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1.
A high performance thin layer chromatographic method was developed and validated for the quantification of fluoxetine in human serum. Fluoxetine was extracted by liquid–liquid extraction method with diethyl ether as extraction solvent. Imipramine was used as internal standard. The chromatographic separation was achieved on precoated silica gel F 254 high performance thin layer chromatographic plates using a mixture of toluene/acetic acid glacial (4:5 v/v) as mobile phase. 4‐Dimethylamino‐azobenzene‐4‐sulphonyl chloride was used as derivatization reagent. Densitometric detection was done at 272 nm. The method was linear between 12.5 and 87.5 ng/spot, corresponding to 0.05 and 0.35 ng/μL of fluoxetine in human serum after extraction process and applying 25 μL to the chromatographic plates. The method correlation coefficient was 0.999. The intra‐assay and inter‐assay precisions, expressed as the RSD, were in the range of 0.70–2.01% (n=3) and 0.81–3.90% (n=9), respectively. The LOD was 0.23 ng, and the LOQ was 0.70 ng. The method proved be accurate, with a recovery between 94.75 and 98.95%, with a RSD not higher than 3.61% and was selective for the active principle tested. This method was successfully applied to quantify fluoxetine in patient serum samples. In conclusion, the method is useful for quantitative determination of fluoxetine in human serum.  相似文献   

2.
《Analytical letters》2012,45(7):1381-1391
Abstract

A rapid, sensitive, and specific liquid chromatography‐electrospray ionization mass spectrometric (LC‐ESI‐MS) method has been developed for quantification of gliclazide in human plasma. The analyte and tolbutamide (internal standard, I.S.) were extracted from plasma samples with n‐hexane–dichloromethane (1:1, v/v) and analyzed on a C18 column. The chromatographic separation was achieved within 4.0 min by using methanol–0.5% formic acid (80:20, v/v) as mobile phase and the flow rate was 1.0 mL/min. Ion signals m/z 324.0 and 271.0 for gliclazide and internal standard were measured in the positive mode, respectively. The method was linear within the range of 2.5–2000 ng/mL. The lower limit of quantification (LLOQ) was 2.5 ng/mL. The intra‐ and inter‐day precisions were lower than 2.8% in terms of relative standard deviation (RSD). The inter‐day relative error (RE) as determined from quality control samples (QCs) ranged from ?1.93% to 1.85%. This validated method was successfully applied for the evaluation of pharmacokinetic profiles of gliclazide modified‐release tablets in 20 healthy volunteers.  相似文献   

3.
A simple, precise, and rapid high‐performance thin‐layer chromatographic (HPTLC) method for the simultaneous quantification of pharmacologically important naphthoquinone shikonin ( 1 ) together with its derivatives acetylshikonin ( 2 ), and β‐acetoxyisovalerylshikonin ( 3 ) in four species of genus Arnebia (A. euchroma, A. guttata, A. benthamii, and A. hispidissima) from the Indian subcontinent has been developed. In addition, the effect of solvents with varying polarity (hexane, chloroform, ethyl acetate, and methanol) for the extraction of these compounds was studied. HPTLC was performed on precoated RP‐18 F254S TLC plates. For achieving good separation, mobile phase consisting of ACN/methanol/5% formic acid in water (40:02:08 v/v/v) was used. The densitometric determination of shikonin derivatives was carried out at 520 nm in reflection/absorption mode. The method was validated in terms of linearity, accuracy, precision, robustness, and specificity. The calibration curves were linear in the range of 100–600 ng for shikonin and acetylshikonin, and 100–1800 ng for β‐acetoxyisovalerylshikonin. Lower LOD obtained for compounds 1 – 3 were 18, 15, and 12 ng, respectively, while the LOQ obtained were 60, 45, and 40 ng, respectively.  相似文献   

4.
A high performance liquid chromatographic method was developed and validated for the quantitative determination of carbamazepine in intravenous nanoemulsions. The method validation yielded good results with respect to linearity, specificity, precision and accuracy. The method was carried out on a RP-18 column with a mobile phase composed of methanol–water (70:30 v/v) subjected to a gradient of acetonitrile after drug elution, and detection at 286 nm. The linearity in the range of 10.0–50.0 μg mL−1 presented a determination coefficient (r 2) of 0.9996, calculated by least-squares regression; the RSD values for intra-day and inter-day precision for % recovered were <0.44 and <1.21%, respectively; and the recovery of carbamazepine from the sample matrix ranged from 94.3 to 104.9%.  相似文献   

5.
An instrumental planar chromatographic (HPTLC) method for quantitative analysis of clozapine in human serum was developed and validated. Clozapine was extracted with n-hexane-isoamyl alcohol (75:25 v/v). The chromatographic separation was achieved on precoated silica gel F 254 HPTLC plates using a mixture of chloroform and methanol (9:1 v/v) as mobile phase. Quantitative analyses were carried out by densitometry at a wavelength of 290 nm. The method was linear between 10 and 100 ng/spot, corresponding to 0.10 and 1.00 ng/microL of clozapine in human serum after extraction process and applying 10 microL to the chromatographic plates. The method correlation coefficient was 0.999. The intra-assay variation was between 2.10 and 3.33% (n = 5) and the interassay was between 2.67 and 4.44% (n = 9). The detection limit was 0.03 ng/microL, and the quantification limit was 0.05 ng/microL. The method proved to be accurate, with a recovery between 97.00 and 99.00%, with an RSD not higher than 7.22%, and was selective for the active principle tested. This method was successfully applied to quantify clozapine in patient serum samples. In conclusion, the method is useful for the quantitative determination of clozapine in serum.  相似文献   

6.
Artemisinin (AN) and artemisinic acid (AA), valuable phyto‐pharmaceutical molecules, are well known anti‐malarials, but their activities against diseases like cancer, schistosomiasis, HIV, hepatitis‐B and leishmaniasis are also being reported. For the simultaneous estimation of AN and AA in the callus and leaf extracts of A. annua L. plants, we embarked upon a simple, rapid, selective, reliable and fairly economical high performance thin layer chromatography (HPTLC) method. Experimental conditions such as band size, chamber saturation time, migration of solvent front and slit width were critically studied and the optimum conditions were selected. The separations were achieved using toluene–ethyl acetate, 9:1 (v/v) as mobile phase on pre‐coated silica gel plates, G 60F254. Good resolution was achieved with Rf values of 0.35 ± 0.02 and 0.26 ± 0.02 at 536 nm for AN and 626 nm for AA, respectively, in absorption–reflectance mode. The method displayed a linear relationship with r2 value 0.992 and 0.994 for AN and AA, respectively, in the concentration range of 300–1500 ng for AN and 200–1000 ng for AA. The method was validated for specificity by obtaining in‐situ UV overlay spectra and sensitivity by estimating limit of detection (30 ng for AN and 15 ng for AA) and limit of quantitation (80 ng for AN and 45 ng for AA) values. The accuracy was checked by the recovery studies conducted at three different levels with the known concentrations and the average percentage recovery was 101.99% for AN and 103.84% for AA. The precision was analyzed by interday and intraday precision and was 1.09 and 1.00% RSD for AN and 1.22 and 6.05% RSD for AA. The analysis of statistical data substantiates that this HPTLC method can be used for the simultaneous estimation of AN and AA in biological samples. Copyright © 2015 John Wiley & Sons, Ltd.  相似文献   

7.
A simple and specific bioanalytical method based on reversed‐phase high‐performance liquid chromatography (RP‐HPLC) coupled with ultraviolet detection was developed and validated for the determination of a novel valproic acid arylamide, N‐(2‐hydroxyphenyl)‐2‐propylpentanamide (HO‐AAVPA) in rat hepatic microsomes (a subcellular fraction containing phase I enzymes, especially cytochrome P450). The chromatographic separation was achieved using a reversed‐phase Zorbax SB‐C18 column and a mobile phase of acetic acid in water (0.2% v/v) and acetonitrile (40:60 v/v) with a flow rate of 0.5 mL/min. The calibration curve was linear over the range of 882–7060 ng/mL (r2 = 0.9987), and the lower limit of quantification and the lower limit of determination were found to be 882 and 127.99 ng/mL, respectively. The method was validated with excellent sensitivity, and intra‐day accuracy and precision varied from 93.79 to 93.12%, and from 2.12 to 4.36%, respectively. The inter‐day accuracy and precision ranged from 93.29 to 97.30% and from 0.68 to 3.60%, respectively. The recovery of HO‐AAVPA was measured between 91.36 and 97.98%. The assay was successfully applied to the analysis of kinetic metabolism and pharmacokinetic parameters in vitro by a substrate depletion approach. Copyright © 2014 John Wiley & Sons, Ltd.  相似文献   

8.
Two simple, selective, precise and highly sensitive high‐performance thin‐layer chromatography (HPTLC) methods have been developed and validated for analysis of five angiotensin II receptor antagonists, namely losartan, irbesartan valsartan, candesartan and olmesartan, which are widely used in clinical practice. HPTLC of the drugs was performed on pre‐coated silica gel HPTLC plates 60 F254 by development using a mobile phase composed of chloroform–acetone–glacial acetic acid (7.8:1.5:0.7m v/v/v), which was suitable for all of the studied drugs. The first method depended on utilizing reflectance/fluorescence mode for detection while the second method depended on using 2,3,5,6‐tetrachloro‐1,4‐benzoquinone as spraying reagent for the first time to form orange spots scanned at 460 nm. A good linear relationship was obtained over the concentration ranges of 1.2–60 and 360–3000 ng/band while detection and quantification limits were in the ranges of 0.07–0.43, 45.2–140.49 and 0.21–1.29, 137.05–425.74 ng/band for reflectance/fluorescence and reflectance/absorbance methods respectively. The developed methods were applied successfully for their determination in tablets and spiked human plasma for reflectance/fluorescence method with good accuracy and precision, and so can be applied in the pharmacokinetic and bioavailability studies.  相似文献   

9.
A densitometric high performance thin-layer chromatographic (HPTLC) method was developed and validated for quantitative analysis of L-DOPA in tablets. Chromatographic separation was achieved on precoated silica gel F 254 HPTLC plates using a mixture of acetone-chloroform-n-butanol-acetic acid glacial-water (60:40:40:40:35 v/v/v/v/v) as mobile phase. Quantitative analysis was carried out at a wavelength of 497 nm. The method was linear between 100 and 500 ng/microL, with a correlation coefficient of 0.999. The intra-assay variation was between 0.26 and 0.65% and the interassay was between 0.52 and 2.04%. The detection limit was 1.12 ng/microL, and the quantification limit was 3.29 ng/microL. The accuracy ranged from 100.40 to 101.09%, with a CV not higher than 1.40%. The method was successfully applied to quantify L-DOPA in real pharmaceutical samples, including the comparison with HPLC measurements. The method was fast, specific, with a good precision, and accurate for the quantitative determination of L-DOPA in tablets.  相似文献   

10.
A fast, sensitive and specific method is presented for the quantification of RSD921 in human plasma by liquid chromatography coupled with tandem mass spectrometry using tri-deuterated RSD921 (3d-RSD921) as an internal standard. A single-step liquid/liquid extraction was performed with diethyl ether/hexane (80 : 20, v/v) using 0.5 ml of plasma. The plasma calibration curves were linear from 0.1 to 20 ng ml(-1) (r > 0.999). Between-run precision, based on the percent relative deviation for replicate (n = 40) quality controls, was < or =7.27% (0.5 ng ml(-1)), < or =7.39% (5.0 ng ml(-1)), and < or =5.06% (20.0 ng ml(-1)). Between-run accuracies, based on the relative error, were +/-2.59%, +/-1.23% and +/-1.64% respectively. The method was developed to evaluate the pharmacokinetic profile after 15 min of intravenous stepwise-ascending infusion dose of RSD921 in 18 healthy volunteers. A dissociation study of protonated RSD921 and 3d-RSD921 by collision-induced dissociation using in-source fragmentation and tandem mass spectrometry is also presented.  相似文献   

11.
We describe a simple, rapid and sensitive high‐performance liquid chromatography–electrospray ionization tandem mass spectrometric method that was developed for the simultaneous determination of carebastine and pseudoephedrine in human plasma using cisapride as an internal standard. Acquisition was performed in multiple‐reaction monitoring mode by monitoring the transitions: m/z 500.43 > 167.09 for carebastine and m/z 166.04 > 147.88 for pseudoephedrine. The devised method involves a simple single‐step liquid–liquid extraction with ethyl acetate. Chromatographic separation was performed on a C18 reversed‐phase chromatographic column at 0.2 mL/min by isocratic elution with 10 mM ammonium formate buffer–acetonitrile (30:70, v/v; adjusted to pH 3.3 with formic acid). The devised method was validated over 0.5–100 ng/mL of carebastine and 5–1000 ng/mL of pseudoephedrine with acceptable accuracy and precision, and was successfully applied to a bioequivalence study involving a single oral dose (10 mg of ebastine plus 120 mg of pseudoephedrine complex) to healthy Korean volunteers. Copyright © 2010 John Wiley & Sons, Ltd.  相似文献   

12.
A new sample treatment for liquid chromatographic analysis of ochratoxin A (OTA) in beer is proposed. Degassed beer is mixed with lead hydroxyacetate, which precipitates some bulk components but does not remove OTA. The precipitate is separated and the acidified liquid is extracted with chloroform. The solvent is evaporated and the residue is dissolved in mobile phase (acetonitrile-water, 40:60, v/v; acidified at pH 3.0 with phosphoric acid) and separated by liquid chromatography using fluorescence detection. The limit of detection was 0.005 ng/ml. The average recovery rate and the average RSD of recovery in the spiking level range 0.01-0.5 ng/ml were 95.5% and about 5%, respectively. The method is cheaper that other alternative ones using immunoaffinity columns or other solid-phase extraction cleanup:The separation was optimised with regard to composition and flow of the mobile phase and no interference from the matrix was found. The method was applied to 88 samples of beer (domestic and imported) marketed in Spain. OTA was detected in 82.9% of them. The range for positive samples was 0.007-0.204 ng of OTA/ml.  相似文献   

13.
A selective, precise, and accurate high-performance thin-layer chromatographic (HPTLC) method has been developed for the analysis of L-dopa in Mucuna pruriens seed extract and its formulations. The method involves densitometric evaluation of L-dopa after resolving it by HPTLC on silica gel plates with n-butanol-acetic acid-water (4.0+1.0+1.0, v/v) as the mobile phase. Densitometric analysis of L-dopa was carried out in the absorbance mode at 280 nm. The relationship between the concentration of L-dopa and corresponding peak areas was found to be linear in the range of 100 to 1200 ng/spot. The method was validated for precision (inter and intraday), repeatability, and accuracy. Mean recovery was 100.30%. The relative standard deviation (RSD) values of the precision were found to be in the range 0.64-1.52%. In conclusion, the proposed TLC method was found to be precise, specific and accurate and can be used for identification and quantitative determination of L-dopa in herbal extract and its formulations.  相似文献   

14.
A simple and rapid high‐performance thin‐layer chromatographic method was developed for the separation and determination of six flavonoids (rutin, luteolin‐7‐O‐β‐glucoside, chamaemeloside, apigenin‐7‐O‐β‐glucoside, luteolin, apigenin) and one coumarin, umbelliferone from chamomile plant samples and dietary supplements. The separation was achieved on amino silica stationary phase using dichloromethane/acetonitrile/ethyl formate/glacial acetic acid/formic acid (11:2.5:3:1.25:1.25 v/v/v/v/v) as the mobile phase. The quantitation of each compound was carried out using densitometric reflection/absorption mode at their respective absorbance maxima after postchromatographic derivatization using natural products reagent (1% w/v methanolic solution of diphenylboric acid‐β‐ethylamino ester). The method was validated for specificity, limits of detection and quantification, precision (intra‐ and interday) and accuracy. The limits of detection and quantification were found to be in the range from 6–18 and 16–55 ng/band for six flavonoids and one coumarin, respectively. The intra‐ and interday precision was found to be <5% RSD and recovery of all the compounds was >90%. The data acquired from high‐performance thin‐layer chromatography was processed by principal component analysis using XLSTAT statistical software. Application of principal component analysis and agglomerative hierarchial clustering was successfully able to differentiate two chamomiles (German and Roman) and Chrysanthemum.  相似文献   

15.
The current paper reports the development and validation of stability‐indicating HPLC and HPTLC methods for the separation and quantification of main impurity and degradation product of Carbimazole. The structures of the degradation products formed under stress degradation conditions, including hydrolytic and oxidative, photolytic and thermal conditions, were characterized and confirmed by MS and IR analyses. Based on the characterization data, the obtained degradation product from hydrolytic conditions was found to be methimazole—impurity A of Carbimazole as reported by the British Pharmacopeia and the European Pharmacopeia. A stability‐indicating HPLC method was carried out using a Zorbax Eclipse Plus CN column (150 × 4.6 mm i.d, 5 μm particle size) and a mobile phase composed of acetonitrile–0.05 m KH2PO4 (20: 80, v/v) in isocratic elution, at a flow rate of 1 mL/min. The method was proved to be sensitive for the determination down to 0.5% of Carbimazole impurity A. Additionally, a stability‐indicating chromatographic HPTLC method was achieved using cyclohexane–ethanol (9:1, v/v) as a developing system on HPTLC plates F254 with UV detection at 225 nm. The proposed HPLC and HPTLC methods were successfully applied to Carbimazole® tablets with mean percentage recoveries of 100.12 and 99.73%, respectively.  相似文献   

16.
Herein a quantitative method for the determination of seven penicillins in bovine plasma and veterinary drugs has been developed. Amoxicillin (AMO), ampicillin (AMP), penicillin G (PENG), penicillin V (PENV), oxacillin (OXA), cloxacillin (CLO) and dicloxacillin (DICLO) were separated on a Perfectsil ODS‐2 (250×4 mm, 5 μm) column, using gradient elution, with a mobile phase of 0.1% v/v TFA and ACN–methanol (90:10 v/v). PDA detection was used at 240 nm. Penicillins were isolated from bovine plasma by SPE on Lichrolut RP‐18 cartridges with mean recoveries from 85.7 to 113.5%. Colchicine (3 ng/μL) was used as an internal standard. The developed method was validated in terms of selectivity, linearity, accuracy, precision, stability and sensitivity. Repeatability (n = 5) and between‐day precision (n = 5) revealed RSD < 12%. The detection limits in the bovine plasma were estimated as 18 ng for AMO and AMP, 25 for PENG, PENV and OXA, 3 ng for CLO and 12 ng for DICLO. Spiked plasma samples were stable for 1 wk, except for AMP and CLO, which were stable for 3 wk and OXA for 4 wk. AMO, PENG and PENV were stable for two freeze–thaw cycles, OXA, CLO and DICLO for four, while AMP only for one.  相似文献   

17.
A rapid, sensitive and specific high‐performance thin‐layer chromatographic (HPTLC) method was developed and validated for determination of gliotoxin in Aspergillus infected immunocompromised patients with invasive aspergillosis (IA). Densitometric analysis of gliotoxin was carried out in the absorbance mode at 254 nm after single‐step extraction with chloroform. The method uses TLC aluminum plates pre‐coated with silica gel 60F‐254 as a stationary phase and toluene–isoamyl alcohol–methanol (10:0.5:0.5, v/v/v) as mobile phase, which gives compact spot of gliotoxin (Rf = 0.51). The calibration curve was linear (r2 ≥ 0.994) between peak area and concentration in the tested range of 100–1000 ng spot?1 with minimum detectable range 0.025 ng μ?1 of serum sample. The mean ± SD value of slope and intercept of the standard chromatogram of gliotoxin were found to be 523.2 ± 1.555635 and 915.8 ± 30.68843, respectively. The developed method is simple, rapid, precise and less costly than earlier diagnostic methods, and different serum samples can be run on a single TLC plate for comparative analysis. The proposed method can be used to analyze gliotoxin in patient serum for easy, rapid and cost‐effective diagnosis of IA. Copyright © 2009 John Wiley & Sons, Ltd.  相似文献   

18.
The present work describes novel methods using densitometry and indirect or off‐line high performance thin‐layer chromatography–mass spectrometry (HPTLC–MS) for the simultaneous detection and quantification of asenapine, propranolol and telmisartan and their phase II glucuronide metabolites. After chromatographic separation of the drugs and their metabolites the analytes were scraped, extracted in methanol and concentrated prior to mass spectrometric analysis. Different combinations of toluene and methanol–ethanol–n‐butanol–iso‐propanol were tested for analyte separation and the best results were obtained using toluene–methanol–ammonia (6.9:3.0:0.1, v/v/v) as the elution solvent. All of the drug–metabolite pairs were separated with a homologous retardation factor difference of ≥22. The conventional densitometric approach was also studied and the method performances were compared. Both of the approaches were validated following the International Conference on Harmonization guidelines, and applied to spiked human plasma samples. The major advantage of the TLC–MS approach is that it can provide much lower limits of detection (1.98–5.83 pg/band) and limit of quantitation (5.97–17.63 pg/band) with good precision (?3.0% coefficient of variation) compared with TLC–densitometry. The proposed indirect HPTLC–MS method is simple yet effective and has tremendous potential in the separation and quantitation of drugs and their metabolites from biological samples, especially for clinical studies.  相似文献   

19.
Abstract

A liquid chromatographic method is described for the determination of the new fluoroquinolone Ro 23–6240 and its N-demethyl and N-oxide metabolites in plasma and urine. The three substances were extracted from aqueous solution with dichloromethane/isopropanol containing sodium dodecyl sulphate. After evaporation and reconstitution, samples were analysed on a reversed-phase column using ion pair chromatography and fluorescence detection. The limit of quantification was 10–20 ng/ml (RSD 4%) using a 0.5 ml plasma sample, and the inter assay precision was 3–10% over the concentration range 50 ng/ml to 20 μg/ml. Recovery from plasma was 81% (RSD 10%) over the range 10 ng/ml to 5 μg/ml. The method has been applied successfully to the analysis of several thousand samples from human pharmacokinetic studies. Care has to be taken to avoid exposure of samples to direct sunlight, and the use of opaque vessels for sample storage and handling is recommended.  相似文献   

20.
A simple chiral analytical method was developed for the enantiomeric determination of cyflumetofen in cucumber, tomato, and apple by normal‐phase HPLC. The effects of mobile phase composition and column temperature on the enantioseparation were evaluated. Excellent separation was achieved at 25°C on a Chiralpak AD‐H column, with a mixture of n‐hexane and 2‐propanol (95:5, v/v) as mobile phase at a flow rate of 1.0 mL/min detecting at 234 nm. The resolution of cyflumetofen enantiomers was up to 5.5. The elution order of the enantiomers was determined by an online OR‐2090 detector, which was performed under the same chromatographic conditions. The first eluted enantiomer was (–)‐cyflumetofen and the second eluted one was (+)‐cyflumetofen. The method was validated for linearity, repeatability, accuracy, LOD, and LOQ. LOD ranged from 0.1 to 0.15 mg/kg, with the LOD varying from 0.33 to 0.5 mg/kg for each enantiomer, respectively. The average recoveries of the pesticide ranged from 71.4 to 102.0% at all fortification levels. The precision values associated with the analytical method, expressed as RSD values, were below 14.8% in all matrices. The method was then successfully applied to detect cyflumetofen enantiomers in real samples.  相似文献   

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