Permeation HPLC Research Articles

Analysis of polycyclic aromatic hydrocarbons and their nitro‐derivatives in farm road and barn dust

Road and barn dust samples were analysed for polycyclic aromatic hydrocarbons (PAHs) and nitrated polycyclic hydrocarbons (nitro‐PAHs). For this purpose, suitability of optimised HPLC with fluorimetric detection and GC/MS, both preceded by Soxhlet extraction of samples, gel permeation chromatography and HPLC with silica gel column for clean‐up, fractionation and preconcentration of the analytes, were compared. The detection limits of GC/MS for 16 PAHs ranged from 5 to 10 pg. The detection limits of GC/MS for 11 nitro‐PAHs ranged from 20 to 60 pg and those of on‐line reduction HPLC with fluorescence detection from 0.5 to 5 pg. Therefore, procedures with a higher separation efficiency and a more selective detection, such as GC/MS should be preferred when low concentrations of nitro‐PAHs are expected. PAH concentrations in road dust and barn dust samples ranged from 527 to 5680 μg/kg, and from 114 to 740 μg/kg, respectively. Nitro‐PAHs were detected in one sample of barn dust and two samples of road dust. The concentrations of 2‐nitrofluorene, 1‐nitropyrene, and 6‐nitrochrysene in barn dust were 0.50, 0.72 and 1.25 μg•kg‐1, respectively. Only 1‐nitropyrene at concentrations of 0.11 and 0.29 μg•kg‐1 was detected in road dust. As far as we know, this is the first report on the occurrence of nitro‐PAHs in the farm environment.

Determination of PAH in airborne particulates using gel permeation chromatography and HPLC ‐fluorescence ‐ UV

A method based on gel permeation chromatography as sample clean‐up followed by HPLC ‐ fluorescence ‐ UV is successfully developed for measuring the contents of 16 PAHs in airborne particulate after Soxhlet extraction of the samples. The clean‐up process is proved to be efficient by the analysis of SRM from NIST. The HPLC is operated at gradient steps with constant flow followed by UV and optimized multi‐step fluorescence detection. A complete run of HPLC requires less than 30 minutes with satisfactory resolution. The detection limits obtained with UV detection ranged from 0.017 to 2.542 ng/m3, and those of fluorescence are from 0.001 to 0.044 ng/m3. Precision of measurement is within 9.0 %. This method is applied to monitor the PAH contents is the airborne particulate samples collected in Taipei city at ten different sampling sites with particle size distribution measurements.

Purification of a candidate gonadotropin surge inhibiting factor from porcine follicular fluid.

Several lines of evidence suggest that the ovaries of many species produce a nonsteroidal substance, termed gonadotropin surge inhibiting factor (GnSIF), which inhibits the midcycle gonadotropin surge and attenuates the pituitary response to endogenous or exogenous GnRH. We have previously reported the partial purification of GnSIF from porcine follicular fluid (pFF) and its differentiation from inhibin. We present now the purification of GnSIF to homogeneity and determination of the partial NH2-terminal amino acid sequence. The bioassay for GnSIF used rat pituitary cells in short-term culture that were incubated with test fractions for 48 h, washed, and then incubated with 10 nM GnRH plus test fractions for 4 h. GnSIF activity is defined as the suppression of GnRH-stimulated LH secretion. GnSIF was purified from 500 ml of pFF using sequential heparin-Sepharose, anion-exchange, and cation-exchange liquid chromatography followed by gel permeation, hydrophobic interaction, and mono-Q HPLC steps. Using these six purification steps, we have obtained an apparently homogenous preparation that stains as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. GnSIF has an apparent mol wt of 69K. Limited NH2-terminal sequence analysis reveals that GnSIF has no sequence homology with other reproductive hormones including the inhibins, activins, and follistatins. Over the dose range tested, GnSIF had no effect on basal LH or FSH secretion by pituitary cells in culture and only slightly inhibited GnRH-stimulated FSH secretion at the highest dose tested. In addition, there was no inhibin or follistatin immunoactivity in the GnSIF preparation. As such, GnSIF appears to be a novel protein in pFF that inhibits GnRH-stimulated LH secretion, and which may participate along with other ovarian proteins and steroids in the regulation of pituitary gonadotropin secretion.

Distribution and characterisation of neuropeptide Y immunoreactivity in the brain and gastrointestinal tract of the rainbow trout, Oncorhynchus mykiss

Abstract 1. Neuropeptide Y (NPY) immunoreactivity has been localised cytochemically in neuronal somata and fibres in rainbow trout brain, nerve fibres and mucosal epithelial endocrine cells within the gastrointestinal tract and in endocrine cells within pancreatic islets. 2. Using a C-terminal specific NPY radioimmunoassay, immunoreactivity was detected in extracts of brain (519 pmol/g), cardiac stomach (37.9 pmol/g), pyloric stomach plus pancreas (37.9 ol/g) and intestine (29.2 pmol/g). 3. Gel permeation and reverse-phase HPLC analysis of brain and intestinal extracts resolved a single NPY immunoreactive peptide.

The Mechanism of the Crosslinking Reaction of Gelatin with Hardener

The reaction, in solution, between gelatin and various hardeners has been examined by gel permeation chromatography, HPLC and amino acid analysis. It is concluded that there are two types of crossl...

Production, isolation and identification of low molecular mass peptides from blue cheese by high performance liquid chromatography

SummaryThe blue cheese nitrogenous fractions soluble in water and in 5% phosphotungstic acid (PTA) were analysed by HPLC after 3–180 d ripening. In the water-soluble fraction, in addition to four or five major peaks corresponding to amino acids, there were many minor peaks, which increased during ripening. The low molecular mass peptides, soluble in 5% PTA, showed ripening-induced increments. A method combining precipitation with 5% PTA, gel permeation and subsequent HPLC was used to isolate some peptides of cheese. Four peptides containing between seven and ten residues were isolated and their amino acid composition and N-terminal residues determined.

Purification of a calcium binding protein (rat SPARC) from primary Sertoli cell-enriched culture medium

A protein with an apparent Mr of 43,000 was purified from Sertoli cell-enriched culture medium by sequential anion-exchange, gel permeation, C4 reversed-phase, and diphenyl reversed-phase HPLC. N-Terminal sequence analysis of this protein revealed a sequence of NH2-XPQTEAAEEMVAEETVV for the first 17 amino acids. Comparison of this sequence with existing protein data base maintained at the Protein Identification Resource revealed that it shares extensive identity with a previously described protein secreted by mouse embryo parietal endoderm, SPARC, which is equivalent to a protein secreted by a basement-membrane-producing tumor, BM-40; and a bone protein, osteonectin. This protein also possesses similar in vitro biological activity of SPARC in which it binds Ca2+. The possible physiological significance of this protein was discussed.