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At least 829 records · Page 46Linked to original sources
PubMed1991

A procedure for the simultaneous determination of lipid and protein in biomembranes and other biological samples.

Very small sample sizes frequently become the limiting factor in biochemical and biomembrane studies in which routine quantification of protein and bulk lipids are required. The procedure described here allows the simultaneous determination of protein and lipid without initial, multiple aliquots. The method is based on the quantitative precipitation of proteins from a defined hexane/isopropanol mixture. The liquid phase resulting after decanting and concentrating to dryness can then be used to assay the lipid content directly. Quantitative assay of protein can be achieved after resuspension of the pelleted material by addition of sodium dodecyl sulfate (0.1%) and deoxycholate (1%). The method is also applicable to other types of lipid- and protein-containing samples with a broad range of protein/lipid ratios and lipid compositions, as they occur, for example, in serum lipoproteins.

1-Propanol↗
PubMed1989

Rapid headspace gas chromatography of hexanal as a measure of lipid peroxidation in biological samples.

A rapid, sensitive and convenient capillary gas chromatographic-headspace method was developed to determine hexanal as an important volatile decomposition product of hydroperoxides formed from n-6 polyunsaturated fatty acids in rat liver samples. Total volatiles were also determined as a measure of overall lipid peroxidation. Samples of headspace taken from sealed serum bottles incubated at 37 degrees C were injected into a gas chromatograph. It was possible to make 15 determinations per hour. This method is convenient because no special sample manipulations are necessary. The addition of 0.5 mM ascorbic acid prior to gas chromatographic analysis significantly increased hexanal production. The applicability of the method was demonstrated in studies of the effect of iron in the presence or absence of hydroperoxides of methyl linoleate and methyl linolenate and tert-butyl hydroperoxide on rat liver homogenates, slices and microsomes. A rapid silica cartridge chromatographic procedure was used to purify hydroperoxides from autoxidized methyl linoleate and methyl linolenate, and hydroperoxy epidioxides (cyclic peroxides) from autoxidized methyl linolenate in 20-40 mg quantities. The hydroperoxides and hydroperoxy epidioxides of methyl linolenate were effective inducers of n-6 polyunsaturated fatty acid peroxidation in liver homogenates. Hexanal and thiobarbituric acid-reacting substances were significantly correlated in liver homogenates and microsomes but not in slices. This specific method for hexanal, a known product of peroxidation of n-6 polyunsaturated fatty acids, can be used as a good measure of lipid peroxidation.

Aldehydes↗
PubMed1991

Factors affecting the determination of total mercury in biological samples by continuous-flow cold vapor atomic absorption spectrophotometry.

The acidic reduction of Hg using a continuous-flow analytical system was evaluated. With 25% SnCl2 as the reductant, characteristic concentrations (sensitivities) of 0.44 microgram/L (open cell) and 0.29 microgram/L (flow-through cell) were obtained using inorganic Hg2+ standards in 1.5% HCl. When CH3Hg+ standards were used, absorption signals were an order of magnitude lower, indicating that Sn(II) is incapable of producing Hg degree from organic Hg in this acidic reduction system. Addition of CdCl2 to the SnCl2 reductant, as suggested by Magos (1) for the reduction of organomercurials under alkaline conditions, was without beneficial effect. Similarly, combining Sn with another reducing agent (hydroxylamine hydrochloride), or a strong alkaline solution (40% NaOH), in the reaction coil of the flow-through system did not significantly enhance the Hg absorption signal for either inorganic or organic Hg. Recovery of Hg from spiked liver homogenates digested at 70-80 degrees C using a HNO3/H2SO4/HCl procedure and stabilized with 0.5 mM K2Cr2O7 was > 85% using either inorganic Hg2+ or CH3Hg+, indicating that this digestion procedure successfully breaks the C-Hg bond to form readily reducible Hg species. Using L-cysteine to stabilize standards of inorganic Hg2+ in HCl caused significant depressions of the Hg absorption signal at L-cysteine concentrations > 0.001% (approximately 0.5 mM); 0.1% L-cysteine caused total suppression of the Hg signal. These results indicate that: (1) acidic reduction of Hg by Sn in this continuous-flow system requires breakdown of organomercurials prior to analysis; (2) tissue digestion using HNO3/H2SO4/HCl followed by the addition of K2Cr2O7 to stabilize Hg2+ achieves this breakdown and allows good recovery of total Hg; and (3) use of L-cysteine to complex and prevent losses of Hg should be avoided in systems using acidic reduction of Hg. Concentrations of endogenous tissue sulfhydryls are generally lower than those associated with depressed absorbance signals during the acidic reduction of Hg.

Animals↗
PubMed2001

Monitoring of total antimony and its species by ICP-MS and on-line ion chromatography in biological samples from patients treated for leishmaniasis.

Results from a study are reported in which patients with leishmaniasis were monitored by whole blood, blood plasma, urine, and hair analysis, before, during, and after intramuscular administration of N-methyl meglumine antimoniate. Quadrupole ICP-MS was used for the detection of antimony and on-line ion chromatography for the separation of its species. After typically 30 consecutive daily injections of 5 mg antimony per kg of body weight, Sb concentrations of up to 250 microg L(-1) in whole blood and plasma, and 60 mg of Sb per gram of creatinine in urine, were measured 24 h after drug administration. Antimony in hair samples of these patients showed concentrations of up to 24 microg g(-1). Speciation studies of Sb5+ and Sb3+ in drug, urine, and plasma samples were performed by ion chromatography using a Hamilton PRP-100X anion exchange column and EDTA (2 or 20 mM, pH 4.7) as the mobile phases. Repeatability of elution time and peak area measurements for a 0.125 ng spike were <1.2% and <3.5%, respectively. Method detection limits for both species, using a 1:10 diluted urine or plasma sample, were typically 1.6 microg L(-1). The procedure was capable of separating the very intense drug peak from its inorganic species, thus permitting the first studies on the bio-transformation of N-methyl meglumine antimoniate to Sb5+ and Sb3+ in the human body.

Adult↗
PubMed2006

Determination of resorcylic acid lactones in biological samples by GC-MS. Discrimination between illegal use and contamination with fusarium toxins.

An EU project, FAIR5-CT-1997-3443, has been undertaken to distinguish illegal use of zeranol from consumption of food contaminated with Fusarium spp. toxin. One of the tasks was development of screening and confirmatory methods of analysis. This paper describes a new method based on two-step clean-up and GC-MS analysis. The first clean-up step is matrix-dependant; the second is applicable to both urine and meat. The MS is operated in negative chemical ionisation mode. The method is quantitative for zeranol and taleranol, alpha- and beta-zearalenol, and zearalenone and qualitative for zearalanone. Validation was performed according to the latest EU performance criteria (Commission Decision 2002/657). For analysis of urine CC(alpha) and CC(beta) for the method (microg L(-1)) were 0.06-0.11 for zeranol, 0.07-0.12 for taleranol, 0.07-0.11 for alpha-zearalenol, 0.21-0.36 for beta-zearalenol, 0.35-0.60 for zearalenone, and 0.19-0.33 zearalanone. Within-laboratory reproducibility was 16.2, 11.2, 31.9, 30.1, 26.6, and 54.2% for zeranol, taleranol, alpha-zearalenol, beta-zearalenol, zearalenone, and zearalanone, respectively. It was found that all the compounds are stable in urine at -20 degrees C for at least a year. Part of the validation program was organisation of a small proficiency study (ringtest) and a correlation study with an LC-MS-MS method developed by the Veterinary Science Division (VSD; Belfast, UK-NI). This study showed there was good correlation between results from both laboratories. The method can be used for quantitative analysis discriminating illegal use of zeranol from consumption of zearalenone-contaminated food.

Animals↗
PubMed2005

Determination of sterols in biological samples by SPME with on-fiber derivatization and GC/FID.

A new procedure for the determination of sterols in serum samples is proposed. The system consists of coating a Solid Phase Microextraction (SPME) microfiber in headspace mode with the derivatizing agent N,O-bis(trimethylsilyl)trifluoracetamide (BSTFA) and then applying this coated fiber to the simultaneous extraction and derivatization of three precursors in the cholesterol biosynthesis pathway (desmosterol, lathosterol and lanosterol) and two phytosterols (sitosterol and sitostanol) in serum samples. Optimization of the analytical procedure via the application of an experimental design, a study of matrix effects, and an analysis of serum pool samples are all described and discussed.

Flame Ionization↗
PubMed2006

Cholinesterase inhibition based determination of pancuronium bromide in biological samples.

Pancuronium bromide (PCBr) inhibition effect on enzyme cholinesterase from pooled human serum (Che, EC 3.1.1.8 acylcholine acylhydrolase) was used for development of a spectrophotometric kinetic method for PCBr determination in human serum and urine. Optimal conditions for the basic and inhibitor reactions were established: pH=7.7 and substrate concentration c(benzoylcholine chloride)=1.33 mmol/L. Kinetic parameters were also determined: Michaelis-Menten's constant K(M)=0.40 mmol/L, maximal reaction rate V(max)=52.2 micromol/L min, inhibition constant K(i)=0,56 micromol/L and IC(50)=1.31 micromol/L. Linear dependence between the reaction rate and inhibitor concentration exists in PCBr concentration range 8.20-68.25 nmol/L, which corresponds to the real sample concentrations from 0.328 to 2.730 micromol/L. The method detection and quantification limits were 2.01 nmol/L and 6.67 nmol/L, respectively. Precision of the method was tested for three pancuronium concentrations (10.70, 29.35 and 51.25 nmol/L). Relative standard deviation (RSD) was in the range 0.15-7.45%. Accuracy was examined by standard addition method. Influence of the substances usually present in serum and urine on the reaction rate was tested. The developed method was applied for PCBr content determination in serum model samples, urine model samples and in urine taken during surgery. The method has good sensitivity, accuracy, precision and it is suitable for clinical practice.

Cholinesterases↗
PubMed1999

A rapid and quantitative method for total fatty acid analysis of fungi and other biological samples.

A quantitative method for the one-step esterification and determination of absolute amounts of polyunsaturated fatty acids in biotechnologically produced fungal mycelia is proposed. A system of two internal standards was used to quantify the total fatty acid content of the samples by gas chromatography. The degree of methylation of the analyzed fatty acids was calculated using the internal standards, and subsequently this quotient was used to validate the derivatization and extraction reactions. By utilizing this degree of methylation, the corrected amounts of the various fatty acids in the sample can be calculated.

Chromatography, Gas↗