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Objective: The objective of this study was to develop and validate a novel ion-pair liquid chromatography method, in order to separate and assay of amlodipine/benazepril combination in capsules. This method was a fast, practical and additional choice in quality control laboratories. Methods: The chromatographic conditions comprised of a classical C18-type stationary phase (250 × 4.6 mm, 5μ), with a mobile phase consisting of: 45% of 10-3 M of cetrimide and 55% acetonitrile. The flow rate was 1 ml/min; the detection wavelength was at 242 nm, under ambient temperature. Results: The method was validated for linearity with correlation coefficients very close to one, the accuracy with mean recovery values between 95.0-105.0%, precision with relative standard deviations of the calculated concentrations less than 5.0% and specificity in the presence of degradation products and excipients. Conclusion: The results presented in this paper showed that the developed method was fast and applicable, for the separation and determination of amlodipine/benazepril combination in capsules.
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Objective: To develop a RP-ion pair HPLC with wavelength switching for the simultaneous determination of 4 components (marine, oxymarine, salicylic acid, benzoic acid) in Fufang Kusheng Shuiyangsuan powder.Methods: An Agilent ZORBAX SB-C18 column(150 mm× 4.6 mm, 5 μm) was used;the mobile phase was acetonitrile (A)-0.1% phosphoric acid solution (0.2 g sodium heptanesulfonate was added to 100 ml solution) at a flow rate of 1.0 ml·min-1;the detection wavelength was 220 nm in 0-12 min and 280 nm in 12-25 min;the column temperature was 30℃.Results: The linear range of marine, oxymarine, salicylic acid and benzoic acid was 0.006 030-0.120 6 μg (r=0.999 4), 0.016 56-0.331 2 μg (r=0.999 9), 0.717 1-14.34 μg (r=0.999 9) and 0.512 0-10.24 μg (r=0.999 9), respectively;the average recovery was 98.14%, 97.20%, 97.05% and 98.39% with the RSDs of 1.38%, 0.32%, 0.81% and 1.26%(n=6) , respectively.Conclusion: The method is simple and rapid, and can be applied in the simultaneous determination of 4 components in Fufang Kusheng Shuiyangsuan powder.
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Simple, selective and highly sensitive spectrophotometric methods are proposed for the rapid and accurate determination of anti-hypertensive drugs namely telmisartan (TEL), propranolol (PRO), bisoprolol (BIS) and carvedilol (CRV) in tablets and biological fluids using bromocressol green (BCG) and bromothymol blue (BTB). The developed methods involve formation of stable yellow colored dichloromethane extractable ion-pair complexes of the amino derivative of four antihypertensive drugs such as TEL, PRO, BIS and CRV with two sulphonphthalein acid dyes, namely; BCG and BTB in acidic buffer. The effect of optimum conditions via pH on the ion-pair formation, reagent concentration, time and temperature and solvent was studied. The composition of the ion-pairs was found 1: 1 by Job’s method. The established methods having high sensitivity and good selectivity could be applied to the determination of the studied drugs in pharmaceutical, urine and blood serum samples with satisfactory results. The results obtained are good agreement with experimental data. The reaction mechanism was also discussed.
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Simple, accurate, precise, and rapid extractive spectrophotometric method was developed for the determination of four antipsychotics drugs, namely sulpiride (SUP), olanzapine (OLP), clozapine (CLP) and aripiprazole (ARP) both in tablets and in biological fluids. The method was based on the formation of red colored ion-pair complex between the studied drugs and eriochrome black T (EBT) with absorption maxima at 514 nm. The stoichiometry of the complexes in either case was found to be 1: 1 and the conditional stability constant (Kf) of the complexes have been calculated. Reaction conditions were optimized to obtain the maximum color intensity. Beer’s law was obeyed in the concentration ranges of 4-30, 4-20, 2-18 and 4-26 μg/ml with SUP, OLP, CLP and ARP, respectively. Various analytical parameters have been evaluated and the results have been validated by satistical data. The proposed method was successfully applied to the analysis of commercial tablets containing the drugs and the results were in good agreement with those obtained with reported methods. The proposed method was further applied to the determination of the studied drugs in spiked human serum and urine. A proposal for the reaction pathway was postulated.
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A simple, accurate and highly sensitive spectrophotometric methods are proposed for the rapid and accurate determination of fluvoxamine maleate (FXA) using bromocressol green (BCG), methyl orange (MO) and bromothymol blue (BTB). The developed methods involve formation of stable yellow colored chloroform extractable ion-associate complexes of the amino derivative (basic nitrogen) of the FXA with three sulphonphthalein acid dyes, namely; BCG, MO and BTB, in potassium hydrogen phthalate buffer pH 3.3, 3.6 and 3.4 respectively. The ion-associates exhibit absorption maxima at 420, 420 and 410 nm for BCG, MO and BTB, respectively. FXA can be determined up to 2.0–16, 2.0–15 and 2.0–20 μgmL−1 for BCG, MO and BTB, respectively. The effect of optimum conditions via pH on the ion pair formation, reagent concentration, time and temperature, and solvent was studied. The composition of the ion pairs was found 1:1 by Job’s method. The low relative standard deviation values indicate good precision and high recovery values. These methods have been successfully applied for the assay of FXA in pure form and in pharmaceutical formulations and the results are in good agreement with those obtained by the official method.
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Objective To establish a HPLC method for the determination of Hyperoside in Bushen-Yijing wan. Methods The HPLC system consisted of the Agilent HC-C18(4.6 mm×250 mm, 5μm)column was adopted. The mobile phase was consisted of acetonitrile(10)-HK2PO4(90)(0.85 g/L sodium heptanesulfonate,H3PO4 adjusted pH to 4.8), the flow rate was 1.0 ml/min, the column temperature was 30 ℃, and the UV detector was set at 360 nm. Results The linear response range was 3.0~60.0μg/ml(r=0.999 9). The average recovery of hyperoside was 98.2%, and RSD1.53%. Conclusions The method is simple, rapid, accurate and repeatable. It can be applied in determination of Hyperoside in Bushen-Yijing wan.
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Simple, rapid and sensitive spectrophotometric procedure is suggested for the determination of ranitidine hydrochloride (RNH) drug in pure form and in pharmaceutical formulations. The method was based on the ionpair formations of RNH with different dyestuff reagents such as methyl orange (MO), bromocrysol purple (BCP), eriochrome cyanine R (ECR) and alizaraine red S (ARS). The obtained ion-pairs were measured spectrophotometrically at 408, 420, 330 and 326 nm by using BCP, MO, ECR and ARS reagents, respectively. Beer’s plots were linear in the concentration range of 5-200, 20-350, 10-150 and 10-180 μg mL−1 RNH, with correlation coefficients not less than 0.9991, 0.9996, 0.9993 and 0.999 using BCP, MO, ECR and ARS reagents, respectively. The Sandell sensitivity was found to be 0.813, 0.462, 0.541and 0.630 μg cm−2 for BCP, MO, ECR and ARS, respectively. Standard deviation (SD = 0.024-0.028, 0.018-0.023, 0.016-0.021 and 0.023–0.029) and relative standard deviation (RSD% = 0.123-0.943, 0.0102-0.82, 0.118-0.145 and 0.132-0.178%) (n = 4) values using BCP, MO, ECR and ARS reagents, respectively, were obtained. These results were also confirmed with percent recovery of 99.78–100.52%, 99.86-101.12%, 99.82–100.31% and 100.18-101.25 % for BCP, MO, ECR and ARS reagents, respectively. This method was successfully applied for determination of RNH in aciloc tablet. The calculated t- and F- values (95% confidence limit) indicate no significant differences between the proposed and official methods.
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An isocratic, selective, and accurate Ion pair - reversed phase liquid chromatographic method of analysis of Furosemide and Zonisamide both as a bulk drug and formulations was developed and validated. An ODS chromatographic column (250mmx4.6mm, 5μm) was used for the separation. The mobile phase consisted of a mixture of methanol and Tetrabutyl ammonium hydrogen sulphate (TBHS) 0.035 molar aqueous solution, pH adjusted to 6.0 using 1 N sodium hydroxide solution. The composition of TBHS with methanol used for Furosemide was 40:60(v/v) and that for Zonisamide was 50:50(v/v) delivered at a flow rate of 1.0ml/min and detection at wave length 240 nm. The developed method was validated in terms of selectivity, Linearity, limit of quantitation, precision, accuracy and solution stability. The proposed LC method achieved satisfactory resolution between Furosemide and 4-Chloro-5-sulphamoyl salicylic acid (CSSA), Zonisamide and CSSA an intermediate product possibly present in Furosemide and Zonisamide. The method can be employed as stability indicating method for both the drugs furosemide and zonisamide and its dosage form.
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Two simple, rapid, sensitive, precise and economic spectrophotometric methods have been developed for the estimation of Erlotinib in bulk and tablet formulation. During the course of study, it was observed that solution of the drug formed colored ion-pair complexes with Bromocresol Green (BCG) and Methyl Orange (MO) in phosphate buffer pH 2.5, and extracted in chloroform. This property of the drug was followed for the development of colorimetric methods for analysis of drug. The complex of etoricoxib with BCG and MO showed λ max at 418.5 nm and 424.4 nm respectively. The complex was stable up to 22 hrs and obeyed Beer's law over the concentration ranges of 10-1000 μg/ml. Correlation coefficient was found to be 0.9985. In addition we have determined the molar absorptivity, Sandell sensitivity and the optimum conditions for quantitative analysis of erlotinib. These methods were validated statistically. Recovery studies gave satisfactory results indicating that none of common additives and excipients interfere the assay method. The proposed methods are found to be simple, accurate and reproducible that was successfully applied for the analysis of tablet formulation.
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Objective: To establish a reversed-phase ion-pair chromatography (RP-IPC) method for determination of sinomenine content in Sinomenium acutum. Methods: The IPC method was established using a column of Eclipse Plus C18(4. 6 mmX250 mm, 5 μm) with a mobile phase of acetonitrile and buffer phosphate (0. 03 mol/L potassium dihydrogen phosphate and 0. 03 mol/L sodium heptane sulfonic acid, adjusted to pH 4. 0 by phosphoric acid) 18 : 82. The flow rate was 1. 0 ml/min; the detection wavelength was set at 262 nm, and the temperature was at 40°C. Results: The linearity was obtained over a concentration range of 0. 025-0. 5 mg/ml(r=1. 000). The low limit of detection was 2. 5 μg/ml. The intra-day precisions of low, medium, and high concentrations of sinomenine were 1.54%, 0. 70%, and 0. 45%, and the inter-day precisions were 1. 54%, 0. 70%, and 0. 45%, respectively. The average recovery was 99. 9%(RSD=2. 9%, n=6), which met the requirement for analysis. Conclusion: The present method is simple, fast, and accurate; it is suitable for determination of sinomenine content in Sinomenium acutum.
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A method was developed to elucidate the structures of sulfated oligosaccharides through establishment of an effective reversed phase ion pair ultra-performance liquid chromatography coupled with electrospray ionization time of flight mass spectrometry( RPIP-UPLC-ESI-TOF-MS). Heptylamine (20 mmol/L,pH4) has been selected as the ion-pairing agent.κ-Carrageenan oligosaccharides have been separated on BEH C_(18) column using MeOH/H_2O with 25% heptylammonium formate as eluent in linear gradient mode. Mass spectra were obtained by ESI-Q-TOF-MS in both positive and negative modes. κ-Carrageenan oligosaccharides were well separated up to pentatetrasaccharide,and ESIMS analysis for κ-carrageenan oligosaccharides up to hepta-cosasccharide. The results showed that all acid hydrolyzed κ-carrageenan oligosaccharides were odd sugars,which was further confirmed by polyacrylamide gel electrophoresis ( PAGE). The characteristic fragmentation pattern of ion-pair oligosaccharides in mass spectra can be applied for rapid structure identification.
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An analytical method of fluorescent dye-labeled oligonucleotides was established by ion pair reversed phase high performance liquid chromatography(IP-RP-HPLC) which was improved by optimizing the effects of triethylamine-acetic acid(TEAA)(0-0.15 mol/L), pH(4.5-7.0) and gradient. Comparing the retention of 5, 10 and 15-mer unlabeled oligonucleotides with that of 5'-carboxyfluorescein(5'FAM) labeled oligonucleotides, the mechanism of fluorescent dye-labeled oligonucleotides retention was studied. In addition, TaqMan~(TM) probes as wellas other common fluorescent dye-labeled oligonucleotides were concerned. The results showed that the best resolution of different length fluorescent dye-labeled oligonucleotides was observed under the condition of 0.01 mol/L TEAA and pH 7.0. The retention behavior of fluorescent dye-labeled oligonucleotides was different from that of unlabeled oligonucleotides significantly, and therefore they can be separated completely. The results indicated that the retention of unlabeled oligonucleotides enhanced with the increase of the length of molecule. In contrast, the retention of fluorescent dye-labeled oligonucleotides was reduced with the increase of the length of molecule. For the hydrophobicity of fluorescent dyes made a great impact on the retention, a longer retention time the labeled oligonucleotides would take while the hydrophobicity of fluorescent dyes was higher. However, the effect of the hydrophobicity was limited as the length was increased to a certain level.
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OBJECTIVE:To establish an ion-pair RP-HPLC method for determination of the concentration of netilmicin sulfate in human serum.METHODS:The determination was performed on Luna C18 column with column temperature set at 40℃;the mobile phase consisted of acetonitrile-water (15∶85,pH 2.0,containing 10 mmol?L-1 sodium heptanesulfonate) at a flow rate of 0.8 mL?min-1.The detection wavelength was set at 205 nm.RESULTS:The linear range of netilmicin was 2～20 mg?L-1(r=0.997 6) with a recovery rate of 93.66%～96.05%.The intra-day and inter-day RSD were both less than 10%,respectively.CONCLUSION:The method is sensitive and simple with accurate and reliable results,and it is able to satisfy the requirement for the determination of serum concentration of netilmicin.
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Objective To develop a method for the determination of berberine hydrochloride,palmatine hydrochloride and jatrorrhizine hydrochloride in Yiqi Zhixue Granules. Methods The samples were extracted by ultrasonication with methanol for 30 min,and an ion pair RP-HPLC method was applied to determine three kinds of alkaloids.The column of C18 with temperature at 30 ℃ was used to separate the target components,the mobile phase consisted of the mixed solution of 0.01 mol/L sodium-heptanesulfonate solution and equal volume of 0.01 mol/L potassium dihydrogen phosphate solution(pH being adjusted at 3.1 with phosphoric acid) combined with acetonitrile(70 ∶ 30),the flow rate was 1.0 mL/min,and the detection wavelength was at 345 nm. Results Three kinds of alkaloids were separated perfectly,the average recoveries(n=6)were 100.95 %(RSD=2.10 % ) for berberine chloride,102.14 %(RSD=2.29 % ) for palmatine chloride,and 100.71 %(RSD=2.65 % ) for jatrorrhizine chloride. Conclusion The developed method is demonstrated to be simple,specific and accurate,which can be used to determine the contents of berberine chloride,palmatine chloride and jatrorrhizine chloride in Yiqi Zhixue Granula,and to control the quality of Folium Mahoniae in Yiqi Zhixue Granules.
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AIM: A RP HPLC was developed to determine the contents of oxymatrine and matrine simultaneously in Compound Kushen Injection(Radix Sophorae Flavescentis, Rhizoma Heterosmilacis Japonicae). METHODS: The analysis was carried out on a column of Hypersil ODS 2(4.6mm?200mm,5?m) with a mobile phase of methanol acetonitrile water phosphate acid(10∶30∶65∶0.05,33mmol?L -1 ), flow rate being 1.0mL?min -1 , detection wavelength being at 210nm and the column temperature at room temperature. RESULTS: The linear range was in the range of 17.44～174.4?g?mL -1 ( r =0.9996, n =6) for oxymatrine and in the range of 1.96～ 39.20 ?g?mL -1 ( r=0.9997,n =6) for matrine, respectively. The average recoveries of oxymatrine and matrine were 100.4%( RSD =2.1%, n =9) and 99.7%( RSD =1.4%, n =9), respectively. CONCLUSION: The method is found to be simple and accurate for simultaneous analysis of oxymatrine and matrine in Compound Kushen Injection, and a reliable way to quality evaluation.
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A rapid and simple chromatographic procedure using HPLC-ECD is described for simultaneousdetermination of NE. E. DA. 5-HT with their precursor amino acid (Tyr. Trp) and their main metabolites (HVA. 5-HIAA). Using this assay,eight substrates are measured in cortex, diencephalon and brain stem of mice duing immunological response challenged by SRB6 3 days after challenged, the DA HVA in cortex, NE in diencephalon decrease obviously (p
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Objective:To determine berberine and palmatine in Phellodendron chinense Schneid.and its granules.Methods: The reversed-phase ion pair high performance liquid chromatography(RP-HPLC) was used and the validation of the method was tested.The chromatography condition was with Lichrospher C18 column(4.6 mm?250 mm,5 ?m), mobile phase was ACN∶25 mmol/L NaH 2PO 4∶25 mmol/L SDS(2∶1∶1),flow speed was 1.0 ml/min,detection wavelength was 345 nm,and temperature of column was 25℃,Phellodendron chinense Schneid.and its granules were extracted with methanol solution of hydrochloric acid.Results: The theoretical plate number of berberine and palmatine were 14 906 and 14 847,the resolution were 2.33 and 2.86,the tailing factor were 1.09 and 1.06,respectively; all the parameters were suitable for determination.The calibration curves were linear in the range of 40-500 ng,Y=698 278X-3 846,r=1.000(berberine) and 20-250 ng, Y= 536 632X- 7 738, r=0.999 9,r=0.999 4(palmatine).The intra-day and inter-day precision(RSD) at low,middle and high injection amount was all less than 2.5%(berberine) and 1.5%(palmatine).The stability(RSD) was 0.66%(berberine) and 0.70% (palmatine) in 48 h.The recurrence(RSD,n=5) was 0.11%(berberine) and 0.12%(palmatine).The limits of detection was 2.0 ng(berberine) and 1.0 ng(palmatine).The recoveries were 100.4% (RSD=0.12%,n=3) for berberine and 99.80%(RSD=0.22%,n=3) for palmatine.The contents of berberine and palmatine in 3 batch of Phellodendron chinense Schneid.and 5 batch of its granules were determined.Conclusion: Our method can be used for determination of berberine and palmatine in Phellodendron chinense Schneid.and its granules, which is simple and reliable.
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A high-performance liquid chromatographic procedure for the simultaneous determination of vitamin B1, B2 and niacin in fortified mixed protein was developed. The sample was heated on water bath by adding 0.1N HC1 at 100℃ for 30 min. Afterward, the vitamins in the filtrate of the sample were separated on a column of u-bondapak C18 with methanol/water containing 0.005M pic B7 ion pair reagents (40:60 V/V) as mobile phase, and then the eluate was measured at 254nm. The detection limits for vitamin B1, B2 and niacin were approximately 15 ng, 5ng and 6ng respectively. The average recoveries of vitamin B1, B2 and Niacin were 92.21,90.25 and 92.33% respectively. The coefficients of variation of these vitamins were all under 6%. The method is simple, rapid and accurate, so it could also be applied to the analysis of other samples such as fortified foods and multivitamin preprati-ons etc.