Effect of different levels of oral pyridoxine supplementation on plasma pyridoxal-5'-phosphate and pyridoxal levels and urinary vitamin B-6 excretion.

Plasma pyridoxal-5'-phosphate and pyridoxal levels increased significantly (p less than 0.05) when single, oral bolus doses of pyridoxine were increased from 10 to 25 and from 25 to 50 mg in nine female volunteers. However, when the dose was increased to 100 mg, plasma pyridoxal-5'-phosphate levels did not differ significantly from those recorded after the 50 mg dose. Within 3 h plasma pyridoxal levels rose with a factor of 3.85 compared with the 50 mg dose but high pyridoxal levels were eliminated from the circulation. Renal clearance of pyridoxal remained a constant, low percentage (less than 2.0%) of each pyridoxine supplement in spite of the observed very high circulating pyridoxal levels. Pyridoxine supplementation is discussed in relation to circulating pyridoxal-5'-phosphate and pyridoxal availability for cellular metabolism.

Determination of urinary 4-pyridoxic acid levels as 4-pyridoxic acid lactone using high-performance liquid chromatography.

A high-performance liquid chromatography method for quantitation of urinary 4-pyridoxic acid excretion is presented. Urine samples were treated with HCl to form 4-pyridoxic acid lactone. Acid-treated urine samples were diluted and analyzed by cation-exchange chromatography involving post-column alkalinization and fluorescence detection. Absence of interfering fluorescent urinary compounds was demonstrated by analyzing urine samples without lactonization and rechromatography of aliquots of 4-pyridoxic acid lactone peaks employing reverse-phase chromatography. The method is suitable for determination of urinary 4-pyridoxic acid excretion in nutritional, metabolic, and pharmacokinetic studies.

Plasma pyridoxal-5-phosphate level as risk index for coronary artery disease.

Preliminary evidence is presented that plasma pyridoxal-5-phosphate levels are considerably and significantly reduced (P less than 0.0001) in patients suffering from myocardial infarction when compared to a healthy control group.

Depressed plasma pyridoxal-5'-phosphate levels in tobacco-smoking men.

Tobacco-smoking men (n = 106) showed considerably lower plasma pyridoxal-5'-phosphate levels (P less than 0.001) than non-smoking controls (n = 143). A previously reported inverse relationship between plasma pyridoxal-5'-phosphate levels and age is confirmed by using new and sensitive high-performance liquid chromatography methodology for pyridoxal-5'-phosphate determinations. Plasma pyridoxal levels were neither lower in tobacco-smoking men nor could any relationship between pyridoxal levels and age be demonstrated. Two major risk factors for coronary artery disease, tobacco-smoking and increasing age, are thus associated with lower plasma pyridoxal-5'-phosphate levels.

Further evidence on the effect of vitamin E on the cholesterol distribution in lipoproteins with special reference to HDL subfractions.

Earlier claims that oral administration of alpha-tocopherol results in significant and considerable increase in the serum total HDL concentration could not be substantiated. The present investigation does, however, show that lipoproteins are affected by alpha-tocopherol in other ways, the main effects being an increase in LDL-cholesterol and also in the HDL3-V cholesterol, an HDL3-subfraction determined by polyacrylamide gel electrophoresis. The established lipoprotein risk indices (LDL/HDL and HDL3/HDL2 ratios) were unfavorably affected as a result of alpha-tocopherol treatment. It is concluded that on the basis of available evidence, there is no reason to believe that alpha-tocopherol has a role to play in the prevention and treatment of atherosclerotic disease.

The direct quantification in whole serum of HDL subfractions.

A method is described for the direct electrophoretic separation in human serum of three HDL subfractions and the quantification of the cholesterol content of these by means of gas-liquid chromatography. Used in conjunction with established procedures for the separation of total HDL, VLDL and LDL on agarose gels (with gas-liquid chromatography quantification of cholesterol in each band) the procedure affords comprehensive information on lipoprotein metabolism in patients. Two technicians can process 10 samples per day, making the method suitable for semi-large scale screening of patients such as may occur in clinical trials or to evaluate in greater detail lipoprotein patterns of patients suspected of having lipoprotein abnormalities on the basis of simpler screening methods. Results obtained by the method correlate well with those obtained by means of ultracentrifugation.

A rapid g.l.c. procedure for the determination of codeine and norcodeine in biological fluids based on micro-phase extraction techniques.

Therapeutic serum concentrations of codeine can be virtually completely extracted and obtained sufficiently concentrated for g.c. measurement without distillation, by acid extraction of the drug from the initial crude organic extracts of alkalinized serum followed by a micro-phase back extraction step. The procedure has simplicity, rapidity and eliminates concentration by distillation and background interference. The method can be readily adapted for the seperate measurements of codeine and norcodeine and it can also be used for similar analysis of other body fluids.

Vitamin B6 nutritional status in asthma: the effect of theophylline therapy on plasma pyridoxal-5'-phosphate and pyridoxal levels.

Plasma pyridoxal-5'-phosphate concentrations were significantly lower (p less than 0.001) in a group of 28 asthmatic women when compared to 33 controls. Plasma pyridoxal levels in the two groups were not different. Theophylline was administered to a group of 17 volunteers and resulted in large reductions in plasma pyridoxal-5'-phosphate levels, while plasma pyridoxal levels and urinary 4-pyridoxic acid excretion were unaffected by theophylline therapy. An in vitro study showed that theophylline did not interfere with the high performance liquid chromatography assay for pyridoxal-5'-phosphate, indicating that theophylline could affect liver metabolism of vitamin B6.

Bismuth toxicity in man II. Review of bismuth blood and urine levels in patients after administration of therapeutic bismuth formulations in relation to the problem of bismuth toxicity in man.

A survey of the leterature on bismuth toxicity in man in relation to blood level data, has revealed the necessity of distinguishing between lipid soluble and water soluble organic complexes of bismuth on the one hand and the simple inorganic salts of bismuth on the other hand. A characteristic feature of the former, illustrated by the water soluble bismuth complex triglycollamate, is the high bismuth levels (due to absorption of the complex as such) and the nephrotoxic properties of the compound in man. Bismuth absorption after administration of the simple inorganic salts of bismuth is postulated to occur in the form of ionic bismuth as such, low bismuth levels being characteristic features of such compounds. Bismuth blood and urine levels obtained from patients after administration of a new anti-ulcer drug (Bicitropeptide) in a well controlled clinical trial are discussed and suggest that that this bismuth containing drug behaves pharmacologically in a manner similar to the inorganic bismuth salts in man, low bismuth blood levels and the absence of toxic side effects being conspicuous features of the drug. Based on these considerations, it is proposed that the pharmacologically active bismuth compounds be divided into four different groups depending on structure, stability and solubility. The question as to what constitutes a "toxic bismuth blood level" can only be discussed in relation to the new proposed sub-division of bismuth compounds and is only meaningful if the term is defined to relate only to ionic bismuth (presumably bound to a large extent to blood proteins). Based on information gleaned from the literature and blood level values reported in the clinical trial referred to, it is suggested that bismuth blood level values below 50 micrograms/ml are highly unlikely to be associated with meaningful toxicity in man. Finally, attention is drawn to the reversibility of bismuth toxicity in man as reported by many authors irrespective of the type of bismuth compound concerned.

A rapid and comprehensive system for the routine identification of drugs in biological material based on microphase extraction and drug colour profiles.

The separation of basic, acidic and neutral drugs from propanol-2 extracts of serum, urine and tissue homogenates at different pH values using a micro-phase extraction technique is described. Following preliminary screening, the various drug-containing fractions obtained are further examined by two-dimensional thin-layer chromatography. The drugs present are identified with reference to documented standards with the aid of a drug colour profile system and RF values relative to three different reference standards. By means of gas chromatographic analysis of the same extracts, semi-quantitative estimates of the amounts of drugs present, which are sufficiently accurate for clinical emergency purposes, can be made in many instances. The main advantages of the system are "clean" extracts with a minimum of background interference, rapidity (4-6 h for a complete analysis) and systematically documented and visually presented behaviour of drugs after spraying with various chromogenic and fluorogenic reagents, allowing the systematic identification of unknown substances.

Monitoring aminopentamide urinary excretion by means of multiple 'microphase' extraction - a rapid method for the extraction and concentration of small amounts of lipophilic drugs from large volumes of biological fluids without distillation.

It has been demonstrated that low concentrations of basic lipophilic drugs in biological fluids may be extracted and concentrated 10(4)-10(6) times by a series of extraction procedures in which the ratio of the extracting solvent to that of the solution to be extracted is of the order of 1:100 (microphase extraction procedure). Typically, basic drugs (atropine, aminopentamide, hyoscine, chloroquine, pyrimethamine) were extracted and concentrated sufficiently for direct GC analysis from 24-hour urine samples by a procedure involving three simple consecutive extraction steps. Using this procedure, it was demonstrated that after administration of aminopentamide (300 micrograms) to patients in the form of anti-diarrhoeal tablets, measurable quantities of the free, unchanged drug can be demonstrated in 24-hour urine samples. The main advantages of the method are simplicity, rapidity and sensitivity due to the low background interference in the GC separations. The principle involved can be extended to the analysis of acidic drugs with suitable solubility properties.

A direct procedure for the micro-scale preparation of acyl derivatives of nucleophilic drugs from carboxylic acids for gc separation and identification.

Free carboxylic acids were converted into mixed anhydrides suitable for the acylation of nucleophilic drugs in a one-step procedure. By preparing more than one acyl derivative and comparing the retention times of the derivatives with those of authentic drug samples with reference to suitable internal standards, the positive identification of unknown drugs was possible. The method is suitable for use in conjunction with the microphase extraction of drugs.

Drug induced biogenesis of nitrosamines.

It has been demonstrated that potentially corcinogenic nitroso compounds are readily formed when certain widely used drugs with secondary alkylaminostructures (e.g. nortriptyline and fenfluramine) or drug metabolities with such structures (e.g. norpropoxyphene) are reacted with nitrite in dilute acid aqueous medium or in human gastric juice under simulated gastric conditions. The reactions occur so readily in vitro that new quantitative procedures for the determination of these drugs have been developed and based on these principles. The potential hazards associated with the long term clinical use of such drugs are discussed.

Bismuth toxicity in man - I. Bismuth blood and urine levels in patients after administration of a bismuth protein complex (Bicitropeptide).

In the course of a double-blind clinical trial of treatment involving a bismuth protein complex (Bicitropeptide), an antacid and placebo, blood samples (33 patients) and urine samples (43 patients) were collected for bismuth analysis from patients on bismuth therapy at the beginning of the trial (week 0) and at weeks 3 and 6. Base line blood (2 - 11 ng/l) and urine (2 - 29 ng/l) values were not zero and appeared to have reached "saturation" values in many cases after 3 weeks (Blood : 4 - 33 ng/l and Urine 60 - 600 ng/l). After 6 weeks, blood values were 5 - 20 ng/l and urine values 63 - 780 ng/l. It is concluded that these blood and urine levels are not associated with significant toxicity in man.

Stability of pyridoxal-5-phosphate semicarbazone: applications in plasma vitamin B6 analysis and population surveys of vitamin B6 nutritional status.

The determination of pyridoxal-5-phosphate (PLP) and pyridoxal (PL) in plasma requires the availability of dark room facilities, due to the photosensitivity of these vitamin B6 vitamers. The fact that the semicarbazone forms of PL and PLP are more strongly fluorescent than the underivatized B6 vitamers has been exploited in plasma analyses, but it was not previously realised that these semicarbazone forms are also very stable even under conditions that lead to rapid decomposition of free PL and PLP. The stabilisation of PLP and PL obtained in this manner is sufficient and fully adequate to meet the practical requirements of clinical field studies. We report a high-performance liquid chromatographic method for plasma PLP and PL determinations based on precolumn semicarbazone formation and fluorescence detection. The method is sensitive enough for quantitative plasma PLP determinations even in B6-deficient patients.