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PubMed1990

Simultaneous liquid chromatographic determination of cholesterol, phytosterols and tocopherols in foods.

A method is described for the simultaneous determination of cholesterol, phytosterols and tocopherols in dairy and non-dairy high-performance liquid chromatography. A facile saponification and extraction scheme is completed rapidly within a single reaction tube. Clean-up is not required, with analysis completed by one of two alternative reversed-phase approaches. Sterols are monitored by short-wave ultraviolet spectrophotometry, and tocopherols by means of a series fluorescence detector. Cholesterol is resolved from the principal plant phytosterols, while the tocopherol congeners are well separated, thereby offering complementary information regarding source-lipid composition in foods under regulatory quality-control conditions. Squalene, an essential metabolic sterol precursor, can also be determined concurrently at the same viewing wavelength.

Animals↗

PubMed1992

High-performance liquid chromatographic and derivative ultraviolet spectrophotometric determination of amoxycillin and dicloxacillin mixtures in capsules.

Two rapid assay procedures based on high-performance liquid chromatography (HPLC) and derivative ultraviolet (UV) spectrophotometry have been developed for the simultaneous determination of amoxycillin and dicloxacillin in two-component capsule formulations. The HPLC determination was carried out on a reversed-phase C8 column with use of a mobile phase consisting of methanol-0.02 mol dm-3 ammonium acetate (pH 5) (50 + 50) at a flow rate of 1.0 cm3 min-1, with UV detection at 230 nm. The fourth-derivative spectrophotometric procedure depends on the measurement of the derivative amplitudes, in 0.1 mol dm-3 NaOH, at 308.5 and 275 nm for amoxycillin and dicloxacillin, respectively. For both procedures, the calibration graphs were linear in the ranges 20-200 and 20-140 micrograms cm-3 for the HPLC and UV derivative methods, respectively, with an almost zero intercept and a correlation coefficient of 0.999. Commercial capsules and laboratory-prepared mixtures containing both penicillins in different proportions were assayed by the developed procedures. The results were of comparable accuracy as indicated by a statistical analysis of the data, using both t- and F-tests.

Amoxicillin↗

PubMed1998

Determination of compatibility and stability of drugs used in palliative care.

BACKGROUND: Drugs for symptom control in the terminal phase of palliative patients may be used in pump systems. OBJECTIVE: To investigate the compatibility and stability of solutions of morphine hydrochloride, haloperidol, metoclopramide hydrochloride, atropine sulphate, butylhyoscine bromide and ranitidine hydrochloride, which may be used together under such circumstances. METHOD: Solutions of the drugs were exposed to ambient light at a temperature of 20 +/- 5 degrees C and +/- 31 degrees C for 24 h and 7 days and the solutions studied by ultraviolet spectrophotometry. RESULTS: The combination of haloperidol or metoclopramide hydrochloride with different concentrations of morphine hydrochloride seemed to be compatible and stable. The concentration of atropine sulphate or butylhyoscine bromide could not be determined by the study method, but morphine hydrochloride seemed to be stable. The combination of ranitidine hydrochloride and morphine hydrochloride showed a change in colour after 7 days. CONCLUSION: The results suggest that the combination of different concentrations of morphine hydrochloride with haloperidol, metoclopramide hydrochloride, atropine sulphate, butylhyoscine bromide or ranitidine hydrochloride do not affect their stability when stored for 7 days up to 30 degrees C under the influence of ambient light. However, more robust stability-indicating methods are required to confirm these results. The proposed method is more useful for identifying combinations that are clearly incompatible than to identify those that are compatible.

Atropine↗

PubMed1996

Toluidine Blue Dye-Binding Method for Measurement of Genomic DNA Extracted From Peripheral Blood Leukocytes.

Background: The toluidine blue DNA assay has characteristics that suggest its suitability for concentration determinations of genomic DNA extracted from peripheral blood leukocytes. These include visible wavelength readings, rapid analytical time, useful sensitivity, and low reagent costs. We validated this assay for genomic DNA and devised a simple method to identify extracts that exceed the linear range of the assay. Methods and Results: DNA was prepared from 109 blood samples by a salt extraction method. The toluidine blue assay was linear for DNA concentrations from 0 to 500 ng/µL. By performing bichromatic absorption readings at 570 and 628 nm, extracts with DNA concentrations exceeding the linear range could be identified up to at least 1300 ng/µL. Analytical precision and accuracy studies gave acceptable results. There was excellent agreement between DNA concentrations determined by the toluidine blue assay and ultraviolet spectrophotometry. RNA was confirmed to be significant positive interferent. The detection limit of genomic DNA was determined to be 15 ng/µL. Conclusions: The toluidine blue DNA assay is suitable for measuring DNA concentrations in extracts obtained from peripheral blood. This assay appears to be suitable for automation as a visible spectrophotometric DNA method.

Journal Article↗

PubMed2002

Antioxidant properties of carboxymethyl glucan: comparative analysis.

Antioxidative capabilities of carboxymethylated (1 --> 3)-beta-D-glucan from Saccharomyces cerevisiae cell wall, alpha-tocopherol, and mannitol against lipid peroxidation in phosphatidylcholine liposomes induced by OH. radicals produced with Fenton's reagent (H(2)O(2)/Fe(2+)) were studied using absorption ultraviolet-visible spectrophotometry. It was found that (1 --> 3)-beta-D-glucan is an antioxidant with the scavenging ability lying between that of alpha-tocopherol, which is known to be incorporated in the lipid bilayer, and the water-soluble antioxidant, mannitol.

Antioxidants↗

PubMed1998

Influence of flow rate on aerosol particle size distributions from pressurized and breath-actuated inhalers.

Particle size distribution of delivered aerosols and the total mass of drug delivered from the inhaler are important determinants of pulmonary deposition and response to inhalation therapy. Inhalation flow rate may vary between patients and from dose to dose. The Andersen Sampler (AS) cascade impactor operated at flow rates of 30 and 55 L/min and the Marple-Miller Impactor (MMI) operated at flow rates of 30, 55, and 80 L/min were used in this study to investigate the influence of airflow rate on the particle size distributions of inhalation products. Total mass of drug delivered from the inhaler, fine particle mass, fine particle fraction, percentage of nonrespirable particles, and amount of formulation retained within the inhaler were determined by ultraviolet spectrophotometry for several commercial bronchodilator products purchased in the marketplace, including a pressurized metered-dose inhaler (pMDI), breath-actuated pressurized inhaler (BAMDI), and three dry powder inhalers (DPIs), two containing salbutamol sulphate and the other containing terbutaline sulphate. Varying the flow rate through the cascade impactor produced no significant change in performance of the pressurized inhalers. Increasing the flow rate produced a greater mass of drug delivered and an increase in respirable particle mass and fraction from all DPIs tested.

Aerosols↗

PubMed2005

Use of the next generation pharmaceutical impactor for particle size distribution measurements of live viral aerosol vaccines.

Aerosol administration of live measles vaccine virus has proven to be extremely efficacious in field trials using an industrial compressor coupled to a disposable nebulizer (IPI). To develop a new system for administration, it is necessary to characterize the operating characteristics of the old system. There are no standardized techniques for measuring particle size of live biological agents. This study evaluated the Next Generation Pharmaceutical Impactor's (NGI) ability to particle size wet aerosols in an effort to measure the particle size distribution of live measles vaccine from the IPI nebulizer. As a control albuterol was aerosolized using a Pari LC Star, since the soluble albuterol is evenly distributed throughout the droplets and laser diffraction measurements should agree with those from the NGI, as long as the NGI is cooled to prevent heat transfer to the aerosol. Albuterol was also used as a control for the IPI using quantitative ultraviolet spectrophotometry. There was close agreement in MMD (mean +/- 95% CI) for the LC Star, measured by laser diffraction (3.24 +/- 0.06 microm) and the NGI (2.93 +/- 0.22 microm) and the IPI (4.26 +/- 0.17 and 4.26 +/- 0.24 microm, respectively). For the measles vaccine assayed for plaque forming units, there were significant differences between the NGI MMD (6.14 +/- 0.39 microm) compared to laser diffraction (4.95 +/- 0.16 microm) indicating that the vaccine is not evenly distributed among the droplets of various sizes. This is likely clumping of the virus due to gelatin in the formulation. These data indicate that the NGI is capable of particle sizing live biological agents.

Administration, Inhalation↗

PubMed2006

In vitro validation of 99mTc-HFA-FP delivered via pMDI-spacer.

The purpose of the study was to label Flixotide (fluticasone propionate [FP] with HFA propellant), with technetium-99m and validate that (99m)Tc acts as a suitable marker for FP when delivered via pMDI-spacer. Sodium pertechnetate was mixed with 5 mL of butanone. (99m)Tc was extracted into butanone and transferred into an empty canister. The (99m)Tc lined canister was heated, and the butanone evaporated to dryness. A supercooled commercial Flixotide canister was decrimped, and the contents transferred to the (99m)Tc lined canister and recrimped. The particle size distribution of FP and (99m)Tc from 10 radiolabeled canisters was measured using an Anderson cascade impactor calibrated to 28.3 L/min, and compared to commercial FP. The drug (FP) content of each particle size fraction was measured using ultraviolet spectrophotometry and the (99m)Tc level in each fraction was measured using an ionization chamber. The percentage of particles in the fine particle fraction (<;4.7 microm) and the percentage of (99m)Tc from commercial and radiolabeled canisters were compared. The mean (SD) % FP in the fine particle fraction, before and after label was 43.2 (1.8) % and 43.9 (2.6) %, respectively. The mean (SD) % (99m)Tc in the fine particle fraction was 42.1 (5.1) %. The mean %FP exiting spacer at (<4.7 microm) before labeling was not significantly different from the mean % FP exiting spacer at (<4.7 microm) after labeling (p > 0.05). The mean % (99m)Tc attached to particles at (<4.7 microm) after radiolabeling was not significantly different from the mean % FP levels (p > 0.05). The validation in this study indicates that (99m)Tc can act as a suitable marker for HFAFP, delivered via pMDI-spacer.

Administration, Inhalation↗

PubMedDate not supplied

A death investigation involving quinidine and propranolol.

A death attributed to quinidine-propranolol intoxication is described. Toxicological analysis of the tissues utilized thin layer (TLC) and gas-liquid (GLC) chromatography, ultraviolet (UV) spectrophotometry and gas chromatography-mass spectrometry (GC-MS). Tissue levels for both drugs, along with their chemical ionization mass spectra, are presented.

Adult↗

PubMedDate not supplied

Formation of 2-(4-chlorophenoxy)-2- methylpropanamide from a clofibrate metabolite during extraction of urine.

Drug isolation from urine by solvent extraction after alkalinization suffers the potential disadvantage of chemically altering desired isolates by hydrolytic reactions. This paper reports a reaction which results in the formation of an amide substance when urine from individuals ingesting clofibrate is treated with ammonium hydroxide or sodium hydroxide and ammonium ion. The amide formed in urine has been compared to synthetically prepared 2-(4-chlorophenoxy)-2-methylpropanamide by gas chromatography, thin layer chromatography, ultraviolet spectrophotometry, infrared spectrophotometry, and mass spectrometry with complete correspondence of chemical properties. Conditions under which this amide, the major clofibrate-related entity, formed in urine are those routinely employed in analytical toxicology.

Chemical Phenomena↗

PubMedDate not supplied

Procainamide depleted tablets not a cause of death.

Sixteen apparently intact procainamide tablets, in varied stages of drug depletion, were found distributed throughout the gastrointestinal tract of a man who died of arteriosclerotic heart disease with pulmonary complications. The tablets were found to contain procainamide by a thin layer chromatography system with cataloged compound identification characteristics. The drug was confirmed by ultraviolet spectrophotometry and gas chromatography/mass spectrometry. The concentration of the drug in the liver was in a therapeutic range and the drug was not attributed as contributing to the cause of death.

Aged↗

PubMed1991

Dietary phosphate deprivation increases renal synthesis and decreases renal catabolism of 1,25-dihydroxycholecalciferol in guinea pigs.

In guinea pigs, dietary phosphate deprivation decreases plasma phosphate concentration, increases plasma 1.25-dihydroxycholecalciferol [1,25-(OH)2D3] concentration and causes hypercalcemia concurrent with the maximal increase in plasma 1,25-(OH)2D3 levels. Our objective was to determine whether increased synthesis or decreased catabolism contributed to the elevation in plasma 1,25-(OH)2D3. Preliminary experiments using renal mitochondria from guinea pigs fed a control diet revealed that 23,25-dihydroxycholecalciferol [23,25-(OH)2D3], not 24,25-dihydroxycholecalciferol [24,25-(OH)2D3], was the reciprocal side-chain metabolite to 1,25-(OH)2D3 in this species. An assay employing guinea pig renal mitochondria was used to measure the renal synthesis of 1,25-(OH)2D3 and 23,25-(OH)2D3 from [3H]25-OH-D3. These metabolites were unequivocally identified by combinations of HPLC, ultraviolet spectrophotometry and mass spectrometry. This renal mitochondrial assay was subsequently used to investigate the effect of dietary phosphate deprivation on guinea pig vitamin D metabolism. Within 1 wk the rate of synthesis of 1,25-(OH)2D3 was maximal in phosphate-deprived guinea pigs. This rate was significantly (P less than 0.005) higher than that achieved in same-day control guinea pigs. Conversely, within 1 d the synthesis of 23,25-(OH)2D3 was significantly (P less than 0.005) decreased in phosphate-deprived guinea pigs. Similarly, the rate of 1,25-(OH)2D3 metabolism was decreased within 1 d of dietary phosphate deprivation and was at a minimum within 1 wk. This rate was significantly (P less than 0.005) less than that attained in same-day control guinea pigs. These results suggest that both increased synthesis and decreased metabolism of 1,25-(OH)2D3 contribute to the plasma 1,25-(OH)2D3 elevation that occurs in response to dietary phosphate deprivation.

Animals↗

PubMed1995

Biochemical characterization of a Ca(2+)-dependent lectin from the hemolymph of a photosymbiotic marine bivalve, Tridacna derasa (Röding).

A Ca(2+)-dependent lectin was purified from the hemolymph of a photosymbiotic bivalve, Tridacna derasa. An electrophoretically homogeneous form was obtained by using affinity chromatography with Sepharose 4B. More than 80% of the hemolymph protein was accounted for by this lectin. The apparent molecular mass of the lectin, in its native form exhibiting hemagglutinating activity, was estimated by gel filtration analysis to be approximately 480 kDa. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis, in the absence of reductants, it migrated as a single band corresponding to a very large size, while in the presence of 2-mercaptoethanol, it migrated as two distinct bands of 23 and 46 kDa. These results indicate that the subunits were linked by disulfide bridges to form the native protein. After reducing pyridylethylation, each of the 23- and 46-kDa polypeptides was isolated by gel filtration in a mobile phase containing guanidine-HCl. The two polypeptides had the same amino-terminal sequence and a similar amino-acid composition, and in the presence of 2-mercaptoethanol gave a single band on isoelectric focusing at a pH of 6.0. The results suggested that the 46-kDa peptide is a homodimer of 23-kDa subunits held together by a covalent bond other than a disulfide linkage. This lectin required calcium ions for its activity. By ultraviolet spectrophotometry the association constant for the calcium ion was determined to be 0.88 mM. The hemagglutinating activity decreased dramatically below pH 6.5, but re-increased to the original level when the solution was neutralized. Such a pH-dependent alteration of the ligand-binding activity was similar to that found in vertebrate asialoglycoprotein receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed2006

A study on eggshell pigmentation: biliverdin in blue-shelled chickens.

Biliverdin is an important pigment in the eggshell of chickens and other avian species. Determination of the biosynthesis site for biliverdin is essential for understanding the biochemical process and genetic basis of eggshell pigmentation. Either blood or the shell gland could be the biosynthesis site of eggshell biliverdin. A segregation population with full-sib sisters genotyped Oo and oo, which laid blue-shelled eggs and light brown eggs, respectively, was constructed in a native Chinese chicken breed. Ultraviolet spectrophotometry and HPLC were used to determine the biliverdin concentration in eggshells, blood, bile, excreta, and shell gland of both groups of chickens. Biliverdin content was significantly different between egg shells of blue-shelled and brown-shelled chickens (P < 0.01). Blood and bile were tested 3 to 4 h before oviposition, and excreta was tested randomly. Results showed no significant difference in biliverdin concentration in blood, bile, and excreta between the 2 groups. In the shell gland, the biliverdin contents for the blue-shelled and brown-shelled chickens were 8.25 +/- 2.55 and 1.29 +/- 0.12 nmol/g, respectively, which showed a significant difference (P < 0.01). Our results demonstrated that blood is not the biosynthesis site of the shell biliverdin. Biliverdin is most likely synthesized in the shell gland and then deposited onto the eggshell of chickens.

Animals↗

PubMed2000

Vitamin A and E supplementation in breast-fed newborns.

BACKGROUND: Vitamins A and E are two potent antioxidant nutrients that play a significant role in immune function. In contrast to the numerous studies of vitamin A and E status in children, adolescents, and adults, information on term infants, particularly breast-fed infants, is scarce. The goals of the present investigation were to examine the vitamins A and E nutritional status of term breast-fed infants at birth and to assess retinol and tocopherol plasma levels during a 3-month supplementation trial. METHODS: The study was a prospective, blinded comparison of a supplementation protocol with a placebo in a group of consecutively recruited term newborns. The supplemented group received 3000 IU vitamin A and 5 IU vitamin E orally. The placebo group received a solution of similar viscosity and organoleptic characteristics. Vitamin A and E were separated by reverse-phase high-performance liquid chromatography on a C18 Spectrasyl column and quantified by ultraviolet spectrophotometry. RESULTS: Vitamin A and E levels steadily increased with age in both groups of infants. However, levels at 3 months were higher in the supplemented than in the control group. CONCLUSION: The data show that supplementation with 3000 IU vitamin A and 5 IU vitamin E for 3 months increases circulating vitamin levels in newborn term babies compared with those in nonsupplemented infants.

Adult↗

PubMed1998

Concurrent high-performance liquid chromatographic measurement of loxapine and amoxapine and of their hydroxylated metabolites in plasma.

The dibenzoxazepine neuroleptic loxapine, its N-demethylated metabolite amoxapine, and their 7- and 8-hydroxymetabolites were measured simultaneously in plasma by reversed-phase high-performance chromatographic method. An original liquid-liquid extraction procedure was performed, consisting in coextraction of the substances together with a water-miscible solvent (acetonitrile) by a non-water-miscible solvent (toluene). The substances were separated on a 5-microm CN 25-cm column, and eluted with a mobile phase consisting of acetonitrile-acetic acid 0.5 N (30:70) and hexylamine (0.05%). They were detected by ultraviolet spectrophotometry at 310 nm. Clozapine was used as internal standard. Linearity was demonstrated in the range of 10 to 250 microg/l, and detection limits were found to be 3.5 to 6.3 microg/l according to the substance. Within-day repeatability ranged from 2.7% to 6.5%, and between-day reproducibility ranged from 0.9% to 20.2%. The extraction procedure provided a mean absolute recovery of 51.1% (range, 40.7% to 58.6%) with a mean coefficient of variation of 4.2%. This technique was applied to the concurrent determination of plasma concentrations of the compounds in 10 patients administered loxapine 75 to 600 mg daily. Steady state plasma levels of loxapine were significantly correlated with oral doses (n = 10, r = 0.858, p < 0.002). In conclusion, the method proved to be a convenient and reproducible procedure allowing the simultaneous measurement of loxapine, amoxapine, and their metabolites in patients.

Amoxapine↗

PubMed2000

A novel mutation in the flavin-containing monooxygenase 3 gene, FM03, that causes fish-odour syndrome: activity of the mutant enzyme assessed by proton NMR spectroscopy.

We have previously shown that primary trimethylaminuria, or fish-odour syndrome, is caused by an inherited defect in the flavin-containing monooxygenase 3 (FMO3) catalysed N-oxidation of the dietary-derived malodorous amine, trimethylamine (TMA). We now report a novel causative mutation for the disorder identified in a young girl diagnosed by proton nuclear magnetic resonance (NMR) spectroscopy of her urine. Sequence analysis of genomic DNA amplified from the patient revealed that she was homozygous for a T to C missense mutation in exon 3 of the FMO3 gene. The mutation changes an ATG triplet, encoding methionine, at codon 82 to an ACG triplet, encoding threonine. A polymerase chain reaction/restriction enzyme-based assay was devised to genotype individuals for the FMO3Thr82 allele. Wild-type and mutant FMO3, heterologously expressed in a baculovirus-insect cell system, were assayed by ultraviolet spectrophotometry and NMR spectroscopy for their ability to catalyse the N-oxidation of TMA. The latter technique has the advantage of enabling the simultaneous, direct and semi-continuous measurement of both of the products, TMA N-oxide and NADP, and of one of the reactants, NADPH. Results obtained from both techniques demonstrate that the Met82Thr mutation abolishes the catalytic activity of the enzyme and thus represents the genetic basis of the disorder in this individual. The combination of NMR spectroscopy with gene sequence and expression technology provides a powerful means of determining genotype-phenotype relationships in trimethylaminuria.

Adult↗

PubMed1976

Rhodanese from Thiobacillus A2: catalysis of reactions of thiosulphate with dihydrolipoate and dihydrolipoamide.

Rhodanese (thiosulphate:cyanide sulphurtransferase EC.2.8.1.1) was purified 25- to 30-fold from thiosulphate-grown Thiobacillus A2. It exhibited a pH optimum between pH 10-2 and 10-4 and apparent Km values of 0-36 mM-Na2S2O3 and 17 mM-KCN. Ultraviolet spectrophotometry and thin-layer chromatography showed that the enzyme catalysed the reaction of S2O3(2-) with dihydrolipoic acid or dihydrolipoamide, producing alpha-lipoate or lipoamide, with the intermediate production of the persulphides of dihydrolipoate and dihydrolipoamide, which were demonstrated chromatographically. This is the first demonstration of catalysis by a thiobacillus rhodanese of reactions which are likely to be physiologically important in the oxidative dissimilation of thiosulphate by a central energy-conserving pathway.

Amides↗