The aim of this study was to develop a simple and derivatization free method for the Quantification of S-Epichlorohydrin in R-Epichlorohydrin by using a gas chromatography coupled with flame ionization detector (FID)....The aim of this study was to develop a simple and derivatization free method for the Quantification of S-Epichlorohydrin in R-Epichlorohydrin by using a gas chromatography coupled with flame ionization detector (FID). Enantiopure epichlorohydrin was a valuable epoxide key starting material for preparing optically active Rivaroxaban. The enantiomeric separations of S-Epichlorohydrin and R-Epichlorohydrin were achieved on Gamaa-Dex-225 (30 meters × 0.25 mm I.D, 0.25 μm) column with a total run time of 30 min. Nitrogen was used as a carrier gas with constant pressure 25.0 psi. The critical experimental parameters such as, column selection, flow rate, injection volume and diluent were studied and optimized. Excellent correlation coeffient between peak responses and concentrations was >0.9998. The recoveries of S-Epichlorohydrin spiked in R-Epichlorohydrin were in the range from 98.2% to 102.8%. Limit of quantitation for S-Epichlorohydrin was sufficiently lower than limits specified by ICH. The method has validated as per International Conference on Harmonization (ICH) guidelines. A precise, accurate, linear and robust Gas Chromatography method was developed for the quantification of S-Epichlorohydrin in R-Epichlorohydrin for Rivaroxaban.展开更多
A new and enantioselective liquid chromatographic method was developed for estimation of S-Linagliptin in Linagliptin (LINA) drug substances. The desired enantiomeric separation was achieved on Chiralpak AD-H (250*4.6...A new and enantioselective liquid chromatographic method was developed for estimation of S-Linagliptin in Linagliptin (LINA) drug substances. The desired enantiomeric separation was achieved on Chiralpak AD-H (250*4.6 mm*5 μm) column with the mobile phase composition of ethanol, methanol and diethylamine in a ratio of 90:10:0.1 (v/v/v) with flow rate of 0.5 mL·min<sup>-</sup><sup>1</sup> and column oven temperature 30°C and the eluted compounds were monitored at 225 nm. In the proposed chiral method, USP resolutions between both the enantiomers were more than 5.0. Limit of detection and Limit of quantitation of S-LINA was found to be 0.03 μg·mL<sup>-1</sup> and 0.10 μg·mL<sup>-1</sup> respectively. Linearity study was conducted from LOQ to 150% and correlation coefficient found to be 0.9997. Accuracy was within the range of 98.6% to 101.5%. To prove selectivity power of the method specificity study was conducted by subjecting drug substance to acid, base, hydrolysis, oxidation and photolysis and ensured the peak purity of analyte in degraded samples. Moreover, the method has been fully validated as per ICH guidelines. The proposed method is precise, accurate, linear, rugged, robust and suitable for accurate quantification of S-LINA in LINA drug substance.展开更多
Background and Objective: Islet autotransplantation is performed to preserve endocrine function in patients undergoing pancreatic resections for painful chronic pancreatitis. We characterized islets isolated from chro...Background and Objective: Islet autotransplantation is performed to preserve endocrine function in patients undergoing pancreatic resections for painful chronic pancreatitis. We characterized islets isolated from chronic pancreatitis patients (CP) of tropical region. Patients and Methods: Pancreatic tissues were obtained from CP patients with and without diabetes undergoing pancreatic resections (n = 35) and brain-dead multi organ donors (n = 6;considered as controls). Islets isolated were assessed for yield, purity, viability and in vitro islet function (Glucose stimulated insulin release, GSIR) as per standard protocols. Results: Islets from CP patients without diabetes were similar to controls in yield (control 4120 - 6100 IE/g, CP 3550 - 5660 IE/g), purity (control 78% ± 12%, CP 70% ± 8.2%) and viability (control 85% ± 8%, CP 81% ± 10%) and islets from CP patients with diabetes showed decreases in yield (3002 - 2300 IE/g), purity (61% ± 16%) and viability (62% ± 21%). Islets measuring 50 - 200 μ were similar in abundance in controls (94.74% ± 3.2%) and CP patients with and without diabetes, 86.31% ± 4.9%, 91.03% ± 3.8%. GSIR of islets from CP patients and controls were similar at 5.5 mM glucose (2.8 - 3.1 μU/ml). However, GSIR at 16.5 mM glucose was decreased in CP patients (control 18.5 ± 0.6, CP without diabetes 11.8 ± 0.3, CP with diabetes 4.3 ± 0.3 μU/ml). Conclusion: Our results demonstrate suitability of islets isolated from CP patients of tropical region for autotransplantation.展开更多
Background/Aim: MicroRNAs with regulatory functions in gene expression are implicated in different diseases. The present study investigated differentially expressed miRNAs that possibly influence transcription factors...Background/Aim: MicroRNAs with regulatory functions in gene expression are implicated in different diseases. The present study investigated differentially expressed miRNAs that possibly influence transcription factors involved in insulin gene expression in Chronic Pancreatitis (CP) employing bioinformatics approaches. Methods: Pancreatic tissues were collected from CP patients undergoing partial pancreatectomy (n = 16) and controls (n = 15) undergoing resections for non-pancreatic malignancies. MiRNA profiles obtained using microarrays were validated by qRT-PCR. Target search involving miRWalk and TarBase as well as functional annotation employing KEGG (Kyoto encyclopedia of genes and genomes) and DAVID (Database for Annotation) databases were performed. Ingenuity pathway analysis (IPA) was used to construct networks relating miRNAs to their target genes. mRNA and proteins related to insulin gene transcription factors and hormones were evaluated by qRT-PCR and western blotting followed by confirmation upon immunofluorescent staining. Results: Microarray data revealed 10 up-regulated and 15 down-regulated miRNAs in CP as compared to controls (Log2 FC > 2). Bioinformatic analysis showed 8399 target genes and KEGG pathway analysis suggested a role for the dysregulated miRNAs in modulating cytokine signaling, fibrosis, JAK-STAT signaling and insulin synthesis. IPA analysis suggested a simplified network attributing dysregulated miRNAs to NFκB-dependent cytokine signaling. Further, associations could be noted between miRNA 200b with Maf A, 138-1 with Neuro D and 27b with FoxO1. Decreases in mRNA levels of Pdx1, Neuro D and increases of Maf A and FoxO1 transcription factors could be noted (P Conclusion: Our results identified dysregulation of miRNAs 138-1, 27b and 200b which were found to be associated with insulin gene transcription factors Neuro D, FoxO1 and Maf A respectively.展开更多
文摘The aim of this study was to develop a simple and derivatization free method for the Quantification of S-Epichlorohydrin in R-Epichlorohydrin by using a gas chromatography coupled with flame ionization detector (FID). Enantiopure epichlorohydrin was a valuable epoxide key starting material for preparing optically active Rivaroxaban. The enantiomeric separations of S-Epichlorohydrin and R-Epichlorohydrin were achieved on Gamaa-Dex-225 (30 meters × 0.25 mm I.D, 0.25 μm) column with a total run time of 30 min. Nitrogen was used as a carrier gas with constant pressure 25.0 psi. The critical experimental parameters such as, column selection, flow rate, injection volume and diluent were studied and optimized. Excellent correlation coeffient between peak responses and concentrations was >0.9998. The recoveries of S-Epichlorohydrin spiked in R-Epichlorohydrin were in the range from 98.2% to 102.8%. Limit of quantitation for S-Epichlorohydrin was sufficiently lower than limits specified by ICH. The method has validated as per International Conference on Harmonization (ICH) guidelines. A precise, accurate, linear and robust Gas Chromatography method was developed for the quantification of S-Epichlorohydrin in R-Epichlorohydrin for Rivaroxaban.
文摘A new and enantioselective liquid chromatographic method was developed for estimation of S-Linagliptin in Linagliptin (LINA) drug substances. The desired enantiomeric separation was achieved on Chiralpak AD-H (250*4.6 mm*5 μm) column with the mobile phase composition of ethanol, methanol and diethylamine in a ratio of 90:10:0.1 (v/v/v) with flow rate of 0.5 mL·min<sup>-</sup><sup>1</sup> and column oven temperature 30°C and the eluted compounds were monitored at 225 nm. In the proposed chiral method, USP resolutions between both the enantiomers were more than 5.0. Limit of detection and Limit of quantitation of S-LINA was found to be 0.03 μg·mL<sup>-1</sup> and 0.10 μg·mL<sup>-1</sup> respectively. Linearity study was conducted from LOQ to 150% and correlation coefficient found to be 0.9997. Accuracy was within the range of 98.6% to 101.5%. To prove selectivity power of the method specificity study was conducted by subjecting drug substance to acid, base, hydrolysis, oxidation and photolysis and ensured the peak purity of analyte in degraded samples. Moreover, the method has been fully validated as per ICH guidelines. The proposed method is precise, accurate, linear, rugged, robust and suitable for accurate quantification of S-LINA in LINA drug substance.
文摘Background and Objective: Islet autotransplantation is performed to preserve endocrine function in patients undergoing pancreatic resections for painful chronic pancreatitis. We characterized islets isolated from chronic pancreatitis patients (CP) of tropical region. Patients and Methods: Pancreatic tissues were obtained from CP patients with and without diabetes undergoing pancreatic resections (n = 35) and brain-dead multi organ donors (n = 6;considered as controls). Islets isolated were assessed for yield, purity, viability and in vitro islet function (Glucose stimulated insulin release, GSIR) as per standard protocols. Results: Islets from CP patients without diabetes were similar to controls in yield (control 4120 - 6100 IE/g, CP 3550 - 5660 IE/g), purity (control 78% ± 12%, CP 70% ± 8.2%) and viability (control 85% ± 8%, CP 81% ± 10%) and islets from CP patients with diabetes showed decreases in yield (3002 - 2300 IE/g), purity (61% ± 16%) and viability (62% ± 21%). Islets measuring 50 - 200 μ were similar in abundance in controls (94.74% ± 3.2%) and CP patients with and without diabetes, 86.31% ± 4.9%, 91.03% ± 3.8%. GSIR of islets from CP patients and controls were similar at 5.5 mM glucose (2.8 - 3.1 μU/ml). However, GSIR at 16.5 mM glucose was decreased in CP patients (control 18.5 ± 0.6, CP without diabetes 11.8 ± 0.3, CP with diabetes 4.3 ± 0.3 μU/ml). Conclusion: Our results demonstrate suitability of islets isolated from CP patients of tropical region for autotransplantation.
文摘Background/Aim: MicroRNAs with regulatory functions in gene expression are implicated in different diseases. The present study investigated differentially expressed miRNAs that possibly influence transcription factors involved in insulin gene expression in Chronic Pancreatitis (CP) employing bioinformatics approaches. Methods: Pancreatic tissues were collected from CP patients undergoing partial pancreatectomy (n = 16) and controls (n = 15) undergoing resections for non-pancreatic malignancies. MiRNA profiles obtained using microarrays were validated by qRT-PCR. Target search involving miRWalk and TarBase as well as functional annotation employing KEGG (Kyoto encyclopedia of genes and genomes) and DAVID (Database for Annotation) databases were performed. Ingenuity pathway analysis (IPA) was used to construct networks relating miRNAs to their target genes. mRNA and proteins related to insulin gene transcription factors and hormones were evaluated by qRT-PCR and western blotting followed by confirmation upon immunofluorescent staining. Results: Microarray data revealed 10 up-regulated and 15 down-regulated miRNAs in CP as compared to controls (Log2 FC > 2). Bioinformatic analysis showed 8399 target genes and KEGG pathway analysis suggested a role for the dysregulated miRNAs in modulating cytokine signaling, fibrosis, JAK-STAT signaling and insulin synthesis. IPA analysis suggested a simplified network attributing dysregulated miRNAs to NFκB-dependent cytokine signaling. Further, associations could be noted between miRNA 200b with Maf A, 138-1 with Neuro D and 27b with FoxO1. Decreases in mRNA levels of Pdx1, Neuro D and increases of Maf A and FoxO1 transcription factors could be noted (P Conclusion: Our results identified dysregulation of miRNAs 138-1, 27b and 200b which were found to be associated with insulin gene transcription factors Neuro D, FoxO1 and Maf A respectively.
