Application of derivative spectrophotometry for determination of enalapril, hydrochlorothiazide and walsartan in complex pharmaceutical preparations

A derivative spectrophotometry method was developed to determine enalapril, hydrochlorothiazide, candesartan and walsartan in complex antihypertensive drugs. The pharmaceutical preparations containing hydrochlorothiazide and one of the angiotensin convertase inhibitors were investigated. It was found that the developed method enables the constituents of the investigated drugs to be determined directly despite evident interference of the zero order absorption spectra. For determination of enalapril and hydrochlorothiazide as well as candesartan and hydrochlorothiazide the first derivative was used, while for walsartan and hydrochlorothiazide the second derivative was employed. The method was of high sensitivity; the LOD accuracy for enalapril was 2.81 microg x mL(-1), 0.56 microg x mL(-1) for candesartan, 4.02 microg x mL(-1) for walsartan and ranged from 0.31 microg x mL(-1) to 1.78 microgxmL(-1) for hydrochlorothiazide, depending on preparation under investigation. The recovery of individual constituents was within the limit of 100% +/- 5%, RSD varied from 1.11% to 2.94%, and the linearity range was from 4.1 microg x mL(-1) to 20.5 microg x mL(-1) for enalapril, from 6.45 microg x mL(-1) to 32.25 microg x mL(-1) for walsartan, from 2.36 microg x mL(-1) to 11.80 microg x mL(-1) for candesartan, and from 0.96 microg x mL(-1) to 26.00 microg x mL(-1) for hydrochlorothiazide.     

Development and validation of a HPLC and a UV derivative spectrophotometric methods for determination of hydroquinone in gel and cream preparations

A high performance liquid chromatographic (HPLC) and a ultraviolet derivative spectrophotometric (UVDS) methods were developed and validated for the quantitative determination of hydroquinone (HQ) in gels and creams containing this compound as a unique active principle. Validation parameters such as linearity, precision, accuracy, specificity, limit of detection (LOD) and limit of quantitation (LOQ) were determined. HPLC was carried out by reversed phase technique on a RP-18 column with a mobile phase composed of methanol and water (20:80, v/v). The linearity in the range of 6.0-30.0 microg/mL present a correlation coefficient (r) of 0.9999, calculated by least square method. The LOD and LOQ were 0.08 and 0.26 microg/mL, respectively. Based on the preliminary spectrophotometric profile of HQ, a signal at 302.0 nm of the first derivative spectrum (1D302.0) was found adequate for validation. The linearity between signal 1D302.0 and concentration of HQ in the range of 10.0-26.0 microg/mL in sulfuric acid (0.1N) present a correlation coefficient (r) of 0.9999. The LOD and LOQ were 0.14 and 0.46 microg/mL, respectively. Statistical analysis by t- and F-tests, showed no significant difference at 95% confidence level between the two proposed methods.     

Quantitative analysis of lovastatin in capsule of Chinese medicine monascus by capillary zone electrophoresis with UV-vis detector

A capillary zone electrophoresis (CZE) method was developed for the quantitative analysis of lovastatin (Lvt) in capsule of monascus-Chinese medicine. Lvt in the capsule was separated using an electrolyte system consisting of 16% ethanol (v/v) in 60 mM Gly-sodium hydroxide buffer, pH 10.5, 16 kV applied voltage, 238 nm detection wavelength with a capillary of 51 cm x 75 microm i.d (43 cm to detector). Under the optimized conditions, the linear response of Lvt concentration ranges from 4.0 to 240 microg/mL with high correlation coefficient (r=0.9998, n=9), the limits of detection (LOD) and quantification (LOQ) for Lvt are 0.73 and 2.42 microg/mL, the precision values (expressed as R.S.D.) of intra-day and inter-day are 1.40-2.12% and 1.47-3.88%, respectively. The recoveries of the analyte at three concentration levels are 90.28-100.71%. The developed method can be well used for the quantification of Lvt in the drug in commercial formulations.     

Total flavonoids quantification from O/W emulsion with extract of Brazilian plants

A new derivative spectrophotometric (DS) method was proposed and validated for quantification of total flavonoids from in O/W emulsion with polyacrylamide (and) C13-14 isoparaffin (and) laureth-7 containing Catuaba (Trichilia catigua Adr. Juss) (and) Marapuama (Ptychopetalum olacoides Bentham) extract. DS method was optimized to perform the assay in most favorable conditions. Linearity, specificity and selectivity, recovery (Rc, %), precision (R.S.D., %), accuracy (E, %), detection (LOD, microg ml(-1)) and quantification limits (LOQ, microg ml(-1)) were established for method validation. First-derivative at 388.0 nm (zero-to-peak; amplitude= +/- 0.12; wavelength range= 300.0-450.0 nm and Deltalambda = 4 nm) offered linearity for rutin concentrations ranging from 10.0 to 60.0 microg ml(-1) in ethanol 99.5%. Second-derivative provided to be unsuitable for interval evaluation obtaining unacceptable accuracy. Analytical method was validated for first-derivative, according to the experimental results: correlation coefficient (r = 0.9999); specificity to total flavonoids quantification, expressed in rutin, at wavelength 388.0 nm and selectivity with elimination of interference from matrix; Rc = 108.78%; intra- and inter-run precision (1.30-3.65% and 3.48-4.68%), and intra- and inter-run accuracy (100.00-112.19% and 101.25-118.44%); LOD = 0.62 microg ml(-1) and LOQ = 1.86 microg ml(-1).     

An efficient separation and method development for the quantifying of two basic impurities of Nicergoline by reversed-phase high performance liquid chromatography using ion-pairing counter ions

A quantification method was developed for the two basic impurities, one of which is also a metabolite, of Nicergoline (NIC), by using reversed-phase high performance liquid chromatography (RP-HPLC) and diode array detector (DAD). One of these compounds,10-methoxy-6-methylergoline-8beta-methanol-5-bromo-3-pyridinecarboxylate (1-DN) is the metabolite as well as the impurity whereas, the other 10-methoxy-1,6-dimethylergoline-8beta-methanol-5-chloro-3-pyridinecarboxylate (5-CN) is only an impurity. The chromatographic column was Phenomenex, Luna, 5 microm, C18 (2), 250 mm x 4.6 mm. Mobile phase was 0.1 M ammonium acetate (NH4Ac) solution containing 4 mM 1-octanesulfonicacid sodium salt (OSASS) and 6 mM tetrabutylammonium hydrogen sulphate (TBAHS) (pH: 5.9)/acetonitrile (ACN) (62:38) for 1-DN and (64:36) for 5-CN. Flow rate was 1.0 mL min-1. The diode array detector was operated at 285 nm, band width: 4 nm. Linearity was obtained in the concentration range of 0.032 x 10-5 to 3.828 x 10-5 M, y = 116.88x + 0.2773 (r2 = 0.99989); the limit of detection (LOD) and limit of quantification (LOQ) were determined as 0.012 x 10-5 and 0.041 x 10-5 M for 1-DN, respectively. Linearity was obtained in the concentration range of 0.034 x 10-5 to 4.092 x 10-5 M, y = 104.24x + 0.7486 (r2 = 0.99996); (LOD) and (LOQ) were determined as 0.014 x 10-5 and 0.046 x 10-5 M for 5-CN, respectively. The recovery was 100.65% for 1-DN and 100.32% for 5-CN. The amount of 1-DN in 30 mg NIC was found as 209.65 microg (0.70%) and the amount of 5-CN in 30 mg NIC was found as 27.62 microg (0.09%).     

Use of ceric ammonium sulphate and two dyes, methyl orange and indigo carmine, in the determination of lansoprazole in pharmaceuticals

Two spectrophotometric methods are proposed for the assay of lansoprazole (LPZ) in bulk drug and in dosage forms using ceric ammonium sulphate (CAS) and two dyes, methyl orange and indigo carmine, as reagents. The methods involve addition of a known excess of CAS to LPZ in acid medium, followed by determination of residual CAS by reacting with a fixed amount of either methyl orange, measuring the absorbance at 520 nm (method A), or indigo carmine, measuring the absorbance at 610 nm (method B). In both methods, the amount of CAS reacted corresponds to the amount of LPZ and the measured absorbance was found to increase linearly with the concentration of LPZ, which is corroborated by the correlation coefficients of 0.9979 and 0.9954 for methods A and B, respectively. The systems obey Beer's law for 0.5-7.0 microg mL(-1) and 0.25-3.0 microg mL(-1) for methods A and B, respectively. The apparent molar absorptivities were calculated to be 3.0 x 10(4) and 4.4 x 10(4) L mol(-1) cm(-1) for methods A and B, respectively. The limits of detection (LOD) and quantification (LOQ) were calculated to be 0.08 and 0.25 microg mL(-1) for method A, and 0.09 and 0.27 microg mLs(-1) for method B, respectively. The intra-day and inter-day precision and accuracy of the methods were evaluated according to the current ICH guidelines. Both methods were of comparable accuracy (er < or = 2 %). Also, both methods are equally precise as shown by the relative standard deviation values < 1.5%. No interference was observed from common pharmaceutical adjuvants. The accuracy of the methods was further ascertained by performing recovery studies using the standard addition method. The methods were successfully applied to the assay of LPZ in capsule preparations and the results were statistically compared with those of the literature UV-spectrophotometric method by applying Student's t-test and F-test.     

Quantitative determination of anti-tumor agent bis(4-fluorobenzyl)trisulfide, fluorapacin and its pharmaceutical preparation by high-performance liquid chromatography

A simple isocratic and stability-indicating HPLC method was developed and validated for the quantitative determination of anti-tumor agent fluorapacin and its pharmaceutical preparation. A Spherisorb ODS II C(18) (250 mm x 4.6 mm, 5 microm) column was eluted with a mobile phase consisting of acetonitrile/water (85:15, v/v). The analyses were performed at 40+/-1 degrees C with a flow rate of 1.0 mL/min and UV detection at 218 nm. The calibration curve was linear over a concentration range of 160-240 microg/mL with the correlation coefficient of 0.9997. The LOD and LOQ were determined to be 1.4 and 7.0 ng/mL, respectively. Average recoveries were 98.27% and 100.40% for fluorapacin API and its drug product with corresponding relative standard deviations (R.S.D.) of 0.41% and 0.30%, respectively. Good repeatability (precision and intermediate precision), accuracy and tolerability were obtained with R.S.D. of <1.0%. This specific and reliable method has been successfully applied for quality control of fluorapacin API and drug product.     

HPLC法同时测定复方利血平氨苯蝶啶片中3种成分含量

目的:建立HPLC法测定复方利血平氨苯蝶啶片中氢氯噻嗪、硫酸双肼屈嗪和氨苯蝶啶的含量.方法:色谱柱:Waters Symmetry C18( 150 mm ×4.6 mm,5μm),流动相:0.1％庚烷磺酸钠溶液(含0.05％磷酸二氢铵,磷酸调pH为3.0)-乙睛(80∶ 20),检测波长:310 nm.结果:氢氯噻嗪、硫酸双肼屈嗪和氨苯蝶啶均在0.06 ～0.18 mg·ml-1浓度范围内线性关系良好(r =0.999 9);平均回收率为100.2％,RSD=0.22％.结论:该方法具有操作简便、结果准确的优点,可用于测定复方利血平片的含量测定.     

Use of Folin-Ciocalteu phenol reagent and 3-methyl-2-benzothiazolinone hydrazine hydrochloride in the determination of oxcarbazepine in pharmaceuticals

Two spectrophotometric methods are proposed for the assay of oxcarbazepine (OXC) in bulk and dosage forms using Folin-Ciocalteu phenol reagent (FCP) and 3-methyl-2-benzothiazolinone hydrazine hydrochloride (MBTH) as reagents. The first method involves addition of FCP reagent to OXC in alkaline medium followed by measurement of absorbance at 760 nm (method A), and the other involves addition of a fixed volume of MBTH after treatment of OXC with ferric chloride and measurement of absorbance at 456 nm (method B). In both methods, the amount of chromogen formed corresponds to the amount of OXC and the measured absorbance was found to increase linearly with the concentration of OXC, which is corroborated by the correlation coefficients of 0.9985 and 0.9984 for method A and B, respectively. The systems obey Beer's law for 5-30 microg mL(-1) and 10-50 microg mL(-1) for methods A and B, respectively. The apparent molar absorptivity was calculated to be 8.06 x 10(3) L mol(-1) cm(-1) and 3.126 x 10(3) L mol(-1) cm(-1) for methods A and B, respectively. The limits of detection (LOD) and limit of quantification (LOQ) were calculated to be 1.6 and 5 microg mL(-1) for method A and 3 and 10 microg mL(-1) for method B. The inter-day and intra-day imprecision of the methods were found to be in the range of 1.1-1.7 and 0.9-1.1% for method A, and 1.1-1.9 and 0.6-0.9% for method B. The accuracy ranged between 98.9-99.7% and 99.3-100.1 for method A and B, respectively. No interference was observed from common pharmaceutical excipients. The methods were successfully applied to the assay of OXC in tablet preparations.     

HPLC法同时测定青天葵药材中7种黄酮的含量

建立同时测定青天葵中鼠李秦素(1)、鼠李柠檬素(2)、鼠李素(3)、鼠李秦素-3-O-β-D-葡萄糖苷(4)、鼠李秦素-3-O-β-D-木糖-(1→4)-β-D-葡萄糖苷(5)、鼠李秦素-3-O-β-D-葡萄糖-(1→4)-β-D-葡萄糖苷(6)和鼠李柠檬素-3-O-β-D-葡萄糖-(1→4)-β-D-葡萄糖苷(7)含量的高效液相色谱法。采用Kromasil C18色谱柱(250 mm×4.6mm,5μm),以0.4%磷酸乙腈为流动相进行梯度洗脱,流速为1.0 mL.min 1,检测波长为256 nm,柱温为40℃。7种黄酮类化合物(1～7)的线性范围分别为0.55～70.00μg.mL 1(r=0.999 7)、0.86～110.00μg.mL 1(r=0.999 7)、0.39～50.00μg.mL 1(r=0.999 7)、0.55～70.00μg.mL 1(r=0.999 5)、1.33～170.00μg.mL 1(r=0.999 8)、1.33～170.00μg.mL 1(r=0.999 8)、0.16～20.00μg.mL 1(r=0.999 5),平均回收率在97.19%～99.45%之间,RS...     

Comparison of UV spectrophotometric method and high performance liquid chromatography for the analysis of flunarizine and its application for the dissolution test

This study aimed to develop a simple UV spectrophotometric method for the analysis and the dissolution test of flunarizine in capsules. The UV absorbance was both measured directly and by the first derivative measurements at 254 and 268 nm, respectively. The developed methods were validated for their linearity, accuracy, precision, limit of detection (LOD) and limit of quantitation (LOQ) in comparison with the reported HPLC method. The UV spectrophotometric method illustrated excellent linearity (r2 > 0.9999) in the concentration range of 6-24 microg/mL. Precision (%R.S.D. < 1.50) and recoveries were good (%R > 99.62). The LOD of direct UV and first derivative measurements were 0.09 and 0.84 microg/mL, respectively, and the LOQ were 0.26 and 2.55 microg/mL, respectively. Results from the assay of flunarizine in capsules by the UV spectrophotometric methods, both direct and first derivative measurements were not significantly different from those of the HPLC method (P > 0.05). Additionally, the method was successfully used for the dissolution test of flunarizine capsule and was found to be reliable, simple, fast, and inexpensive.     

Chromatographic determination of polymerized impurities in meropenem

A method for the separation of polymerized impurities in meropenem has been developed by gel filtration chromatography (GFC). The chromatographic conditions included a Superdex peptide HR 10/30 column and the mobile phase consisting of 0.01 mol L(-1) sodium phosphate buffer (pH 7.0), at a flow rate of 0.8 mL min(-1). The wavelength of UV detector was set at 220 nm. The linear range was 0.08-10 microg (r=0.9998); relative standard deviation (R.S.D.)=0.5-1.5%; the LOD and LOQ were 2.4 and 12.4 ng, respectively. Specificity of the GFC was checked by switching the effluent of each peak on the GFC to a C18 trap column and analyzing the effluent by LC-MS. The result shows that for the determination of polymers, the gel filtration chromatography is a rather simple separation mode as compared to RPLC.     

大鼠一次性灌服酸枣仁汤剂提取物后芒果苷的药代动力学研究

沈阳药科大学 药学院, 辽宁 沈阳 110016     

Simultaneous determination of benazepril hydrochloride and hydrochlorothiazide by micro-bore liquid chromatography

A micro-bore liquid chromatographic method was developed for the simultaneous determination of benazepril hydrochloride and hydrochlorothiazide in pharmaceutical dosage forms. The use of a BDS C-18 micro-bore analytical column, results in substantial reduction in solvent consumption and increased sensitivity. The mobile phase consisted of a mixture of 0.025 M sodium dihydrogen phosphate (pH 4.8) and acetonitrile (55:45, v/v), pumped at a flow rate of 0.40 ml min(-1). Detection was set at 250 nm using an ultraviolet detector. Calibration graphs are linear (r better than 0.9991, n = 5), in concentration range 5.0-20.0 microg ml(-1) for benazepril hydrochloride and 6.2-25.0 microg ml(-1) for hydrochlorothiazide. The intra- and interday R.S.D. values were <1.25% (n = 5), while the relative percentage error (Er) was <0.9% (n = 5). The detection limits attained according to IUPAC definition were 0.88 and 0.58 microg ml(-1) for benazepril hydrochloride and hydrochlorothiazide, respectively. The method was applied in the quality control of commercial tablets and content uniformity test and proved to be suitable for rapid and reliable quality control.     

Sensitive fluorimetric method based on sequential injection analysis technique used for dissolution studies and quality control of prazosin hydrochloride in tablets

This report introduces a fully automated flow system for drug-dissolution studies based on the coupling of the sequential injection analysis (SIA) technique with a conventional dissolution apparatus. The methodology described was used for monitoring of dissolution profiles of prazosin hydrochloride (PRH) in pharmaceutical formulation. The very sensitive fluorimetric detection of PRH was performed at lambda(ex)=244 nm (lambda(em)>or=389 nm). Under the optimal conditions, the calibration curve was linear over the range 0.02-2.43 mg x l(-1) of PRH with R.S.D. 1.89, 1.23, and 1.80% (n=10) when determining 0.02, 1.22, and 2.43 mg x l(-1) of PRH in standard solutions, respectively. Equation of the calibration curve was calculated giving the following values: F=4.108 c-3.9 (n=6), r=0.9996. Detection limit was calculated 0.007 mg x l(-1) of PRH. The dissolution test of Deprazolin tablets was programmed for 60 min, with a continuous sampling rate of 70 h(-1) under conditions required by USP 26. Results obtained by SIA technique compared well with HPLC standard method.     

The determination of levofloxacin by flow injection analysis using UV detection,potentiometry, and conductometry in pharmaceutical preparations

A flow injection analysis (FIA) using UV detection, potentiometry and conductometry for levofloxacin (LVF) are described in this study. The best solvent system was found to consist of 0.2 M acetate buffer at pH 3 having 10% MeOH. A flow rate of 1 ml min(-1) was pumped and active material was detected at 288 nm. The detection limit (LOD) and limit of quantification (LOQ) for FIA were calculated to be 3 x 10(-7) M (S/N = 3) and 1 x 10(-7) M (S/N = 10), respectively. In the analysis of tablets, the RSD values were found to be 0.83, 0.98 and 0.99 for FIA, potentiometric and conductometric methods, respectively.     

HPLC method for the determination of ecdysterone in extractive solution from Pfaffia glomerata

A RP-LC method was developed and validated to quantify ecdysterone in extractive solution from subterraneous parts of Pfaffia glomerata. The analysis was performed using a RP-18 column with acetonitrile:water isocratic elution and the detection was carried out by UV at 242 nm. The standard curve for ecdysterone was linear over the range of 5.2-41.6 microg/ml (R2=0.9995). The extractive solution showed linear response in the range of 25.05-175.35 microg/ml (R2=0.9977). This method showed excellent repeatability (relative standard deviation, R.S.D.<2.0%), intermediary precision (R.S.D.=2.13%) and accuracy (101.04; R.S.D.=1.51%). The limit of detection (LOD) was 0.036 microg/ml and the limit of quantification (LOQ) was 0.110 microg/ml, demonstrating the sensitivity of the method. This assay can be readily utilized as quality controlled method for P. glomerata preparations.     

LC method for the analysis of cefetamet pivoxil hydrochloride in drug substance and powder for oral suspension

A high-performance liquid chromatography isocratic procedure was developed for the assay of cefetamet pivoxil hydrochloride in drug substance and powder for oral suspension. The method validation yielded good results and included the range, linearity, precision intra- inter-day, accuracy, specificity, LOD and LOQ values. The chromatographic system consisted of a C(18) absorbosphere column (150 x 4.6 mm i.d., 5 microm particle size), a mobile phase composed of water-acetonitrile-methanol-phosphate buffer, pH 3.5 (50:35:10:5, v/v), flow rate of 1.5 ml min(-1) and UV detection at 254 nm. The relative standard deviation varied between 0.03 and 1.76%, and accuracy of 100.09% was found. Calibration curve was linear from 30.0-80.0 microg ml(-1); its correlation coefficient was 0.99989.     

Chiral separation of ketoprofen on an achiral C8 column by HPLC using norvancomycin as chiral mobile phase additives

A high-performance liquid chromatographic method for chiral separation of ketoprofen racemate was developed. (R)- and (S)-ketoprofen enantiomers were separated on a Hypersil BDS C8 column (150 mm x 4.6 mm i.d., 5 microm) at 25 degrees C, using acetonitrile-triethylamine acetate (TEAA) buffer (pH 5.2, 20 mM) (35:65, v/v) containing 2.0 mM norvancomycin as the mobile phase. Effects of norvancomycin concentration, content of acetonitrile and TEAA buffer pH on the enantioseparation were investigated. The method was validated for linearity, repeatability, limits of detection (LOD) and limits of quantification (LOQ). Calibration curves (r2 = 0.999) were constructed in the range of 2.01-200.8 microg ml(-1) for (S)-ketoprofen and 2.04-152.4 microg ml(-1) for (R)-ketoprofen, respectively. Repeatability (n = 5) showed less than 2% relative standard deviation (R.S.D.). LOD and LOQ for the two enantiomers were found to be 0.20 and 0.78 ng for (S)-ketoprofen, 0.20 and 0.86 ng for (R)-ketoprofen, respectively. Norvancomycin and vancomycin as chiral mobile phase additives (CMPAs) in the chiral separation showed similar abilities of enantioseparation. However, to obtain the optimum enantioseparation, a lower concentration of norvancomycin than that of vancomycin is required.     

An enantiospecific HPLC method for the determination of (S)-enantiomer impurities in (R)-tolterodine tartarate

A high-performance liquid chromatographic method was developed for the separation of the enantiomers of tolterodine tartarate. The proposed method was applied to the determination of (S)-isomer in (R)-tolterodine tartarate, and satisfactory results were obtained. The enantiomers of tolterodine tartarate were separated on a Chiralpak AD-H (250 mm x 4.6 mm) column containing amylose tris-(3,5-dimethylphenyl-carbamate) at room temperature. The mobile phase consisted of n-hexane and isopropyl alcohol in the ratio of 85:15 (v/v) with 0.075% triethylamine (TEA) and 0.05% trifluoroacetic acid (TFA) as the additive. The flow rate was kept at 0.5 ml/min, and UV detection wavelength was set at 283 nm. The calibration curves of (S)-enantiomer in the concentration range from 0.05 microg/ml to 1 microg/ml range were linear. The relative standard deviations of within-day and between-day were less than 2% (n = 3). The limit of detection (LOD) was 0.75 ng (S/N = 3) and the limit of quantification (LOQ) was 0.05 microg/ml (RSD < 4.1%, n = 3). The determination recoveries of the (S)-enantiomer were in the range of 98.2-104.8%. The results demonstrated that the developed HPLC method was a reliable, simple technique and was applicable to the purity determination of (R)- tolterodine tartarate.