Quantitative determination of acetyl glucoside isoflavones and their metabolites in human urine using combined liquid chromatography-mass spectrometry.

To investigate whether the bioavailability of isoflavones could be an alternative to fermented soy foods, the conjugated forms of soy nutritional supplement (containing 98% acetyl glucoside isoflavones) were consumed by eight human volunteers (three were Asian people and five were British). Their daily urine samples were collected before and after a 5-week consumption of supplementation period. Conjugated isoflavones of genistein, daidzein and glycitein were hydrolyzed by enzyme, extracted with methyl tert-butyl ether and analysed using liquid chromatography coupled with electrospray tandem mass spectrometry. Daidzein, genistein, glycitein, dihydrogenistein, dihydrodaidzein and O-desmethylangolensin were identified and quantified simultaneously with high recoveries. The levels of free isoflavones and total isoflavones were compared, and isoflavone glucuronides were identified much higher than the corresponding sulfates or aglycone isoflavones. This method provided the measurement of isoflavones with high sensitivity and specificity and simplified the sample pre-treatment procedure. The limitation of detections of dihydrodaidzein, 3'-hydroxydaidzein, glycitein, daidzein, genistein, dihydrogenistein and O-desmethylangolensin were 37, 23.5, 12.2, 15.4, 14.8, 2.20 and 0.31 pmol, respectively. Only 0.5 ml of urine sample was needed.

Application of atmospheric pressure chemical ionisation mass spectrometry in the analysis of barbiturates by high-speed analytical countercurrent chromatography.

Four barbiturates (barbital, allobarbital, phenobarbital and butalbital) were analysed using high-speed analytical countercurrent chromatography (HSACCC) and high-performance liquid chromatography (HPLC) interfaced with mass spectrometry, using negative mode atmospheric pressure chemical ionisation (APCI). The polar biphasic solvent system of butyronitrile/acetonitrile/water (1:1:1) was used, in the upper-stationary, lower-mobile mode of operation, at a flow rate of 1 mL/min and a rotational speed of 1200 rpm, equating to an applied "g"-field of 177 g. The fractional stationary phase retention (S(F)) was 0.58. Representative mass spectral data are presented from the HPLC and the HSACCC analyses. Structural information was obtained using source-induced fragmentation at increased source block voltages. The effect of increasing g-field on chromatographic resolution is illustrated using the binary base system of butyronitrile/water (1:1), under electrospray ionisation.

Analysis of urinary nucleosides. III. Identification of 5'-deoxycytidine in urine of a patient with head and neck cancer.

Modified nucleosides in the urine have been postulated to be diagnostic indices of disease, particularly of cancer. The urine of a patient with terminal head and neck cancer has been found to contain a modified nucleoside with a protonated molecule of m/z 228. By means of high-performance liquid chromatography/ion trap mass spectrometry (HPLC/ITMS) and capillary liquid chromatography/triple quadruple mass spectrometry (CapLC/TQMS) we have identified the compound as 5'-deoxycytidine. This is the first report of 5'-deoxycytidine in man: in addition to the elucidation of its structure, its possible origins and the potential significance of its occurrence are discussed.

Isolation and identification of four flavonoid constituents from the seeds of Oroxylum indicum by high-speed counter-current chromatography.

Four flavonoids, chrysin, baicalein, baicalein-7-O-glucoside, baicalein-7-O-diglucoside (Oroxylin B) and one unknown flavonoid have been isolated and purified for the first time in the seeds of Oroxylum indicum by high-speed counter-current chromatography with a two-phase solvent system composed of chloroform-methanol-water (8:10:5, v/v). Then, 50 mg baicalein-7-O-glucoside, 10.5 mg baicalein-7-O-diglucoside, 4.5 mg chrysin-7-O-diglucoside, 25 mg baicalein and 45 mg chrysin could be obtained after injecting 20 mg/ml sample extract ten times and their purities were 96, 90, 85, 95 and 98%, respectively. All these constituents were identified by high-performance liquid chromatography-mass spectrometry and nuclear magnetic resonance.

Cyclic nucleotide content of tobacco BY-2 cells.

The cyclic nucleotide content of cultured tobacco bright yellow-2 (BY-2) cells was determined, after freeze-killing, perchlorate extraction and sequential chromatography, by radioimmunoassay. The identities of the putative cyclic nucleotides, adenosine 3',5'-cyclic monophosphate (cyclic AMP), guanosine 3',5'-cyclic monophosphate (cyclic GMP) and cytidine 3',5'-cyclic monophosphate (cyclic CMP) were unambiguously confirmed by tandem mass spectrometry. The potential of BY-2 cell cultures as a model system for future investigations of cyclic nucleotide function in higher plants is discussed.