A new reversed-phase high performance liquid chromatography method was developed to quantitate the activity of xanthine oxidase involved in milk fat globule membrane with xanthine as the substrate and the separation o...A new reversed-phase high performance liquid chromatography method was developed to quantitate the activity of xanthine oxidase involved in milk fat globule membrane with xanthine as the substrate and the separation of product (uric acid). The increment of uric acid in the reaction system was used to calculate the total activity of XO. The optimized assay conditions, linearity of detection, recovery of uric acid and chromatogram were developed in text, indicating this method is simple, rapid and efficient. It is an alternative potential method for the determination of the activity of XO in milk.展开更多
A rapid, fast and precise method has been developed and validated for the simultaneous determination of amlodipine with H1-receptor antagonists (cetirizine, fexofenadine, and buclizine) from dosage forms. The chromato...A rapid, fast and precise method has been developed and validated for the simultaneous determination of amlodipine with H1-receptor antagonists (cetirizine, fexofenadine, and buclizine) from dosage forms. The chromatography was performed on a Purospher? Star, C18 (5 mm, 250 × 4.6 mm) column using acetonitrile: buffer (0.01 mM) (40:60, v/v, pH adjusted to 3.0), as a mobile phase. The mobile phase was pumped at a flow rate of 1.0 mL·min-1 and UV detection was performed at 240 nm. The method was validated for linearity, accuracy, precision and specificity. The method was applied to study the interaction between amlodipine and H1-receptor antagonists. These interactions were carried out in simulated gastric juice (pH 1), simulated full stomach (pH 4), blood pH (pH 7.4) and simulating GI (pH 9). The interacting drugs were heated at 37℃ with intermit-tent shaking and the samples were withdrawn every thirty minutes for three hours and drug contents were analyzed by RP-HPLC techniques. In most cases the in vitro availability of amlodipine was decreased. It was observed that the change in in vitro availability was pH dependent.展开更多
AIM:To determine moxonidine in aqueous humor and iris-ciliary body by reversed-phase high performance liquid chromatography(RP-HPLC),and to evaluate the retinal neuroprotective effect after topical administration with...AIM:To determine moxonidine in aqueous humor and iris-ciliary body by reversed-phase high performance liquid chromatography(RP-HPLC),and to evaluate the retinal neuroprotective effect after topical administration with moxonidine in a high intraocular pressure(IOP)model.METHODS:The eyes of albino rabbits were administered topically and ipsilaterally with 0.2%moxonidine.A RPHPLC method was employed for the identification and quantification of moxonidine between 2 and 480 min,which presented in the aqueous humor and iris-ciliary body.Flash electroretinography(F-ERG)amplitude and superoxide dismutase(SOD)level were measured between day 1 and day 15 after topical administration with moxonidine in a rabbit model of high IOP.Histological and ultrastructural observation underwent to analyze the changes of retinal morphology,the inner retinal layers(IRL)thickness,and retinal ganglion cell(RGC)counting.RESULTS:Moxonidine was detectable between 2 and 480 min after administration,and the peak concentration developed both in the two tissues at 30 min,0.51μg/m Lin aqueous humor and 1.03μg/g in iris-ciliary body.In comparison to control,F-ERG b-wave amplitude in moxonidine eyes were significantly differences between day 3 and day 15(P<0.01)in the high IOP model;SOD levels were significantly higher at all time-points(P<0.01)with a maximum level of 20.29 U/mgprot at day 15;and RGCs were significantly higher(P<0.05).CONCLUSION:Moxonidine is a viable neuroprotective agent with application to high IOP model.All layers of retina,including RGC layer,retinal nerve fiber layer and INL,are more preserved after moxonidine administration.SOD plays a neuroprotective role in ocular hypertension-mediated RGC death.展开更多
Objective: A highly sensitive SPE-liquid/liquid extraction RPLC method has been developed for the analysis of 6β-hydroxycortisol and cortisol in the urine of cancer patients. Methods: After SPE column purification an...Objective: A highly sensitive SPE-liquid/liquid extraction RPLC method has been developed for the analysis of 6β-hydroxycortisol and cortisol in the urine of cancer patients. Methods: After SPE column purification and liquid-liquid extraction, the sample test solutions were analyzed with RPLC using a C18 analytical column. This improved analytical method has been validated for linearity, accuracy (recovery from urine), repeatability (within-day and between-day precision), specificity, sensitivity, and stability. This SPE-liquid/liquid extraction-RPLC is rapid, simple, accurate and reproducible. The technique is particularly useful for monitoring the CYP3A activity of cancer patients in clinical settings. The results are expressed as the ratio of 6β-hydroxycortisol to cortisol. Results: The CYP3A activity from a total of 153 samples was measured using this improved method. Considerable variation in the CYP3A activity of different cancer patients has been documented. Thus, personalized medical treatment based on the individual metabolic enzyme activity level is necessary. Conclusion: This new analytical method facilitates such individualized medical treatments.展开更多
The research was carried out for establishing a new reverse phase-HPLC stability indicating method for the quantification of Rucaparib. The experiment was determined on Waters HPLC instrument using 996 photo-diode arr...The research was carried out for establishing a new reverse phase-HPLC stability indicating method for the quantification of Rucaparib. The experiment was determined on Waters HPLC instrument using 996 photo-diode array detector. The separation was done by using symmetry C-18 ODS (25 cm × 0.46 cm internal diameter) 5 μm analytical column containing mobile phase of Phosphate buffer (0.02 M) and methanol [65:35% v/v] adjusted pH to 4.8 by adding dilute ortho phosphoric acid. The method was run at 1 ml·min<sup>-1</sup> at 286 nm detection. The drug was eluted at 5.484 min. After developing the method, it was assured for the intended use by validation which was done according to ICH Q2B guidelines. The analytical parameters checked were linearity, accuracy, repeatability, intermediate precision, limit of detection, limit of quantitation, ruggedness and robustness. It was observed that the response of the detector was linear in the range of 6 - 14 μg/ml with correlation coefficient of 0.999. The results of all the parameters were found to be within the acceptance criteria. The stability indicating assay method was established by using the samples generated by forced degradation process. The forced degradation was carried out by subjecting the drug to acid, alkali, thermal, oxidative and photolytic degradation and the results showed that the degradation products were successfully separated from the drug. Hence, this can be applied perfectly later for the analysis of quality of the rucaparib drug.展开更多
The research was carried out to establish a new reverse phase-HPLC stability indicating method for quantifying Bimatoprost & Timolol in ophthalmic solution. The experiment of Bimatoprost & Timolol in ophthalmi...The research was carried out to establish a new reverse phase-HPLC stability indicating method for quantifying Bimatoprost & Timolol in ophthalmic solution. The experiment of Bimatoprost & Timolol in ophthalmic solution method development was determined on Waters HPLC instrument using a UV Detector. The separation was done by using L11, Zorbex SB phenyl (4.6 mm × 250 mm internal diameter) 5 μm analytical column, containing mobile phase of Phosphate buffer (0.02 M), methanol, and acetonitrile [50:30:20 % v/v]. The method was run at 1 ml·min<sup>-1</sup> at 210 nm for Bimatoprost and 295 nm for Timolol for detection. The drug was eluted at 10.81 min for Bimatoprost and 3.77 min for Timolol. After developing the method, it was assured for the intended use by validation, which was done according to ICH Q2B guidelines. The analytical parameters checked were Specificity/Selectivity, linearity, Range, accuracy, ruggedness, and robustness. It was observed that the response of the detector was linear in the range of 6 - 18 μg/ml with a correlation coefficient of 0.999. The results of all the parameters were found to be within the acceptance criteria. The stability-indicating assay method was established by using the samples generated by the forced degradation process. The forced degradation was carried out by subjecting the drug to acid, alkali, thermal, oxidative, and photolytic degradation, and the results showed that the degradation products were successfully separated from the drug. Hence, this can be applied perfectly later for the quantitative analysis of Bimatoprost 0.3% + Timolol 0.5% Ophthalmic Solution drugs for pharmaceutical use. Currently, there is no official method for Bimatoprost & Timolol combination products in USP or BP. Available research work related to single Bimatoprost or Timolol products was not suitable for testing Bimatoprost and Timolol combination drugs. Additionally, there is no stability-indicating method to test Bimatoprost & Timolol combination products which insist us to do research and develop a new reverse phase-HPLC indicating method which will be faster and more accurate.展开更多
The interaction index c which was derived from the fundamental retention equationlogk’ =a + cC_B in reversed--phase high--performancc liquid chromatography (RP-HPLC) quan-titatively describes the difference in the in...The interaction index c which was derived from the fundamental retention equationlogk’ =a + cC_B in reversed--phase high--performancc liquid chromatography (RP-HPLC) quan-titatively describes the difference in the interaction between solute--strong. solvent andsolute--weak solvent; it has shown to be a constant for a specific solute even when columnsystems with different C18 packings are used. The theoretical basis for peak identificationby using interaction index has been proposed, which was based on the a,c values on stan-dard C18 column by utilizing linear a-a plots on column pairs and the linear relationship be-tween parameters a and c for the structural related compounds. Through the establishmentof parameters a,c data based on the standard C18 column for a certain type of compounds,the retention of thesc compounds on various C18 columns can be predicted. Typical exam-ples have been given to verify the correctness of this method.展开更多
In this work, we developed a highly sensitive method to detect melamine based on reversed phase liquid chromatography mass spectrometry. A mass spectrometry compatible ion pair, heptafluorobu-tyric acid(HFBA), was use...In this work, we developed a highly sensitive method to detect melamine based on reversed phase liquid chromatography mass spectrometry. A mass spectrometry compatible ion pair, heptafluorobu-tyric acid(HFBA), was used to separate melamine by reversed phase liquid chromatography prior to electrospray mass spectrometry. The incorporation of isotope internal standard and multiple reaction monitoring improved the accuracy and linearity of quantification. Based on this strategy, the method limit of quantification was 0.1 ng/g. The limits of quantification were 8 ng/g for liquid milk and 15 ng/g for dry milk powder. This method provided a reproducible and stable approach to sensitive detection and quantification of melamine.展开更多
With insulin methanol water, and the ion pairing agent, hydrochloric acid and trifluroacetic acid (TFA), the character of the first plateau (FP) on the elution curve of frontal analysis in reversed phase liquid chro...With insulin methanol water, and the ion pairing agent, hydrochloric acid and trifluroacetic acid (TFA), the character of the first plateau (FP) on the elution curve of frontal analysis in reversed phase liquid chromatography (RPLC) was investigated by on line UV spectrometry and identified with nuclear magnetic resonance (NMR) spectrometry and mass spectrometry. The profile of the FP is the same as that of a usual elution curve of methanol in frontal analysis (FA). When the insulin concentration was limited to a certain range, the height of the FP was found to be proportional to the insulin concentration in mobile phase and its length companying to shorten. The FP profile on the intersection of two tangents reflects the components of the microstructure in the depth direction of the bonded stationary phase layer and the desorption dynamics of the displaced components. The displaced methanol was quantitatively determined by NMR and on line UV spectrometries. TFA with high UV absorbance can not be used as an ion pairing agent for the investigation of the FP in RPLC, but it can be used as a good marker to investigate the complicated transfer process of components in the stationary phase in RPLC. A stoichiometric displacement process between solute and solvent was proved to be valid in both usual and FA in RPLC. From the point of view of dynamics of mass transfer, the solutes can only contact to the surface of stationary phase in usual RPLC, while solute can penetrate into it in FA of RPLC. The solvation of insulin in methanol and water solution as an example indicating the usage of the FP in the FA was also investigated in this paper.展开更多
Rapid and sensitive reversed phase high performance liquid chromatography (RP-HPLC) and ultra performance liquid chromatography (RP-UPLC) method with UV detection has been developed and validated for quantification of...Rapid and sensitive reversed phase high performance liquid chromatography (RP-HPLC) and ultra performance liquid chromatography (RP-UPLC) method with UV detection has been developed and validated for quantification of parathyroid hormone (PTH) in presence of meta-cresol as a stabilizer in a pharmaceutical formulation.Chromatography was performed with mobile phase containing 0.1% Trifluoroacetic acid (TFA) in MilliQ water and 0.1% TFA in acetonitrile with gradient program and flow rate at 0.3 mL/min for HPLC and 0.4 mL/min for UPLC.Quantification was accomplished with internal reference standard (qualified against innovator product and National Institute for Biological Standards and Control (NIBSC) standard).The methods were validated for linearity (correlation coefficient 0.99),range,accuracy,precision and robustness.Robustness was confirmed by considering three factors;mobile phase composition,column temperature and flow rate/age of mobile phase.Intermediate precision was confirmed on different equipments,different columns and on different days.The relative standard deviation (RSD) (<2% for RP-HPLC and <1% for UPLC,n=30) indicated a good precision.Retention time was found about 17 min and 2 min by HPLC and UPLC methods,respectively.Both methods are simple,highly sensitive,precise and accurate and have the potential of being useful for routine quality control.展开更多
A simple, sensitive, and specific method was developed for simultaneous determination of Amlodipine besylate (AML), Valsartan (Vals) and Hydrochlorothiazide (HCT) by high performance liquid chromatography without prev...A simple, sensitive, and specific method was developed for simultaneous determination of Amlodipine besylate (AML), Valsartan (Vals) and Hydrochlorothiazide (HCT) by high performance liquid chromatography without previous separation. Satisfactory resolution was achieved using a RP-C18 chromatographic column, Phenomenex Kinetex (150 mm × 4.6 mm i.d) and a mobile phase consisting of acetonitrile-phosphate buffer (0.05 M) with pH 2.8 in the proportion of (40/60, v/v) at a flow rate 0.8 mL/min and the wavelength detection was 227 nm. The retention time for HCT, AML and VAls was 2.26, 3.16 and 11.19 min;respectively. The described method was linear over a range of 4-28 μg /ml, 5-40 μg /ml and 1-12 μg /ml for AML, Vals and HCT;respectively. The mean percent recoveries were 99.94%, 99.96% and 99.78% for AML, Vals and HCT;respectively. F-test and t-test at 95%con?dence level were used to check the intermediate precision data obtained under different experimental setups. The method could be used for analysis of combined dose tablet formulation containing AML, Vals, HCT as well as spiked human plasma.展开更多
To identify acute renal allograft rejection biomarkers in human serum, two-dimensional differential in-gel electrophoresis (2-D DIGE) and reversed phase high-performance liquid chromatography (RP-HPLC) followed by...To identify acute renal allograft rejection biomarkers in human serum, two-dimensional differential in-gel electrophoresis (2-D DIGE) and reversed phase high-performance liquid chromatography (RP-HPLC) followed by electrospray ionization mass spectrometry (ESI-MS) were used. Serum samples from renal allograft patients and normal volunteers were divided into three groups: acute rejec- tion (AR), stable renal function (SRF) and normal volunteer (N). Serum samples were firstly processed using Multiple Affinity Removal Column to selectively remove the highest abundance proteins. Differentially expressed proteins were analyzed using 2-D DIGE. These differential protein spots were excised, digested by trypsin, and identified by RP-HPLC-ESI/MS. Twenty-two differentially expressed proteins were identified in serum from AR group. These proteins included complement C9 precursor, apolipoprotein A-IV precursor, vitamin D-binding protein precursor, beta-2-glycoprotein 1 precursor, etc. Vitamin D-binding protein, one of these proteins, was confirmed by ELISA in the independent set of serum samples. In conclusion, the differentially expressed proteins as serum biomarker candidates may provide the basis of acute rejection noninvasive diagnosis. Confirmed vitamin D-binding protein may be one of serum biomarkers of acute rejection. Furthermore, it may provide great insights into understanding the mechanisms and potential treatment strategy of acute rejection.展开更多
Present study was aimed to develop and validate a reverse-phase high-performance liquid chromatography method for simultaneous determination of sitagliptin phosphate and metformin hy-drochloride in a marketed formulat...Present study was aimed to develop and validate a reverse-phase high-performance liquid chromatography method for simultaneous determination of sitagliptin phosphate and metformin hy-drochloride in a marketed formulation. The drug separation was performed on Hibar-240, Li-chrosphere-100 C18 ODS (250 × 4.6 mm, 5 μm) column, at a flow rate of 1 mL/min. The mobile phase used was a mixture of methanol: potassium di-hydrogen phosphate buffer at a ratio of 70:30 v/v. The detection was carried out at a wavelength of 266 nm. The retention times of sitagliptin phosphate and metformin hydrochloride were found as 6.1 and 4.9 min respectively. Linear calibration curves with good correlation coefficients were obtained over the concentration ranges of 10 - 50 μg/mL for sitagliptin and 20 - 100 μg/mL for metformin. The limit of detection was 0.016 and 0.14 μg/mL and the limit of quantification was 0.048 and 0.42 μg/mL for sitagliptin phosphate and metformin hydrochloride respectively. Validation of the method demonstrated system selectivity, specificity, linearity, accuracy and precision. The developed method was found useful in the simultaneous analysis of sitagliptin phosphate and metformin hydrochloride in formulation.展开更多
[ Objective] The aim was to establish effective method for endogenous hormone extraction and explore conditions of chromatographic analysis for three endogenous hormones in rhizome of Alhagi sparsifolia. [ Methed ] Ac...[ Objective] The aim was to establish effective method for endogenous hormone extraction and explore conditions of chromatographic analysis for three endogenous hormones in rhizome of Alhagi sparsifolia. [ Methed ] Activol (GA3 ), zeatin (ZR) and indole-3-acetic acid (IAA) in rhizome were separated and measured as per RP-HPLC. [Result] The average recovery rates of GA3, ZR and IAA were 98.3%, 90.3% and 101.3%, respectively, indicating that the method is suitable for quantitative analysis with little errors. The chromatographic conditions were as follows: methanol/0. 75% of acetic acid at 45:55 (mobile phase) ; flow speed at 0.7 ml/min; wavelength at 254 nm; column temperature at 25 ℃. [Conclusion] The research preliminarily established HPLC conditions for separation of endogenous hormones in rhizome of Alhagi sparsifolia.展开更多
文摘A new reversed-phase high performance liquid chromatography method was developed to quantitate the activity of xanthine oxidase involved in milk fat globule membrane with xanthine as the substrate and the separation of product (uric acid). The increment of uric acid in the reaction system was used to calculate the total activity of XO. The optimized assay conditions, linearity of detection, recovery of uric acid and chromatogram were developed in text, indicating this method is simple, rapid and efficient. It is an alternative potential method for the determination of the activity of XO in milk.
文摘A rapid, fast and precise method has been developed and validated for the simultaneous determination of amlodipine with H1-receptor antagonists (cetirizine, fexofenadine, and buclizine) from dosage forms. The chromatography was performed on a Purospher? Star, C18 (5 mm, 250 × 4.6 mm) column using acetonitrile: buffer (0.01 mM) (40:60, v/v, pH adjusted to 3.0), as a mobile phase. The mobile phase was pumped at a flow rate of 1.0 mL·min-1 and UV detection was performed at 240 nm. The method was validated for linearity, accuracy, precision and specificity. The method was applied to study the interaction between amlodipine and H1-receptor antagonists. These interactions were carried out in simulated gastric juice (pH 1), simulated full stomach (pH 4), blood pH (pH 7.4) and simulating GI (pH 9). The interacting drugs were heated at 37℃ with intermit-tent shaking and the samples were withdrawn every thirty minutes for three hours and drug contents were analyzed by RP-HPLC techniques. In most cases the in vitro availability of amlodipine was decreased. It was observed that the change in in vitro availability was pH dependent.
基金Supported by the Key Science and Technology Program of Shaanxi Province,China(No.2015SF146).
文摘AIM:To determine moxonidine in aqueous humor and iris-ciliary body by reversed-phase high performance liquid chromatography(RP-HPLC),and to evaluate the retinal neuroprotective effect after topical administration with moxonidine in a high intraocular pressure(IOP)model.METHODS:The eyes of albino rabbits were administered topically and ipsilaterally with 0.2%moxonidine.A RPHPLC method was employed for the identification and quantification of moxonidine between 2 and 480 min,which presented in the aqueous humor and iris-ciliary body.Flash electroretinography(F-ERG)amplitude and superoxide dismutase(SOD)level were measured between day 1 and day 15 after topical administration with moxonidine in a rabbit model of high IOP.Histological and ultrastructural observation underwent to analyze the changes of retinal morphology,the inner retinal layers(IRL)thickness,and retinal ganglion cell(RGC)counting.RESULTS:Moxonidine was detectable between 2 and 480 min after administration,and the peak concentration developed both in the two tissues at 30 min,0.51μg/m Lin aqueous humor and 1.03μg/g in iris-ciliary body.In comparison to control,F-ERG b-wave amplitude in moxonidine eyes were significantly differences between day 3 and day 15(P<0.01)in the high IOP model;SOD levels were significantly higher at all time-points(P<0.01)with a maximum level of 20.29 U/mgprot at day 15;and RGCs were significantly higher(P<0.05).CONCLUSION:Moxonidine is a viable neuroprotective agent with application to high IOP model.All layers of retina,including RGC layer,retinal nerve fiber layer and INL,are more preserved after moxonidine administration.SOD plays a neuroprotective role in ocular hypertension-mediated RGC death.
基金supported by the Shanghai Pharmaceutical Association
文摘Objective: A highly sensitive SPE-liquid/liquid extraction RPLC method has been developed for the analysis of 6β-hydroxycortisol and cortisol in the urine of cancer patients. Methods: After SPE column purification and liquid-liquid extraction, the sample test solutions were analyzed with RPLC using a C18 analytical column. This improved analytical method has been validated for linearity, accuracy (recovery from urine), repeatability (within-day and between-day precision), specificity, sensitivity, and stability. This SPE-liquid/liquid extraction-RPLC is rapid, simple, accurate and reproducible. The technique is particularly useful for monitoring the CYP3A activity of cancer patients in clinical settings. The results are expressed as the ratio of 6β-hydroxycortisol to cortisol. Results: The CYP3A activity from a total of 153 samples was measured using this improved method. Considerable variation in the CYP3A activity of different cancer patients has been documented. Thus, personalized medical treatment based on the individual metabolic enzyme activity level is necessary. Conclusion: This new analytical method facilitates such individualized medical treatments.
文摘The research was carried out for establishing a new reverse phase-HPLC stability indicating method for the quantification of Rucaparib. The experiment was determined on Waters HPLC instrument using 996 photo-diode array detector. The separation was done by using symmetry C-18 ODS (25 cm × 0.46 cm internal diameter) 5 μm analytical column containing mobile phase of Phosphate buffer (0.02 M) and methanol [65:35% v/v] adjusted pH to 4.8 by adding dilute ortho phosphoric acid. The method was run at 1 ml·min<sup>-1</sup> at 286 nm detection. The drug was eluted at 5.484 min. After developing the method, it was assured for the intended use by validation which was done according to ICH Q2B guidelines. The analytical parameters checked were linearity, accuracy, repeatability, intermediate precision, limit of detection, limit of quantitation, ruggedness and robustness. It was observed that the response of the detector was linear in the range of 6 - 14 μg/ml with correlation coefficient of 0.999. The results of all the parameters were found to be within the acceptance criteria. The stability indicating assay method was established by using the samples generated by forced degradation process. The forced degradation was carried out by subjecting the drug to acid, alkali, thermal, oxidative and photolytic degradation and the results showed that the degradation products were successfully separated from the drug. Hence, this can be applied perfectly later for the analysis of quality of the rucaparib drug.
文摘The research was carried out to establish a new reverse phase-HPLC stability indicating method for quantifying Bimatoprost & Timolol in ophthalmic solution. The experiment of Bimatoprost & Timolol in ophthalmic solution method development was determined on Waters HPLC instrument using a UV Detector. The separation was done by using L11, Zorbex SB phenyl (4.6 mm × 250 mm internal diameter) 5 μm analytical column, containing mobile phase of Phosphate buffer (0.02 M), methanol, and acetonitrile [50:30:20 % v/v]. The method was run at 1 ml·min<sup>-1</sup> at 210 nm for Bimatoprost and 295 nm for Timolol for detection. The drug was eluted at 10.81 min for Bimatoprost and 3.77 min for Timolol. After developing the method, it was assured for the intended use by validation, which was done according to ICH Q2B guidelines. The analytical parameters checked were Specificity/Selectivity, linearity, Range, accuracy, ruggedness, and robustness. It was observed that the response of the detector was linear in the range of 6 - 18 μg/ml with a correlation coefficient of 0.999. The results of all the parameters were found to be within the acceptance criteria. The stability-indicating assay method was established by using the samples generated by the forced degradation process. The forced degradation was carried out by subjecting the drug to acid, alkali, thermal, oxidative, and photolytic degradation, and the results showed that the degradation products were successfully separated from the drug. Hence, this can be applied perfectly later for the quantitative analysis of Bimatoprost 0.3% + Timolol 0.5% Ophthalmic Solution drugs for pharmaceutical use. Currently, there is no official method for Bimatoprost & Timolol combination products in USP or BP. Available research work related to single Bimatoprost or Timolol products was not suitable for testing Bimatoprost and Timolol combination drugs. Additionally, there is no stability-indicating method to test Bimatoprost & Timolol combination products which insist us to do research and develop a new reverse phase-HPLC indicating method which will be faster and more accurate.
文摘The interaction index c which was derived from the fundamental retention equationlogk’ =a + cC_B in reversed--phase high--performancc liquid chromatography (RP-HPLC) quan-titatively describes the difference in the interaction between solute--strong. solvent andsolute--weak solvent; it has shown to be a constant for a specific solute even when columnsystems with different C18 packings are used. The theoretical basis for peak identificationby using interaction index has been proposed, which was based on the a,c values on stan-dard C18 column by utilizing linear a-a plots on column pairs and the linear relationship be-tween parameters a and c for the structural related compounds. Through the establishmentof parameters a,c data based on the standard C18 column for a certain type of compounds,the retention of thesc compounds on various C18 columns can be predicted. Typical exam-ples have been given to verify the correctness of this method.
基金Supported by the National Natural Science Foundation of China (Grant No. 30425021)National High Technology Research and Development Program of China (Grant No. 2007AA02Z334)
文摘In this work, we developed a highly sensitive method to detect melamine based on reversed phase liquid chromatography mass spectrometry. A mass spectrometry compatible ion pair, heptafluorobu-tyric acid(HFBA), was used to separate melamine by reversed phase liquid chromatography prior to electrospray mass spectrometry. The incorporation of isotope internal standard and multiple reaction monitoring improved the accuracy and linearity of quantification. Based on this strategy, the method limit of quantification was 0.1 ng/g. The limits of quantification were 8 ng/g for liquid milk and 15 ng/g for dry milk powder. This method provided a reproducible and stable approach to sensitive detection and quantification of melamine.
文摘With insulin methanol water, and the ion pairing agent, hydrochloric acid and trifluroacetic acid (TFA), the character of the first plateau (FP) on the elution curve of frontal analysis in reversed phase liquid chromatography (RPLC) was investigated by on line UV spectrometry and identified with nuclear magnetic resonance (NMR) spectrometry and mass spectrometry. The profile of the FP is the same as that of a usual elution curve of methanol in frontal analysis (FA). When the insulin concentration was limited to a certain range, the height of the FP was found to be proportional to the insulin concentration in mobile phase and its length companying to shorten. The FP profile on the intersection of two tangents reflects the components of the microstructure in the depth direction of the bonded stationary phase layer and the desorption dynamics of the displaced components. The displaced methanol was quantitatively determined by NMR and on line UV spectrometries. TFA with high UV absorbance can not be used as an ion pairing agent for the investigation of the FP in RPLC, but it can be used as a good marker to investigate the complicated transfer process of components in the stationary phase in RPLC. A stoichiometric displacement process between solute and solvent was proved to be valid in both usual and FA in RPLC. From the point of view of dynamics of mass transfer, the solutes can only contact to the surface of stationary phase in usual RPLC, while solute can penetrate into it in FA of RPLC. The solvation of insulin in methanol and water solution as an example indicating the usage of the FP in the FA was also investigated in this paper.
文摘Rapid and sensitive reversed phase high performance liquid chromatography (RP-HPLC) and ultra performance liquid chromatography (RP-UPLC) method with UV detection has been developed and validated for quantification of parathyroid hormone (PTH) in presence of meta-cresol as a stabilizer in a pharmaceutical formulation.Chromatography was performed with mobile phase containing 0.1% Trifluoroacetic acid (TFA) in MilliQ water and 0.1% TFA in acetonitrile with gradient program and flow rate at 0.3 mL/min for HPLC and 0.4 mL/min for UPLC.Quantification was accomplished with internal reference standard (qualified against innovator product and National Institute for Biological Standards and Control (NIBSC) standard).The methods were validated for linearity (correlation coefficient 0.99),range,accuracy,precision and robustness.Robustness was confirmed by considering three factors;mobile phase composition,column temperature and flow rate/age of mobile phase.Intermediate precision was confirmed on different equipments,different columns and on different days.The relative standard deviation (RSD) (<2% for RP-HPLC and <1% for UPLC,n=30) indicated a good precision.Retention time was found about 17 min and 2 min by HPLC and UPLC methods,respectively.Both methods are simple,highly sensitive,precise and accurate and have the potential of being useful for routine quality control.
文摘A simple, sensitive, and specific method was developed for simultaneous determination of Amlodipine besylate (AML), Valsartan (Vals) and Hydrochlorothiazide (HCT) by high performance liquid chromatography without previous separation. Satisfactory resolution was achieved using a RP-C18 chromatographic column, Phenomenex Kinetex (150 mm × 4.6 mm i.d) and a mobile phase consisting of acetonitrile-phosphate buffer (0.05 M) with pH 2.8 in the proportion of (40/60, v/v) at a flow rate 0.8 mL/min and the wavelength detection was 227 nm. The retention time for HCT, AML and VAls was 2.26, 3.16 and 11.19 min;respectively. The described method was linear over a range of 4-28 μg /ml, 5-40 μg /ml and 1-12 μg /ml for AML, Vals and HCT;respectively. The mean percent recoveries were 99.94%, 99.96% and 99.78% for AML, Vals and HCT;respectively. F-test and t-test at 95%con?dence level were used to check the intermediate precision data obtained under different experimental setups. The method could be used for analysis of combined dose tablet formulation containing AML, Vals, HCT as well as spiked human plasma.
基金supported by a grant from National Basic Research 973 Program of China (No.2009CB522407)
文摘To identify acute renal allograft rejection biomarkers in human serum, two-dimensional differential in-gel electrophoresis (2-D DIGE) and reversed phase high-performance liquid chromatography (RP-HPLC) followed by electrospray ionization mass spectrometry (ESI-MS) were used. Serum samples from renal allograft patients and normal volunteers were divided into three groups: acute rejec- tion (AR), stable renal function (SRF) and normal volunteer (N). Serum samples were firstly processed using Multiple Affinity Removal Column to selectively remove the highest abundance proteins. Differentially expressed proteins were analyzed using 2-D DIGE. These differential protein spots were excised, digested by trypsin, and identified by RP-HPLC-ESI/MS. Twenty-two differentially expressed proteins were identified in serum from AR group. These proteins included complement C9 precursor, apolipoprotein A-IV precursor, vitamin D-binding protein precursor, beta-2-glycoprotein 1 precursor, etc. Vitamin D-binding protein, one of these proteins, was confirmed by ELISA in the independent set of serum samples. In conclusion, the differentially expressed proteins as serum biomarker candidates may provide the basis of acute rejection noninvasive diagnosis. Confirmed vitamin D-binding protein may be one of serum biomarkers of acute rejection. Furthermore, it may provide great insights into understanding the mechanisms and potential treatment strategy of acute rejection.
文摘Present study was aimed to develop and validate a reverse-phase high-performance liquid chromatography method for simultaneous determination of sitagliptin phosphate and metformin hy-drochloride in a marketed formulation. The drug separation was performed on Hibar-240, Li-chrosphere-100 C18 ODS (250 × 4.6 mm, 5 μm) column, at a flow rate of 1 mL/min. The mobile phase used was a mixture of methanol: potassium di-hydrogen phosphate buffer at a ratio of 70:30 v/v. The detection was carried out at a wavelength of 266 nm. The retention times of sitagliptin phosphate and metformin hydrochloride were found as 6.1 and 4.9 min respectively. Linear calibration curves with good correlation coefficients were obtained over the concentration ranges of 10 - 50 μg/mL for sitagliptin and 20 - 100 μg/mL for metformin. The limit of detection was 0.016 and 0.14 μg/mL and the limit of quantification was 0.048 and 0.42 μg/mL for sitagliptin phosphate and metformin hydrochloride respectively. Validation of the method demonstrated system selectivity, specificity, linearity, accuracy and precision. The developed method was found useful in the simultaneous analysis of sitagliptin phosphate and metformin hydrochloride in formulation.
基金Initial Funding of Doctor in Xinjiang University(7020428027)National Innovation Experiment Program for University Students of China (101075512)
文摘[ Objective] The aim was to establish effective method for endogenous hormone extraction and explore conditions of chromatographic analysis for three endogenous hormones in rhizome of Alhagi sparsifolia. [ Methed ] Activol (GA3 ), zeatin (ZR) and indole-3-acetic acid (IAA) in rhizome were separated and measured as per RP-HPLC. [Result] The average recovery rates of GA3, ZR and IAA were 98.3%, 90.3% and 101.3%, respectively, indicating that the method is suitable for quantitative analysis with little errors. The chromatographic conditions were as follows: methanol/0. 75% of acetic acid at 45:55 (mobile phase) ; flow speed at 0.7 ml/min; wavelength at 254 nm; column temperature at 25 ℃. [Conclusion] The research preliminarily established HPLC conditions for separation of endogenous hormones in rhizome of Alhagi sparsifolia.