A simple efficient isocratic reversed-phase HPLC method was developed and validated for the determination of clindamycin palmitate hydrochloride (CPH) and its commercially available oral solution products. Separation ...A simple efficient isocratic reversed-phase HPLC method was developed and validated for the determination of clindamycin palmitate hydrochloride (CPH) and its commercially available oral solution products. Separation was achieved on a Phenomenex Zorbax (Luna) cyano column (150 × 4.6 mm, 5 μm) with a Phenomenex cyano guard cartridge (4 × 3.0 mm) on Agilent 1050 series HPLC system. CPH and its resolution standard lincomycin were eluted isocratically at a flow rate of 1 mL/min with a simplified mobile phase (potassium phosphate buffer (5 mM, pH 3.0)—acetonitrile—tetrahydrofuran (20:75:5, v/v/v)) and detected at 210 nm. The column was maintained at 25?C. The method was validated according to USP category I requirements. Robustness and forced degradation studies were also conducted. CPH marketed drug products were obtained from a drug distributor and assayed for potency using the validated method. Validation acceptance criteria were met in all cases. The analytical range for CPH was 15 - 500 μg/mL and the linearity was r2 > 0.999 over three days. The method was determined to be specific and robust. Both accuracy (92.0% - 103.8%) and precision (0.67% - 1.52%) were established across the analytical range for low, intermediate and high QC concentrations. Method applicability was demonstrated by analyzing two marketed products of CPH, in which results showed potency >98%. The method was determined to be an enhancement over the current USP methodology for assay as a result of increased efficiency, reduced organic solvents and the elimination of matrix modifiers. This method was successfully applied for the quality assessment of: 1) currently marketed drug products and 2) will in future assess the product quality of novel dosage forms of CPH for pediatric use.展开更多
The mechanism of degradation of dantrolene in aqueous buffer solutions was studied at various pH values in the range of pH 1.2-9.5 and at temperatures ranging from 25℃ to 75℃ to determine the optimum pH and temperat...The mechanism of degradation of dantrolene in aqueous buffer solutions was studied at various pH values in the range of pH 1.2-9.5 and at temperatures ranging from 25℃ to 75℃ to determine the optimum pH and temperature requirements for its stability and eventual product performance over the human gastrointestinal pH range. Dantrolene was analyzed by reversed phase ultra-performance liquid chromatographic (UPLC). Chromatographic separation was achieved on a Waters Acquity UPLC system using a Waters BEH C18 analytical column and Waters BEH C18 guard column. The compounds were eluted with a linear acetonitrile gradient (25%-75%) over three minutes with a buffer composition of 2.0 mM of sodium acetate at pH 4.5 for degradation studies. The flow rate was maintained at 0.5 mL/min. Column temperature was maintained at 35℃. Injection volume was 4 μL and the degradation products were detected by a photodiode array (PDA) detector at 375 nm. Degradation products, including compound B and C were analyzed by mass spectroscopy (MS) and nuclear magnetic resonance spectroscopy (NMR) and the degradation pathways were proposed. Degradation of dantrolene followed pseudo first–order kinetics and a V-shaped pH-rate profile over the pH range 1.2-9.5. The maximum stability was observed at pH 7.4 and 37℃. Although the focus of this paper was on the mechanism of hydrolysis of dantrolene, the poor aqueous solubility of dantrolene, the developed understanding can be utilized to improve the quality of the formulation and the risk associated with the extravasation of dantrolene sodium solution in its current form.展开更多
The purpose of this study was to evaluate the impact of polystyrene divinylbenzene copolymer HPLC columns on the chromatographic performance of the USP compendial method for doxycycline hyclate. The compendial method ...The purpose of this study was to evaluate the impact of polystyrene divinylbenzene copolymer HPLC columns on the chromatographic performance of the USP compendial method for doxycycline hyclate. The compendial method was implemented based on the assessment of the chromatographic performance of six USP defined L21 polystyrene divinylbenzene HPLC columns. Modifications to the method were based on USP for chromatography. The method was validated for the determination of doxycycline hyclate and its impurities in commercially available drug products. A number of different polystyrene-divinylbenzene columns were tested and failed to provide selectivity for the resolution of doxycycline and its impurities. Separation was optimally achieved on an Agilent PLPR-S column (250 × 4.6 mm, 8 μm) by using an Agilent 1260 series HPLC system. Doxycycline hyclate and its impurities were eluted isocratically at a flow rate of 1 mL/min with mobile phase and detected at 270 nm. The column temperature was maintained at 60oC. The method was validated according to USP category I requirements for Assay. Validation acceptance criteria were met in all cases. The analytical range for doxycycline hyclate was 50 - 250 μg/mL and the linearity was r2 > 0.999 over three days. The method was determined to be specific. Both accuracy (95.1% - 102.4%) and precision (0.50% - 4.8%) were established across the analytical range for low, intermediate and high QC concentrations. Method applicability was demonstrated by analyzing marketed products of doxycycline hyclate, in which results showed potency meeting USP acceptance criteria. In conclusion, this study described the remarkable differences in selectivity that were encountered during the implementation phase for the compendial methods for doxycycline and its impurities in marketed products and it could be used in the future to assss the product quality of doxycycline hyclate capsules stored in the National stockpiles.展开更多
The development and implementation of advanced analytical technologies is essential for extending the expiry for complex drug products stored in the Strategic National Stockpiles. Consequently, a novel Ultra High-Perf...The development and implementation of advanced analytical technologies is essential for extending the expiry for complex drug products stored in the Strategic National Stockpiles. Consequently, a novel Ultra High-Performance Liquid Chromatographic (UHPLC) method has been developed for the analysis of atropine and its respective impurities to support the analytical research platform for auto-injectors. This study is part of a larger research effort to improve the efficiency and broaden the applicability of advanced analytical methods for medical counter-measure medications. The current HPLC compendial methodology for atropine sulfate injection requires an analysis time of 40 minutes for atropine. In comparison, the novel gradient UHPLC method required only 8 minutes to evaluate both atropine and its major pharmaceutical impurities. Improved separation was achieved on a Waters Acquity UHPLC BEH C18 1.7 μm, 2.1 × 100 mm column employing gradient elution of mobile phase solvent A (0.1% H3PO4) and solvent B (0.1% H3PO4, 90% ACN, and 10% H2O). The method was validated according to USP Category I requirements for assay. The daily standard calibration curves were linear over a concentration range from 50 μg/mL to 250 μg/mL with a correlation coefficient of >0.999. The detection limit (LOD) and quantitation limit (LOQ) were 3.9 μg/ml and 13.1 μg/ml, respectively. Resolution results indicate that atropine and the following impurities, degradants and a preservative can also be separated and analyzed using this proposed method: noratropine, 4,4’-di-hy-droxydiphenyl ether, 2,4’-dihydroxydiphenyl ether, 4-bromophenol, 4-hydro-xyatropine, tropic acid, apoatropine HCl, atropic acid, hydroquinone, nitroethane, phenol and catechol. The UHPLC method demonstrated enhanced selectivity and significantly reduced the analysis time when compared with the traditional USP compendial HPLC method. The method was successfully applied to the evaluation of atropine in ATNAA auto-injectors lots from the Strategic National Stockpiles.展开更多
Compared to small molecule process analytical technology （PAT） applications, biotechnology product PAT applications have certain unique challenges and opportunities. Understanding process dynamics of bioreactor cell...Compared to small molecule process analytical technology （PAT） applications, biotechnology product PAT applications have certain unique challenges and opportunities. Understanding process dynamics of bioreactor cell culture process is essential to establish an appropriate process control strategy for biotechnology product PAT applications. Inline spectroscopic techniques for real time monitoring of bioreactor cell culture process have the distinct potential to develop PAT approaches in manufac- turing biotechnology drug products. However, the use of inline Fourier transform infrared （FTIR） spectroscopic techniques for bioreactor cell culture process monitoring has not been reported. In this work, real time inline FTIR Spectroscopy was applied to a lab scale bioreactor mAb IgG3 cell culture fluid biomolecular dynamic model. The technical feasibility of using FTIR Spectroscopy for real time tracking and monitoring four key cell culture metabolites （including glucose, glutamine, lactate, and ammonia） and protein yield at increasing levels of complexity （simple binary system, fully formulated media, actual bioreactor cell culture process） was evaluated via a stepwise approach. The FTIR fingerprints of the key metabolites were identified. The multivariate partial least squares （PLS） calibration models were established to correlate the process FTIR spectra with the concentrations of key metabolites and protein yield of in-process samples, either individually for each metabolite and protein or globally for all four metabolites simultaneously. Applying the 2＇ld derivative pre-processing algorithm to the FTIR spectra helps to reduce the number of PLS latent variables needed significantly and thus simplify the interpretation of the PLS models. The validated PLS models show promise in predicting the concentration profiles of glucose, glutamine, lactate, and ammonia and protein yield over the course of the bioreactor cell culture process. Therefore, this work demonstrated the technical feasibility of real time monitoring of the bioreactor cell culture process via FTIR spectroscopy. Its implications for enabling cell culture PAT were discussed.展开更多
文摘A simple efficient isocratic reversed-phase HPLC method was developed and validated for the determination of clindamycin palmitate hydrochloride (CPH) and its commercially available oral solution products. Separation was achieved on a Phenomenex Zorbax (Luna) cyano column (150 × 4.6 mm, 5 μm) with a Phenomenex cyano guard cartridge (4 × 3.0 mm) on Agilent 1050 series HPLC system. CPH and its resolution standard lincomycin were eluted isocratically at a flow rate of 1 mL/min with a simplified mobile phase (potassium phosphate buffer (5 mM, pH 3.0)—acetonitrile—tetrahydrofuran (20:75:5, v/v/v)) and detected at 210 nm. The column was maintained at 25?C. The method was validated according to USP category I requirements. Robustness and forced degradation studies were also conducted. CPH marketed drug products were obtained from a drug distributor and assayed for potency using the validated method. Validation acceptance criteria were met in all cases. The analytical range for CPH was 15 - 500 μg/mL and the linearity was r2 > 0.999 over three days. The method was determined to be specific and robust. Both accuracy (92.0% - 103.8%) and precision (0.67% - 1.52%) were established across the analytical range for low, intermediate and high QC concentrations. Method applicability was demonstrated by analyzing two marketed products of CPH, in which results showed potency >98%. The method was determined to be an enhancement over the current USP methodology for assay as a result of increased efficiency, reduced organic solvents and the elimination of matrix modifiers. This method was successfully applied for the quality assessment of: 1) currently marketed drug products and 2) will in future assess the product quality of novel dosage forms of CPH for pediatric use.
文摘The mechanism of degradation of dantrolene in aqueous buffer solutions was studied at various pH values in the range of pH 1.2-9.5 and at temperatures ranging from 25℃ to 75℃ to determine the optimum pH and temperature requirements for its stability and eventual product performance over the human gastrointestinal pH range. Dantrolene was analyzed by reversed phase ultra-performance liquid chromatographic (UPLC). Chromatographic separation was achieved on a Waters Acquity UPLC system using a Waters BEH C18 analytical column and Waters BEH C18 guard column. The compounds were eluted with a linear acetonitrile gradient (25%-75%) over three minutes with a buffer composition of 2.0 mM of sodium acetate at pH 4.5 for degradation studies. The flow rate was maintained at 0.5 mL/min. Column temperature was maintained at 35℃. Injection volume was 4 μL and the degradation products were detected by a photodiode array (PDA) detector at 375 nm. Degradation products, including compound B and C were analyzed by mass spectroscopy (MS) and nuclear magnetic resonance spectroscopy (NMR) and the degradation pathways were proposed. Degradation of dantrolene followed pseudo first–order kinetics and a V-shaped pH-rate profile over the pH range 1.2-9.5. The maximum stability was observed at pH 7.4 and 37℃. Although the focus of this paper was on the mechanism of hydrolysis of dantrolene, the poor aqueous solubility of dantrolene, the developed understanding can be utilized to improve the quality of the formulation and the risk associated with the extravasation of dantrolene sodium solution in its current form.
文摘The purpose of this study was to evaluate the impact of polystyrene divinylbenzene copolymer HPLC columns on the chromatographic performance of the USP compendial method for doxycycline hyclate. The compendial method was implemented based on the assessment of the chromatographic performance of six USP defined L21 polystyrene divinylbenzene HPLC columns. Modifications to the method were based on USP for chromatography. The method was validated for the determination of doxycycline hyclate and its impurities in commercially available drug products. A number of different polystyrene-divinylbenzene columns were tested and failed to provide selectivity for the resolution of doxycycline and its impurities. Separation was optimally achieved on an Agilent PLPR-S column (250 × 4.6 mm, 8 μm) by using an Agilent 1260 series HPLC system. Doxycycline hyclate and its impurities were eluted isocratically at a flow rate of 1 mL/min with mobile phase and detected at 270 nm. The column temperature was maintained at 60oC. The method was validated according to USP category I requirements for Assay. Validation acceptance criteria were met in all cases. The analytical range for doxycycline hyclate was 50 - 250 μg/mL and the linearity was r2 > 0.999 over three days. The method was determined to be specific. Both accuracy (95.1% - 102.4%) and precision (0.50% - 4.8%) were established across the analytical range for low, intermediate and high QC concentrations. Method applicability was demonstrated by analyzing marketed products of doxycycline hyclate, in which results showed potency meeting USP acceptance criteria. In conclusion, this study described the remarkable differences in selectivity that were encountered during the implementation phase for the compendial methods for doxycycline and its impurities in marketed products and it could be used in the future to assss the product quality of doxycycline hyclate capsules stored in the National stockpiles.
文摘The development and implementation of advanced analytical technologies is essential for extending the expiry for complex drug products stored in the Strategic National Stockpiles. Consequently, a novel Ultra High-Performance Liquid Chromatographic (UHPLC) method has been developed for the analysis of atropine and its respective impurities to support the analytical research platform for auto-injectors. This study is part of a larger research effort to improve the efficiency and broaden the applicability of advanced analytical methods for medical counter-measure medications. The current HPLC compendial methodology for atropine sulfate injection requires an analysis time of 40 minutes for atropine. In comparison, the novel gradient UHPLC method required only 8 minutes to evaluate both atropine and its major pharmaceutical impurities. Improved separation was achieved on a Waters Acquity UHPLC BEH C18 1.7 μm, 2.1 × 100 mm column employing gradient elution of mobile phase solvent A (0.1% H3PO4) and solvent B (0.1% H3PO4, 90% ACN, and 10% H2O). The method was validated according to USP Category I requirements for assay. The daily standard calibration curves were linear over a concentration range from 50 μg/mL to 250 μg/mL with a correlation coefficient of >0.999. The detection limit (LOD) and quantitation limit (LOQ) were 3.9 μg/ml and 13.1 μg/ml, respectively. Resolution results indicate that atropine and the following impurities, degradants and a preservative can also be separated and analyzed using this proposed method: noratropine, 4,4’-di-hy-droxydiphenyl ether, 2,4’-dihydroxydiphenyl ether, 4-bromophenol, 4-hydro-xyatropine, tropic acid, apoatropine HCl, atropic acid, hydroquinone, nitroethane, phenol and catechol. The UHPLC method demonstrated enhanced selectivity and significantly reduced the analysis time when compared with the traditional USP compendial HPLC method. The method was successfully applied to the evaluation of atropine in ATNAA auto-injectors lots from the Strategic National Stockpiles.
文摘Compared to small molecule process analytical technology （PAT） applications, biotechnology product PAT applications have certain unique challenges and opportunities. Understanding process dynamics of bioreactor cell culture process is essential to establish an appropriate process control strategy for biotechnology product PAT applications. Inline spectroscopic techniques for real time monitoring of bioreactor cell culture process have the distinct potential to develop PAT approaches in manufac- turing biotechnology drug products. However, the use of inline Fourier transform infrared （FTIR） spectroscopic techniques for bioreactor cell culture process monitoring has not been reported. In this work, real time inline FTIR Spectroscopy was applied to a lab scale bioreactor mAb IgG3 cell culture fluid biomolecular dynamic model. The technical feasibility of using FTIR Spectroscopy for real time tracking and monitoring four key cell culture metabolites （including glucose, glutamine, lactate, and ammonia） and protein yield at increasing levels of complexity （simple binary system, fully formulated media, actual bioreactor cell culture process） was evaluated via a stepwise approach. The FTIR fingerprints of the key metabolites were identified. The multivariate partial least squares （PLS） calibration models were established to correlate the process FTIR spectra with the concentrations of key metabolites and protein yield of in-process samples, either individually for each metabolite and protein or globally for all four metabolites simultaneously. Applying the 2＇ld derivative pre-processing algorithm to the FTIR spectra helps to reduce the number of PLS latent variables needed significantly and thus simplify the interpretation of the PLS models. The validated PLS models show promise in predicting the concentration profiles of glucose, glutamine, lactate, and ammonia and protein yield over the course of the bioreactor cell culture process. Therefore, this work demonstrated the technical feasibility of real time monitoring of the bioreactor cell culture process via FTIR spectroscopy. Its implications for enabling cell culture PAT were discussed.
