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Reverse-phase High Performance Liquid Chromatography (reverse-phase + high_performance_liquid_chromatography)
Selected AbstractsIdentification of proNeuropeptide FFA peptides processed in neuronal and non-neuronal cells and in nervous tissueFEBS JOURNAL, Issue 20 2003Elisabeth Bonnard Peptides which should be generated from the neuropeptide FF (NPFF) precursor were identified in a neuronal (human neuroblastoma SH-SY5Y) cell line and in COS-7 cells after transient transfection of the human proNPFFA cDNA and were compared with those detected in the mouse spinal cord. After reverse-phase high performance liquid chromatography of soluble material, NPFF-related peptides were immunodetected with antisera raised against NPFF and identified by using on-line capillary liquid chromatography/nanospray ion trap tandem mass spectrometry. Neuronal and non-neuronal cells generated different peptides from the same precursor. In addition to NPFF, SQA-NPFF (Ser-Gln-Ala-Phe-Leu-Phe-Gln-Pro-Gln-Arg-Phe-amide) and NPAF were identified in the human neuroblastoma while only NPFF was clearly identified in COS-7 cells. In mouse, in addition to previously detected NPFF and NPSF, SPA-NPFF (Ser-Pro-Ala-Phe-Leu-Phe-Gln-Pro-Gln-Arg-Phe-amide), the homologous peptide of SQA-NPFF, were characterized. These data on intracellular processing of proNeuropeptide FFA are discussed in regard to the known enzymatic processing mechanisms. [source] ORGANIC ACIDS PROFILE IN TOMATO JUICE BY HPLC WITH UV DETECTIONJOURNAL OF FOOD QUALITY, Issue 1 2007OMBRETTA MARCONI ABSTRACT A simple method was developed to determine 10 organic acids simultaneously in tomato products using reverse-phase high performance liquid chromatography (HPLC) column with the diode array detector set at 210 nm. After centrifugation and filtration, the samples were passed through to an anion exchange resin and the organic acids were released using 0.1 N HCl. The chromatographic separation was achieved with isocratic analysis in a 20-min run. The method was reliable and sensitive. The coefficient of determination of the standard calibration curve is 0.9925 , r2 , 0.9999 and the limit of detection ranged from 0.08 to 6.00 mg/kg for trans -aconitic acid and acetic acid, respectively. The limit of quantification ranged from 0.19 to 15.18 mg/kg for trans-aconitic and acetic acid, respectively. To establish the efficiency of the anion resin the procedure was applied to a standard solution of a mixture of organic acids. The organic acids recovery ranged from 87.0% ± 1.9 for citramalic acid to 109.9% ± 5.2 for fumaric acid. [source] Novel angiotensin I-converting enzyme inhibitory peptides isolated from Alcalase hydrolysate of mung bean proteinJOURNAL OF PEPTIDE SCIENCE, Issue 8 2006Guan-Hong Li Abstract Mung bean protein isolates were hydrolyzed for 2 h by Alcalase. The generated hydrolysate showed angiotensin I-converting enzyme (ACE) inhibitory activity with the IC50 value of 0.64 mg protein/ml. Three kinds of novel ACE inhibitory peptides were isolated from the hydrolysate by Sephadex G-15 and reverse-phase high performance liquid chromatography (RP-HPLC). These peptides were identified by amino acid composition analysis and matrix assisted-laser desorption/ionization time-of-flight tandem mass spectrometry (MALDI-TOF MS/MS), as Lys-Asp-Tyr-Arg-Leu, Val-Thr-Pro-Ala-Leu-Arg and Lys-Leu-Pro-Ala-Gly-Thr-Leu-Phe with the IC50 values of 26.5 µM, 82.4 µM and 13.4 µM, respectively. Copyright © 2006 European Peptide Society and John Wiley & Sons, Ltd. [source] Prostaglandin D2 pathway and peroxisome proliferator-activated receptor ,-1 expression are induced by mechanical loading in an osteoblastic cell lineJOURNAL OF PERIODONTAL RESEARCH, Issue 2 2006Chitpol Siddhivarn Objective:, The hypothesis underlying the current study was that the arachidonic acid cascade, specifically activation of the prostaglandin (PG) D2 pathway in osteoblasts, is an anabolic signal induced by mechanical loading. Background:, Previous studies have shown that mechanical loading of osteoblasts triggers cyclooxygenase (COX)-2, PGE2 and prostacyclin (PGI2) synthesis. Since modest mechanical loading of osteoblasts promotes bone formation, we sought to determine whether mechanical stress activates the osteoblastic PGD2 pathway resulting in the synthesis of osteogenic cyclopentenones, including ,12PGJ2. Methods:, Osteoblast monolayers were stretched using a Bioflex apparatus at a frequency of 1 Hz with 1% elongation. Cells and cell media were collected at various time points: 5, 10, 15, 30 min; and 1, 4, 16, 24 h. RNA was extracted for quantitative reverse transcriptase,polymerase chain reaction (RT,PCR). In certain experiments, cells were pre-labeled with 14C arachidonic acid prior to stretching. Radiolabeled metabolites in cell media were identified by reverse-phase high performance liquid chromatography (RP-HPLC). Osteoblasts were evaluated for an induction in bone nodule formation by stretching. Results:, Mechanical strain significantly increased mRNA expression of COX-1, COX-2, PGD2 synthase and peroxisome proliferator-activated receptor (PPAR) ,-1, but not of PPAR,-2 as compared to control unstretched cells (p < 0.05). Mechanical loading stimulated the release of PGE2, PGD2 and the PGD2 metabolite ,12PGJ2. Mechanical strain resulted in the induction of bone nodules. Conclusions:, This report indicates that mechanical loading of osteoblasts results in activation of PGD2 and the concomitant expression of transcription factor PPAR,-1 mRNA. The coordinated synthesis of ,12PGJ2, a natural ligand for PPAR,-1, with the increased expression of PPAR,-1, suggests that biomechanical transduction pathways that initially involve the activation of cyclooxygenases may also involve the activation of the ,12PGJ2,PPAR pathway. [source] | ||||||||||