids by the gut flora, leading to an increased formation of products known to have promoting or co-carcinogenic properties.

Amino Acids↗

Purple urine bags.

Purple urine drainage bags were found in 7 of 71 chronically catheterized elderly women. The purple staining of the bags is due to a violet discoloration (indirubin) of the plastic of the catheter bag and fine blue crystals of indigo in the urine. The colors are formed from the substrate indoxyl sulfate (indican) and all 7 patients had bacteria in the urine that would produce blue colonies on agar enriched with the urine (filter sterilized) of the patients involved. Organisms identified were Providencia or Klebsiella species. Indican excretion was higher in patients with purple urinary catheter bags than in controls.

Aged↗

Microbial amino acid metabolites and bladder cancer: no evidence of promoting activity in man.

1. Indole, p-cresol and phenol are microbial amino acid metabolites which show co-carcinogenic or promoting activity in animal studies. Their involvement in the development of human bladder cancer has been determined by measuring the urinary excretion of indican (indoxyl sulphate) and conjugated phenols. 2. Thirty-two patients (22 males, 10 females) with histologically confirmed carcinoma of the urinary bladder and a similar number of age and sex matched controls took part in the study. The excretion of indican, p-cresol and phenol showed wide interindividual variability, but did not differ significantly between the two groups. 3. The findings indicate that these endogenous metabolites do not contribute significantly to the development of human bladder cancer.

Aged↗

Liquid-chromatographic study of fluorescent compounds in hemodialysate solutions.

We have separated and identified three endogenous, naturally fluorescent substances in uremic hemodialysate by using reversed-phase liquid chromatography with fluorescence detection. Co-chromatography with authentic standards, monitoring peak shifts after enzymic treatment, and spectrofluorescence measurements were used to confirm the identity of indican, tryptophan, and indole-3-acetic acid. Concentrations of indican were about 1.5 those of tryptophan and considerably greater than those of indole-3-acetic acid in hemodialysate samples from 12 renal patients. These three compounds, as well as eight unidentified components, were consistently present in dialysate samples from each of the 12 patients.

Chromatography, High Pressure Liquid↗

The role of protein metabolism in 204 liver cirrhotics with and without hepatic encephalopathy. II. Amino acids, free phenols and indoles.

Toxic protein metabolites are assumed to play an important role in the multifactorial pathogenesis of hepatic encephalopathy (HE). To investigate this, we examined the serum levels of free amino acids, free phenols and indoles in 100 healthy adults, and in 124 liver cirrhotics with HE and 80 without HE. We found a significant increase in free serum phenols and indican already in liver cirrhosis without portal hypertension (PH) and HE. In stage III and IV HE large amounts of p-hydroxy-phenyl lactic acid were detected, which was not the case in cirrhotics without HE. In HE the increase in free serum phenols and indican was much higher than that of the mother substances tyrosine and tryptophan. The quotient BCAA/AAA was decreased significantly already in PH without HE. In addition to the increased formation by intestinal bacteria, a diminished oxidative capacity of the cirrhotic liver seems to be one of the main causes of the increased serum levels of toxic protein metabolites in HE.

Adult↗

Biochemical activity of intestinal microflora in adult coeliac disease.

In order to answer up to now open questions regarding the role of the microflora in the pathophysiology of adult coeliac disease the excretion of bacterial metabolites in urine has been followed. Unusual high outputs of p-cresol and/or phenol were found in almost all patient, whereas an increased excretion of indican could be observed in a few persons, only. The response to short-term antibacterial therapy was variable.

Adolescent↗

Efficacy of probiotics and nutrients in functional gastrointestinal disorders: a preliminary clinical trial.

The purpose of this study was to evaluate the efficacy and safety of commonly used probiotics and nutrients available for functional gastrointestinal disorders (FGID). Five different combinations of probiotics and nutrients, or placebo, were taken daily over 12 weeks. In this randomized controlled clinical trial, men and women 21 to 72 years of age with FGID symptoms of indigestion, bloating, and abdominal discomfort were assigned to one of six groups, 12 patients per group. Gastrointestinal Quality of Life Index (GIQLI) and visual analogue scale (VAS) for GI symptoms, SF-36, lactulose and mannitol test (LMT), and urine indican levels were evaluated. GIQLI, VAS scores, and LMT did not change significantly (P > 0.05). There were clinically notable improvements in two of the combination formulations. While the nonsignificant improvements in GI symptoms could suggest that combination probiotics and nutrients may be beneficial in conditions such as FGID, no conclusive evidence was found in this pilot trial. Further investigations to explore the findings are discussed.

Abdominal Pain↗

Deficient intestinal absorption of L-tryptophan in progressive myoclonus epilepsy without Lafora bodies.

L-Tryptophan was administered orally to 7 patients with progressive myoclonus epilepsy (PME) without Lafora bodies and 7 controls. Total and free plasma tryptophan wre subsequently measured at intervals. Concentration of free tryptophan was equal in the two groups. Total tryptophan was significantly lower in PME patients and the difference became marked at 120 and 240 min after the oral dose. At the same time urinary excretion of indican rose in PME patients but not in controls but the change was not statistically significant. Urinary 5-HIAA excretion was a little lower in PME patients. The increase of 5-HIAA after administration of L-tryptophan was equal in both groups. Intravenous administration of L-tryptophan produced an equal increase of plasma concentrations in PME patients as in controls, measured at 2-h intervals. These findings suggest incomplete absorption of L-tryptophan through the intestinal tract in PME patients.

Epilepsies, Myoclonic↗

Blue collection bag after ileal diversion.

Five children with ileal diversions have shown asymptomatic blue staining of the urine collection bags. A tryptophan derivative (indican) in the urine that oxidizes to indigo blue on exposure to air is thought to be the cause of this benign transient phenomenon.

Adolescent↗

The clearance of protein-bound solutes by hemofiltration and hemodiafiltration.

BACKGROUND: Hemofiltration in the form of continuous venovenous hemofiltration (CVVH) is increasingly used to treat acute renal failure. Compared to hemodialysis, hemofiltration provides high clearances for large solutes but its effect on protein-bound solutes has been largely ignored. METHODS: Standard clinical systems were used to remove test solutes from a reservoir containing artificial plasma. Clearances of the protein-bound solutes phenol red (C(PR)) and indican (C(IN)) were compared to clearances of urea (C(UREA)) during hemofiltration and hemodiafiltration. A mathematical model was developed to predict clearances from values for plasma flow Q(p), dialysate flow Q(d), ultrafiltration rate Q(f), filter size and the extent of solute binding to albumin. RESULTS: When hemofiltration was performed with Q(p) 150 mL/min and Q(f) 17 mL/min, clearance values were C(PR) 1.0 +/- 0.1 mL/min; C(IN) 3.7 +/- 0.5 mL/min; and C(UREA) 14 +/- 1 mL/min. The clearance of the protein-bound solutes was approximately equal to the solute-free fraction multiplied by the ultrafiltration rate corrected for the effect of predilution. Addition of Q(d) 42 mL/min to provide HDF while Q(p) remained 150 mL/min resulted in proportional increases in the clearance of protein-bound solutes and urea. In contrast, the clearance of protein-bound solutes relative to urea increased when hemodiafiltration was performed using a larger filter and increasing Q(d) to 300 mL/min while Q(p) was lowered to 50 mL/min. The pattern of observed results was accurately predicted by mathematical modeling. CONCLUSION: In vitro measurements and mathematical modeling indicate that CVVH provides very limited clearance of protein-bound solutes. Continuous venous hemodiafiltration (CVVHDF) increases the clearance of protein-bound solutes relative to urea only when dialysate flow rate and filter size are increased above values now commonly employed.

Acute Kidney Injury↗

Findings in routine laboratory examination in progressive myoclonus epilepsy.

Thirty-one patients suffering from progressive myoclonus epilepsy, (also called Unverricht-Lundborg's disease) without Lafora bodies, were examined to check the findings reported in literature and to chart out the main abnormalities in routine laboratory findings. Many alterations could be pointed out, but a high proportion of them were due to factors which are secondary to the syndrome of progressive myoclonus epilepsy: continuous anticonvulsive medication; immobilization; frequent infections; and the patient's poor nutritional condition. The most remarkable finding, and the only one which supported the abnormality reported earlier, was the raised excretion of indican which could not be explained by fermentation in the bowels. The urinary 5-hydroxyindoleacetic acid excretion was a little low, but still within the normal range. The tryptophan and 5-hydroxytryptamine metabolism deserves further investigation in attempting to discover the aetiology of progressive myoclonus epilepsy.

Adolescent↗

Plasma protein binding of frusemide in renal failure rabbits: investigation of endogenous protein binding inhibitors.

The reduction mechanism of frusemide-protein binding in the plasma of renal failure was investigated. The drug-albumin binding was inhibited by the low molecular-weight fraction obtained from acute renal failure rabbits, suggesting the presence of the inhibitors in the plasma. Further, this fraction was divided into six subfractions by Bio-Gel P-2. Fractions II and V2 showed significant inhibition of the protein binding of frusemide. Among uraemic toxins, four indole derivatives markedly inhibited the protein binding. Analysis by hplc confirmed that the concentration of indican was markedly increased in acute renal failure rabbit plasma. It is suggested that this compound could be one of the major inducers of the protein binding defect.

Acute Kidney Injury↗

Circadian periodicity of tryptophan metabolism.

Rhythmicity of tryptophan metabolism via the kynurenine pathway has been demonstrated in man. Normal subjects given 3 g of tryptophan at 0900 hours excreted almost three times the quantity of kynurenine, kynurenic acid, and xanthurenic acid than did subjects given the same dose at 2100 hours. Other metabolites of the kynurenine pathway varied in the same fashion but with lesser magnitude. In contrast, indican, a tryptophan metabolite not in the kynurenine pathway, varied inversely with the other metabolites measured. The data suggest that the liver enzyme tryptophan pyrrolase has a circadian rhythm in man similar to that already described in mice in a previous study. Tryptophan tolerance tests in the future should be controlled relative to time of amino acid administration.

Adult↗

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.

Adult↗

Intestinal epithelial function and villus surface area in rats with bile fistulae.

We have previously found reduced absorption of vitamin B12 in rats with choledochocolic fistulae. To investigate whether the reduction is caused by epithelial dysfunction or mucosal hypoplasia, choledochocolic fistulae were made in 11 rats, whereas 10 rats were sham-operated. The epithelial function was evaluated 9 days later by measuring the uptake of 57CoB12 and glucose in perfused intestinal segments and by determining the activities of 11 mucosal enzymes. Hypoplasia was investigated by performing morphometric measurements of the villus surface area and by measuring the weight, protein, and DNA in mucosal scrapings. The results suggest that choledochocolic fistulae in rats do not impair the epithelial function or cause mucosal hypoplasia. The urinary excretion of indican was increased in the fistula-operated rats, but further studies are needed to establish the significance of this observation.

Animals↗