Simultaneous determination of binary mixtures of trimethoprim and sulfamethoxazole or sulphamethoxypyridazine by the bivariate calibration spectrophotometric method

The bivariate calibration algorithm was applied to the spectrophotometric simultaneous determination of trimethoprim (TMP), sulfamethoxazole (SMX) or sulphamethoxypyridazine (SMP) binary mixtures in pharmaceutical and veterinary products. The results obtained were compared with those from derivative spectrophotometry. The statistical evaluation of the method bias showed that the proposed procedure is comparable with commonly used first-derivative spectrophotometry. However, the advantage of bivariate calibration is its simplicity, due to the minimal spectra manipulation when compared with derivative techniques.     

Simultaneous determination of trimethoprim and sulphamethoxazole in veterinary formulations by chromatographic multivariate methods

A comparative chromatographic study was developed for the simultaneous quantitative resolution of trimethoprim (TMP) and sulphamethoxazole (SMX) in veterinary formulations. Multi-wavelength chromatograms were recorded by using diode array detector (DAD) system at the five-wavelength set consisting of 220, 230, 240, 250 and 260 nm. In the first step, five different calibration equations at the above wavelengths for each drug were obtained by using the relationship between concentration and peak area. These calibration graphs were used for the quantitative evaluation of TMP and SMX in samples. These single-wavelength applications were called traditional LC method. In the second step, principal component regression (PCR) and partial least-squares (PLS) calibrations were applied to the above mentioned multi-wavelength chromatograms. The amount of two investigated drugs in samples was determined by the constructed PCR and PLS calibrations. The experimental results obtained from each single-wavelength calibration graph were compared with those obtained by the chemometric approaches and chromatographic multivariate approaches give successful results more than traditional LC method.     

近红外光谱—偏最小二乘方法同时测定复方磺胺甲唑片的两种有效成分

目的建立复方磺胺甲口恶唑片的两种有效成分磺胺甲口恶唑(SMZ)和甲氧苄啶(TMP)的快速同时测定方法。方法基于近红外漫反射光谱技术,利用偏最小二乘方法建立该复方中SMZ和TMP的定量分析多元校正模型。结果对于所建立的SMZ与TMP模型,相关系数分别为:100.00%与100.00%;校正集残差分别为:0.0163与0.008 36;预测均方差分别为:0.156与0.0815。结论本方法在样品不经任何预处理的情况下,实现了该制剂的两种有效成分SMZ和TMP的简单、快速、两组分同时准确测定。     

Simultaneous determination of dorzolamide HCL and timolol maleate in eye drops by two different spectroscopic methods

Two-component mixtures of dorzolamide hydrochloride and timolol maleate were assayed by first derivative and ratio derivative spectrophotometric methods. The first method, derivative spectrophotometry, by the zero-crossing measurements, was used due to the drugs closely overlapping absorption spectra. Linear calibration graphs of first derivative values at 250.3 nm for dorzolamide hydrochloride and 315.8 nm for timolol maleate. The second method, is based on ratio first derivative spectrophotometry, the amplitudes in the first derivative of the ratio spectra at 242.9 and at 223.5 nm were selected to determine dorzolamide and timolol maleate in the binary mixture. Calibration graphs were established for 8.0-30.0 microg ml(-1) for dorzolamide hydrochloride and 3.0-24.6 microg ml(-1) for timolol maleate in binary mixture. Good linearity, precision and selectivity were found, and the proposed methods were applied successfully to the pharmaceutical dosage from containing the above-mentioned drug combination without any interference by the excipients. Vierordt's method was also developed for a comparison method.     

Chemometric resolution of a mixture containing hydrochlorothiazide and amiloride by absorption and derivative spectrophotometry

Four chemometric techniques, classical least squares (CLS) and inverse least squares (ILS) and principal component regression (PCR) and partial least squares regression (PLSR) were applied to the absorption and derivative spectrophotometric determinations of amiloride and hydrochlorothiazide in a pharmaceutical preparation. Four chemometric calibrations for both zero-order and first derivative spectra were constructed by measuring the absorbance and their dA/dlambda values at 34 points in the wavelength range 205-395 nm for a training set containing 2-10 microg/ml amiloride and 4-28 microg/ml hydrochlorothiazide corresponding to 25 point mixture design. The building chemometric calibrations were confirmed by using the synthetic mixtures containing two drugs. The results obtained by the proposed techniques based on the use of the measurements at the absorption spectra and at the first derivative spectra were statistically compared with each other.     

Content uniformity and dissolution tests of triplicate mixtures by a double divisor-ratio spectra derivative method

The use of a UV double divisor-ratio spectra derivative calibration for the simultaneous analysis of synthetic samples and commercial tablet preparations without prior separation is proposed. The method was successfully applied to quantify three ternary mixtures, chlorpheniramine maleate and caffeine combined with paracetamol or acetylsalicylic acid and a mixture of acetylsalicylic acid combined with paracetamol and caffeine, using the information in the absorption spectra of appropriate solutions. Beer's law was obeyed in the concentration range of 0.84-4.21 microg/ml for chlorpheniramine maleate, 1.60-15.96 microg/ml for caffeine, 2.0-20.0 microg/ml for acetylsalicylic acid and 1.58-15.93 microg/ml for paracetamol. The whole procedure was applied to synthetic mixtures of pure drugs as well as to commercial preparations (Algon) by using content uniformity and dissolution tests (USP 24) and was found to be precise and reproducible. According to the dissolution profile test more than 84% of paracetamol and caffeine were dissolved within 20 min. Acetylsalicylic acid dissolved more slowly, taking about 45-60 min to dissolve completely. A chemometric method partial least squares (PLS) and a HPLC method were also employed to evaluate the same mixtures. The results of the proposed method were in excellent agreement with those obtained from PLS and HPLC methods and can be satisfactorily used for routine analysis of multicomponent dosage forms.     

Validated stability-indicating methods for determination of cilostazol in the presence of its degradation products according to the ICH guidelines

Sensitive and selective stability-indicating assay methods (SIAMs) are suggested for the determination of cilostazol (CIL) in the presence of its acid, alkaline and oxidative degradation products. Developing SIAMs is necessary to carry out any stability study. Stress testing of CIL was performed according to the International Conference on Harmonization (ICH) guidelines in order to validate the stability-indicating power of the analytical procedures. Stress testing showed that CIL underwent acid, alkaline and oxidative degradation; on the other hand, it showed stability towards photo- and thermal degradation. Two chromatographic SIAMs were developed, namely HPLC and HPTLC methods. The concentration range and the mean percentage recovery were 1.0-31.0 microg/ml and 99.96+/-0.46 and 0.6-14.0 microg/spot and 99.88+/-1.10 for HPLC and HPTLC methods, respectively. In addition, derivative spectrophotometric methods were developed in order to determine CIL in the presence of its acid degradation product; these were performed by using the third derivative spectra (3D) and the first derivative of the ratio spectra (1DD) methods. The linearity range and the mean percentage recovery were 2.0-34.0 microg/ml and 100.27+/-1.20 for the (3D) method, while they were 2.0-30.0 microg/ml and 99.94+/-1.18 for the (1DD) method. Also, two chemometric-assisted spectrophotometric methods, based on using partial least squares (PLS) and concentration residual augmented classical least squares method (CRACLS), for the determination of CIL were developed. Both methods were applied on zero order spectra of the mixtures of CIL and its acid degradation product, the mean percentage recovery was 100.03+/-1.09 and 99.91+/-1.27 for PLS and CRACLS, respectively. All methods were validated according to the International Conference on Harmonization (ICH) guidelines and applied on bulk powder and pharmaceutical formulations.     

Simultaneous spectrofluorimetric and spectrophotometric determination of melatonin and pyridoxine in pharmaceutical preparations by multivariate calibration methods

Partial least-squares (PLS) calibration and principal component regression (PCR) methods were utilized for the simultaneous spectrofluorimetric and spectrophotometric determination of pyridoxine (PY) and melatonin (MT). Since emission and adsorption spectra of these drugs overlap, PY and MT cannot be directly determined by fluorimetric nor by spectrophotometric methods. Full-spectrum multivariate calibration PLS and PCR methods were developed for both fluorimetry and spectrophotometry. The conditions were optimized for fluorimetric as well as for spectrophotometric determination of both drugs. The simultaneous determination of PY and MT was carried out in mixtures by recording the emission fluorescence spectrum between 324 and 500 nm (lambda(ex) 285 nm) for fluorimetry, and by recording the absorption spectrum between 250 and 350 nm for spectrophotometry (lambda(max(PY)) 310 nm, lambda(max(MT)) 278 nm). The experimental calibration matrixes were designed orthogonally. At the optimum conditions, dynamic ranges were 0.04-1.3 and 0.1-4 microg ml(-1) for fluorimetry and 1-22 and 1-24 microg ml(-1) for spectrophotometry for MT and PY, respectively. The calibration concentrations were prepared in the dynamic ranges. The parameters of the chemometrics procedure for the simultaneous determination of MT and PY were optimized, and the proposed methods were validated with prediction set. Finally the procedures were successfully applied to simultaneous spectrofluorimetric and spectrophotometric determination of PY and MT in synthetic mixtures and in a pharmaceutical formulation.     

Simultaneous spectrophotometric determination of pseudoephedrine hydrochloride and ibuprofen in a pharmaceutical preparation using ratio spectra derivative spectrophotometry and multivariate calibration techniques

Spectrophotometric methods are described for the simultaneous determination of pseudoephedrine hydrochloride and ibuprofen in their combination. The obtained data were evaluated by using five different methods. In the first method, ratio spectra derivative spectrophotometry, analytical signals were measured at the wavelengths corresponding to either maximums and minimums for both drugs in the first derivative spectra of the ratio spectra obtained by using each other spectra as divisor in their solution in 0.1 M HCl. In the other four spectrophotometric methods using chemometric techniques, classical least-squares, inverse least-squares, principal component regression and partial least-squares (PLS), the concentration data matrix were prepared by using the synthetic mixtures containing these drugs in methanol:0.1 M HCl (3:1). The absorbance data matrix corresponding to the concentration data matrix was obtained by the measurements of absorbances in the range 240-285 nm in the intervals with deltalambda = 2.5 nm at 18 wavelengths in their zero-order spectra, then, calibration or regression was obtained by using the absorbance data matrix and concentration data matrix for the prediction of the unknown concentrations of pseudoephedrine hydrochloride and ibuprofen in their mixture. The procedures did not require any separation step. The linear range was found to be 300-1300 microg/ml for ibuprofen and 100-1300 microg/ml for pseudoephedrine hydrochloride in all five methods. The accuracy and the precision of the methods have been determined and they have been validated by analyzing synthetic mixtures. The five methods were successfully applied to tablets and the results were compared with each other.     

Rapid and simultaneous determination of sulfamethoxazole and trimethoprim in human plasma by high-performance liquid chromatography

A simple and reproducible high-performance liquid chromatographic method was developed for simultaneous determination of sulfamethoxazole (SMX) and trimethoprim (TMP) in human plasma. The method entailed injection of the samples after deproteination with perchloric acid and subsequent neutralizing. Primidone was used as internal standard. Chromatography was performed on a C(18) column (250 mm x 4.6 mm, 5 microm) under isocratic elution with 50 mM aqueous sodium dihydrogen phosphate-acetonitrile-triethylamine (100:25:0.5, v/v), pH 5.9. Detection was made at 240 nm and analyses were run at a flow-rate of 1.5 ml/min at a temperature of 35 degrees C. The recovery was 83.4, 88.5 and 98.2% for TMP, SMX and internal standard, respectively. The precision of the method was 2.6-9.8% over the concentration range of 0.125-2 microg/ml for TMP and 0.39-50 microg/ml for SMX. The limit of quantification (LOQ) in plasma was 0.125 and 0.39 microg/ml for TMP and SMX, respectively. The method was used for a bioequivalence study.     

A comparative study of the ratio spectra derivative spectrophotometry, Vierordt's method and high-performance liquid chromatography applied to the simultaneous analysis of caffeine and paracetamol in tablets

Two spectrophotometric methods and high-performance liquid chromatography were proposed for the simultaneous analysis of caffeine and paracetamol in a tablet formulation. The ratio spectra derivative method is based on the use of the analytical signals obtained by measuring at 267.9 and 291.0 nm for caffeine and 237.0 and 251.8 nm for paracetamol in the first derivative of the ratio spectra. Calibration graphs were prepared in the range 4-40 microg/ml for caffeine and 8-48 microg/ml for paracetamol. In Vierordt's method, A1(1) (1%, 1cm) values of paracetamol and caffeine were determined at 242.9 and 273.0 nm in zero-order spectra. The matrix for A1(1) (1%, 1cm) values was written and the amounts of both drugs were calculated by means of the program 'Matlab' software. The results obtained by these spectrophotometric methods were compared with the results of HPLC method.     

A flow injection sensor for simultaneous determination of sulfamethoxazole and trimethoprim by using Sephadex SP C-25 for continuous on-line separation and solid phase UV transduction

A flow-through sensor based on integration of spectrophotometric detection and the different kinetics of retention/elution of analytes on a solid support is proposed for the simultaneous determination of sulfamethoxazole (SMZ) and trimethoprim (TMP). The solid support (Sephadex SP C-25) fills both, a microcolumn placed on-line and the sensing microzone. The intrinsic absorbance of both compounds is monitored directly on the solid phase at 269 nm and so, no derivatization step is required. Using two alternate solutions, 10(-4) M hydrochloric acid and 0.20 M NaAc/HAc (pH 5.0) buffer, the sensor responds linearly in the measuring range of 50-250 and 10-70 microg ml(-1) with detection limits of 9.5 and 0.6 microg ml(-1) (500 microl of sample volume) for SMZ and TMP, respectively. The main advantages of the sensor are simplicity, rapidity and low reagents consumption. Its application to SMZ and TMP determination in synthetic samples and pharmaceutical preparations is demonstrated. The results obtained by the proposed method were compared with those obtained by a standard HPLC method.     

Application of TLC-densitometry, first-derivative UV-spectrophotometry and ratio derivative spectrophotometry for the determination of dorzolamide hydrochloride and timolol maleate

Three methods are described for the simultaneous determination of dorzolamide hydrochloride (DORZ) and timolol maleate (TIM) in ophthalmic solutions. The first method is based on application of thin layer chromatographic separation of both drugs followed by the densitometric measurements of their spot areas. After separation on silica gel GF(254) plates, using methanol-ammonia 25% (100:1.5 v/v) as the mobile phase, the chromatographic zones corresponding to the spots were scanned at 253 and 297 nm, respectively. The calibration function was established in the ranges of 2-18 microg for DORZ and 0.5-4.5 microg for TIM. The second method depends on first derivative ultraviolet spectrophotometry, with zero-crossing measurement method. The first derivative values D(1) at 250.2 and 312.5 nm were selected for the assay of DORZ and TIM, respectively. Calibration graphs follow Beer's law in the range 10-64 and 2.5-16 microg ml(-1), respectively. The third method is based on ratio first derivative spectrophotometry. The signals in the first derivative of the ratio spectra at 244 and 306.2 nm were selected to determine DORZ and TIM in the mixture and calibration graphs are linear in the range of 5-40 and 5.0-17.5 microg ml(-1), respectively. The proposed methods were successfully applied to the determination of these compounds in synthetic mixtures and in pharmaceutical preparations. The proposed methods are simple, rapid and suitable for quality control application.     

Simultaneous determination of loratadine and pseudoephedrine sulfate in pharmaceutical formulation by RP-LC and derivative spectrophotometry

Highly sensitive, simple and accurate reversed phase liquid chromatographic and first derivative spectrophotometric methods for determination of antihistaminic drug loratadine [I] and nasal decongestant drug pseudoephedrine sulfate [II] are described. The HPLC method involves separation of [I] and [II] on micro-BondaPak C18 column using mixture of (methanol:H(2)O:phosphoric acid:ammonium dihydrogen phosphate) (220:300:2:3 g) (V/V/V/W), 60 and 40% acetonitrile as mobile phase flowing at 2 ml/min with ultraviolet detection at 247 nm. The calibration graphs are linear from 5 to 100 microg/ml for [I] and from 120 to 1200 microg/ml for [II] the detection limits are 0.5 microg/ml for [I] and 60 microg/ml for [II]. The spectrophotometric method is based on recording the first derivative spectra for [I] and [II] at 307, 266 nm, respectively, of their solutions in 0.1 M hydrochloric acid using the acid as blank. The calibration graphs are linear in the range of 5-25 microg/ml for [I] and 240-720 microg/ml for [II]; the limits of detection are 0.16 microg/ml for [I] and 10 microg/ml for [II]. The mean percentage recoveries obtained for different synthetic mixtures by using this method are 97.6% with coefficient of variation 1.79 for [I] and 101.6% with coefficient of variation 1.95 for [II]. The two methods have been applied successfully for the determination of [I] in its combination with [II] Clarinase tablets and [I] alone in different pharmaceutical dosage forms.     

Simultaneous determination of sulphamethoxazole and trimethoprim in powder mixtures by attenuated total reflection-Fourier transform infrared and multivariate calibration

A partial least-squares calibration (PLS) procedure in combination with infrared spectroscopy has been developed for simultaneous determination of sulphamethoxazole (SMZ) and trimethoprim (TMP) in raw material powder mixtures used for manufacturing commercial pharmaceutical products. Multivariate calibration modeling procedures, interval partial least squares (iPLS) and synergy partial least squares (siPLS), were applied to select a spectral range that provided the lowest prediction error in comparison to the full-spectrum model. The experimental matrix was constructed using 49 synthetic samples and 15 commercial samples. The considered concentration ranges were 400–900 mg g⁻¹ SMZ and 80–240 mg g⁻¹ TMP. Spectral data were recorded between 650 and 4000 cm⁻¹ with a 4 cm⁻¹ resolution by Fourier transform infrared spectroscopy coupled with attenuated total reflectance (ATR-FTIR) accessory. The proposed procedure was compared with conventional procedure by high performance liquid chromatography (HPLC) using 15 commercial samples containing SMZ and TMP. The results showed that PLS regression model combined to ATR-FTIR is a relatively simple, rapid and accurate procedure that could be applied to the simultaneous determination of SMZ and TMP in routine quality control of powder mixtures. A root mean square error of prediction (RMSEP) of 13.18 mg g⁻¹ for SMZ and 6.03 mg g⁻¹ for TMP was obtained after selection of better intervals by siPLS. Using the proposed procedure it is possible to analyze each sample in less than 3 min considering two replicates (excluding the grinding step). Accuracy was checked by comparison to HPLC method and agreement better than 98.8% was achieved.     

Application of some chemometric methods in conventional and derivative spectrophotometric analysis of acetaminophen and ascorbic acid

The multivariate methods, principal component regression (PCR) and partial least squares (PLS) were tested as a calibration procedure for simultaneous ultraviolet spectrophotometric determination of acetaminophen (AC) and ascorbic acid (AA). Determination of these compounds is important because of their pharmacotherapeutic advantages. Due to spectral overlapping of AC and AA, PCR and PLS were used for construction of the calibration sets. The concentration linear range of AC and AA were 1.5–24.2 and 1.8–21.1 µg mL^?1 respectively. The absorption spectra were recorded from 215–310 nm. The minimum root mean square error of prediction (RMSEP) was 1.3507 and 0.4088 for AC and AA, by PLS, 0.7525 and 0.4015 by PCR in original data and 0.9454 and 0.2875, by PLS and 1.0386 and 0.4000 by PCR in derivative data. The procedure allows the simultaneous determination of AC and AA in synthetic mixtures and real sample solutions made up from pharmaceutical products, human serum and urine. Copyright © 2010 John Wiley & Sons, Ltd.     

Two derivative spectrophotometric methods for the simultaneous determination of lovastatin combined with three antioxidants

A zero crossing and an algorithm bivariate calibration derivative method for the simultaneous determination of lovastatin combined separately with three antioxidants (ascorbic acid, quercetin and gallic acid) in synthetic mixtures are described. The aqueous or methanolic solutions obeyed Beer's law in the concentration ranges of 3.20–17.36 μg/ml for lovastatin, 1.76–8.80 μg/ml for ascorbic acid, 1.41–7.04 μg/ml for gallic acid and 1.84–9.20 μg/ml for quercetin, for both methods, respectively. In the second derivative (²D) zero crossing method measurements were carried out at 238.4 nm for lovastatin and 265.6 nm for ascorbic acid, 247.7 nm for lovastatin and 281.1 nm for quercetin, 251.8 nm for lovastatin and 267.6 nm for gallic acid. In the first derivative (¹D) bivariate spectrophotometric method an optimum pair of wavelengths was chosen for the determination of different binary mixtures. The proposed procedures were successfully applied to the simultaneous determination of lovastatin and different antioxidants in mixtures with high percentage of recovery, 98.3–100.4% for lovastatin, 98.3–98.6% for ascorbic acid, 99.0–99.8% for quercetin and 100.5–101.1% for gallic acid and good precision. In addition, the results from the above procedures were verified by using partial least-squares (PLS) multivariate calibration method.     

非相关成分参比一倍率导数光谱法测定制剂中甲氧苄啶和磺胺甲(口恶)唑

提出了非相关成分参比一倍率导数光谱法用于测定混合组分体系的基本原理和实验方法。本法可用于待测成分和非相关成分导数光谱严重重叠且待测成分含量较低时的不经分离直接测定。试用本法消除了甲氧苄啶(TMP)和磺胺甲嚼唑(SMZ)及附加剂之间的相互干扰,从而测定了部分制剂中的TMP:用“零交法”测定了SMZ,平均回收率分别为:102.5±1.63%(CV)和100.3±0.99%(CV)。     

Simultaneous spectrophotometric determination of mefenamic acid and paracetamol in a pharmaceutical preparation using ratio spectra derivative spectrophotometry and chemometric methods

Four new methods are described for the simultaneous determination of mefenamic acid (MEF) and paracetamol (PAR) in their combination. In the first method, ratio spectra derivative method, analytical signals were measured at the wavelengths corresponding to either maximums or minimums for both drugs in the first derivative spectra of the ratio spectra obtained by dividing the standard spectrum of one of two drugs in 0.1 M NaOH:methanol (1:9). In the chemometric techniques, classical least-squares, inverse least-squares and principal component regression (PCR), the training was randomly prepared by using the different mixture compositions containing two drugs in 0.1 M NaOH:methanol (1:9). The absorbance data was obtained by the measurements at 13 points in the wavelength range 235–355 nm in the absorption spectra. Chemometric calibrations were constructed by the absorbance data and training set for the prediction of the amount of MEF and PAR in samples. In the third chemometric method, PCR, the covariance matrix corresponding to the absorbance data was calculated for the basis vectors and matrix containing the new coordinates. The obtained calibration was used to determine the title drugs in their mixture. Linearity range in all the methods was found to be 2–10 μg/ml of MEF and 4–20 μg/ml of PAR. Mean recoveries were found satisfactory (>99%). The procedures do not require any separation step. These methods were successfully applied to a pharmaceutical formulation, tablet, and the results were compared with each other.     

Simultaneous determination of paracetamol and methocarbamol in tablets by ratio spectra derivative spectrophotometry and LC

The application of the ratio spectra derivative spectrophotometry and high-performance liquid chromatography (HPLC) to the simultaneous determination of paracetamol (PAR) and methocarbamol (MET) in combined pharmaceutical tablets is presented. The spectrophotometric procedure is based on the use of the first derivative of the ratio spectra obtained by dividing the absorbtion spectrum of the binary mixtures by a standard spectrum of one of the compounds. The first derivative amplitudes were measured at 243.0 and 230.3 nm for the assay of PAR and MET, respectively. Calibration graphs were established for 2-30 microg ml for PAR and 2-36 microg/ml for MET in binary mixture. The detection limits for PAR and MET were found 0.097 and 0.079 microg/ml, respectively; while the quantification limits were 0.573 microg/ml for PAR and 1.717 microg/ml for MET. For the HPLC procedure, a reversed-phase column with a mobile phase of methanol-water (60:40, v/v), was used to separate both compounds with a detection of 274.0 nm. Linearity was obtained in the concentration range of 2 300 and 1.5-375 microg/ml for PAR and MET, respectively. The detection and quantification limits were found to be 0.42 and 1.4 microg/ml for PAR and 0.36 and 1.2 microg/ml for MET, respectively. The relative standard deviations were found to be less than 0.52%, indicating reasonable repeatibility of both methods. The proposed methods were successfully applied to the determination of these drugs in commercial tablets.