Simultaneous determination of pteridines in multicomponent mixtures using derivative spectrophotometry and partial least-squares calibration

Simple binary mixtures composed of very similar pteridines (neopterin and pterin) have been resolved by derivative spectrophotometry. Detection limits of 0.30 μg ml⁻¹ and 0.12 μg ml⁻¹ for pterin and neopterin, respectively, have been calculated. Also, different mixtures of pteridines considered as disease markers, such as pterin, neopterin, xanthopterin and isoxanthopterin, have been determined by using a partial least-squares (PLS-2) model. Calibration set containing 0–7 μg ml⁻¹ for each component was used. The resolution of several mixtures and single determination was tested in artificial samples.     

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.     

Simultaneous diffuse reflectance infrared determination of clavulanic acid and amoxicillin using multivariate calibration techniques

A method for simultaneous determination of clavulanic acid (CA) and amoxicillin (AMO) in commercial tablets was developed using diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS) and multivariate calibration. Twenty-five samples (10 commercial and 15 synthetic) were used as a calibration set and 15 samples (10 commercial and 5 synthetic) were used for a prediction set. Calibration models were developed using partial least squares (PLS), interval PLS (iPLS), and synergy interval PLS (siPLS) algorithms. The best algorithm for CA determination was siPLS model with spectra divided in 30 intervals and combinations of 2 intervals. This model showed a root mean square error of prediction (RMSEP) of 5.1 mg g(-1). For AMO determination, the best siPLS model was obtained with spectra divided in 10 intervals and combinations of 4 intervals. This model showed a RMSEP of 22.3 mg g(-1). The proposed method was considered as a suitable for the simultaneous determination of CA and AMO in commercial pharmaceuticals products.     

Simultaneous spectrophotometric determination of diclofenac potassium and methocarbamol in binary mixture using chemometric techniques and artificial neural networks

In this study, the simultaneous determination of diclofenac potassium (DP) and methocarbamol (MT) by chemometric approaches and artificial neural networks using UV spectrophotometry has been reported as a simple alternative to using separate models for each component. Three chemometric techniques-classical least-squares (CLS), principal component regression (PCR), and partial least-squares (PLS)-along with radial basis function-artificial neural network (RBF-ANN) were prepared by using the synthetic mixtures containing the two drugs in methanol. A set of synthetic mixtures of DP and MT was evaluated and the results obtained by the application of these methods were discussed and compared. In CLS, PCR, and PLS, the absorbance data matrix corresponding to the concentration data matrix was obtained by the measurements of absorbances in the range 260-310 nm in the intervals with Δλ = 0.2 nm 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 DP and MT in their mixtures. In RBF-ANN, the input layer consisting of 251 neurons, 9 neurons in the hidden layer, and 2 output neurons were found appropriate for the simultaneous determination of DP and MT. The accuracy and the precision of the four methods have been determined and they have been validated by analyzing synthetic mixtures containing the two drugs. The proposed methods were successfully applied to a pharmaceutical formulation containing the examined drugs.     

Simultaneous determination of metronidazole and spiramycin in bulk powder and in tablets using different spectrophotometric techniques

Metronidazole (MZ) is an anti-infective drug used in the treatment of anaerobic bacterial and protozoa infections in humans. It is also used as a veterinary antiparasitic drug. Spiramycin (SP) is a medium-spectrum antibiotic with high effectiveness against Gram-positive bacteria. Three simple, sensitive, selective and precise spectrophotometric methods were developed and validated for the simultaneous determination of MZ and SP in their pure form and in pharmaceutical formulations. In methods A and B, MZ was determined by the application of direct spectrophotometry and by measuring its zero-order (D(0)) absorption spectra at its λ(max) = 311 nm. In method A, SP was determined by the application of first derivative spectrophotometry (D(1)) and by measuring the amplitude at 218.3 nm. In method B, the first derivative of the ratio spectra (DD(1)) was applied, and SP was determined by measuring the peak amplitude at 245.6 nm. Method C entailed mean centering of the ratio spectra (MCR), which allows the determination of both MZ and SP. The methods developed were used for the determination of MZ and SP over a concentration range of 5-25 μg ml(-1). The proposed methods were used to determine both drugs in their pure, powdered forms with mean percentage recoveries of 100.16 ± 0.73 for MZ in methods A and B, 101.10 ± 0.90 in method C, 100.09 ± 0.70, 100.02 ± 0.88 and 100.49 ± 1.26 for SP in methods A, B and C, respectively. The proposed methods were proved using laboratory-prepared mixtures of the two drugs and were successfully applied to the analysis of MZ and SP in tablet formulation without any interference from each other or from the excipients. The results obtained by applying the proposed methods were compared statistically with a reported HPLC method and no significant difference was observed between these methods regarding both accuracy and precision.     

Simultaneous determination of chlordiazepoxide and clidinium bromide in pharmaceutical formulations by derivative spectrophotometry.

A direct and simple first derivative spectrophotometric method has been developed for the simultaneous determination of clidinium bromide and chlordiazepoxide in pharmaceutical formulations. Acetonitrile was used as solvent for extracting the drugs from the formulations and subsequently the samples were evaluated directly by derivative spectrophotometry. Simultaneous determination of the drugs can be carried out using the zero-crossing method for clidinium bromide at 220.8 nm and the graphical method for chlordiazepoxide at 283.6 nm. The calibration graphs were linear in the ranges from 0.983 to 21.62 mg/l of clidinium bromide and from 0. 740 to 12.0 mg/l of chlordiazepoxide. The ingredients commonly found in commercial pharmaceutical formulations do not interfere. The proposed method was applied to the determination of these drugs in tablets.     

Simultaneous determination of fosinopril and hydrochlorothiazide in pharmaceutical formulations by spectrophotometric methods

Three new spectrophotometric procedures for the simultaneous determination of fosinopril and hydrochlorothiazide are described. The first method, derivative-differential spectrophotometry, comprised of measurement of the difference absorptivities derivatized in the first-order (ΔD₁) of a tablet extract in 0.1 N NaOH relative to that of an equimolar solution in methanol at wavelengths of 227.6 and 276.4 nm, respectively. The second method, depends on the application ratio spectra derivative spectrophotometric method to resolve the interferance due to spectral overlapping. The analytical signals were measured at 237.9, 243.8 nm for fosinopril and 262.4, 269.3 and 278.6 nm for hydrochlorothiazide in the binary mixture, in the first derivative of the ratio spectra of the mixture solutions in methanol. Calibration graphs were established for 4.0–50.0 μg ml⁻¹ fosinopril and 2.0–14.0 μg ml⁻¹ hydrochlorothiazide in binary mixture. The third method, absorbance ratio method, the determination of fosinopril and hydrochlorothiazide was performed by using the absorbances read at 210.0, 219.5 and 271.7 nm in the zero-order spectra of their mixture. The developed methods were compared with absorbance ratio method. Application of the suggested procedures were successfully applied to the determination of this compound in synthetic mixtures and in pharmaceutical preparations, with high percentage of recovery, good accuracy and precision.     

Simultaneous spectrophotometric determination of paracetamol and para-aminophenol in pharmaceutical dosage forms using two novel multivariate standard addition methods based on net analyte signal and rank annihilation factor analysis

Recently we proposed two multivariate standard addition methods using net analyte signal concept and rank annihilation factor analysis, which we named SANAS and SARAF, respectively. These methods could model both indirect sample matrix effects and direct interference effects of the coexisting analytes. Here, the potential of these methods for simultaneous determination of paracetamol and para-aminophenol in the synthetic mixtures and pharmaceutical products was examined. It was found that both models could predict the concentration of analytes in synthetic mixtures with relative errors of prediction lower than 7%. Moreover, the employed methods exhibited the recoveries in the range of 93.11 and 108.67 for analysis of the drugs in the real matrices. From SANAS plots, the figures of merit including sensitivity, selectivity and limit of detection were calculated as 0.143 μg/ml, 0.587 and 0.018 μg/ml for paracetamol and 0.128 μg/ml, 0.547 and 0.020 μg/ml for para-aminophenol, respectively.     

Simultaneous determination of rifampicin, isoniazid and pyrazinamide in tablet preparations by multivariate spectrophotometric calibration

The use of multivariate spectrophotometric calibration is presented for the simultaneous determination of the active components of tablets used in the treatment of pulmonary tuberculosis. The resolution of ternary mixtures of rifampicin, isoniazid and pyrazinamide has been accomplished by using partial least squares (PLS-1) regression analysis. Although the components show an important degree of spectral overlap, they have been simultaneously determined with high accuracy and precision, rapidly and with no need of nonaqueous solvents for dissolving the samples. No interference has been observed from the tablet excipients. A comparison is presented with the related multivariate method of classical least squares (CLS) analysis, which is shown to yield less reliable results due to the severe spectral overlap among the studied compounds. This is highlighted in the case of isoniazid, due to the small absorbances measured for this component.     

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 phenobarbital, phenytoin and methylphenobarbital in pharmaceutical preparations by using partial least-squares and principal component regression multivariate calibration

Two multivariate calibration methods, partial least squares (PLS-2) and principal component regression (PCR) have been applied to the simultaneous spectrophotometric analysis of ternary mixtures of phenytoin (DPH), phenobarbital (PBT) and methylphenobarbital (MPBT) in the Comital-L pharmaceutical formulation. The PLS-2 and PCR procedures were employed to evaluate the data of a variable number of calibration solutions measured over the wavelength range 400-700 nm. The concentration ranges used to construct the calibration matrix were varied between 5 and 30 microg ml(-1). The proposed methods were validated by applying them to the analysis of the Comital-L pharmaceutical formulation and the average relative errors were less than 6% for each one of the analyzed compounds. The results obtained by both proposed methods have been compared with the results obtained by application of a RPLC reference method.     

HPLC法测定复方罗布麻片Ⅰ中盐酸异丙嗪和氯氮(艹卓)的含量

目的:采用高效液相色谱法考察并建立了测定复方罗布麻片Ⅰ中盐酸异丙嗪和氯氮(艹卓)含量的方法.方法:色谱柱为Phenomenex-ODS3柱(250×4.60mm,5μm),甲醇-磷酸盐缓冲溶液(取磷酸二氢胺2.64g,加水1 000mL溶解后,用磷酸调节pH值至3.0)(55:45)为流动相,流速1.0mL/min,检测波长为250nm,进样量20μL.结果:盐酸异丙嗪的线性范围为5.1～60.8μg/mL(r=1.000 0),氯氮(艹卓)的线性范围为5.2～60.4μg/mL(r=0.999 9),平均回收率分别为99.5%和101.2%.结论:本法精密度好,结果准确可靠,适用于该复方制剂的质量检验分析.     

HPLC法测定复方罗布麻片Ⅰ中盐酸异丙嗪和氯氮的含量

目的:采用高效液相色谱法考察并建立了测定复方罗布麻片Ⅰ中盐酸异丙嗪和氯氮(?)含量的方法。方法:色谱柱为Phenomenex-ODS3柱(250 ×4.60mm,5μm),甲醇-磷酸盐缓冲溶液(取磷酸二氢胺2.64g,加水1 000mL溶解后,用磷酸调节pH值至3.0)(55:45)为流动相,流速1.0mL/min,检测波长为250nm,进样量20μL。结果:盐酸异丙嗪的线性范围为5.1-60.8μg/mL(r=1.000 0),氯氮(?)的线性范围为5.2-60.4μg/mL(r=0.999 9),平均回收率分别为99.5％和101.2％。结论:本法精密度好,结果准确可靠,适用于该复方制剂的质量检验分析。     

Utility of certain π-acceptors for the spectrophotometric determination of perindopril

Simple, rapid, accurate and sensitive spectrophotometric methods are described for the determination of perindopril. The methods are based on the reaction of this drug as n-electron donor with 2,3-dichloro-5,6-dicyano-p-benzoquinone(DDQ)-7,7,8,8-tetracyanoquinodimethane (TCNQ), tetracyanoethylene (TCNE), chloranil (CL) and p-chloranilic acid (p-CA) as π-acceptors to give highly coloured complex species. The coloured products are measured spectrophotometrically at 588, 843, 419, 550 and 520 nm for DDQ, TCNQ, TCNE, CL and p-CA, respectively, optimization of different experimental conditions is described. Beer's law is obeyed in the range of 20–200 μg ml⁻¹ and colours were produced in non-aqueous media and were stable for at least 1 h. Application of the suggested methods to perindopril tablets are presented.     

Simultaneous determination of potassium and total fluoride in toothpastes using a SIA system with two potentiometric detectors

A sequential injection analysis (SIA) system has been developed for the first time to quantify potassium and total fluoride in toothpastes and gels used to prevent both dentinal hypersensitivity and dental caries. To enable this simultaneous determination, potentiometric detection, using a conventional fluoride electrode and a tubular potassium selective electrode, formed by a PVC membrane containing valinomycin as ionophore, was carried out. A manifold that uses a three-way solenoid valve was designed. The former under binary sampling conditions, provides reproducible mixing ratios of two solutions. This fact facilitates that the system automatically generates, on-line, the calibration curves required by the analytical procedure. The calibration ranged from 1.0 × 10⁻⁴ to 1.0 × 10⁻³ mol L⁻¹ for both potassium and total fluoride determinations. The R.S.D. (11 readings) resulted to be less than 1.5% for both determinations. Off-line studies related to the dissolution of the solid samples, the transformation of monofluorophosphate in fluoride, the elimination of organic matrix interference onto the plastic membrane of the potassium electrode, and the selection of the most adequate TISAB solution for fluoride determination, were also considered. A sampling rate of 18 samples h⁻¹ for both determinations was attained, their precisions and accuracies being statistically indistinguishable from those achieved by atomic emission spectroscopy (for potassium determination) and by a conventional batch potentiometry (for total fluoride determination) adopted as reference techniques.     

Use of xylenol orange and cetylpyridinium chloride in rapid spectrophotometric determination of zinc in pharmaceutical products

A simple, rapid, and sensitive spectrophotometric method for the determination of zinc(II) is performed, based on colour reaction between the metal ion and xylenol orange in the presence of surfactant cationic cetylpyridinium chloride. The important analytical parameters and their effects on the reported system are investigated. Zinc(II) reacts with the reagent and surfactant in the ratio 1:2:4 (metal:ligand:surfactant) in the pH range 5.0–6.0 to form a ternary complex with an absorption maximum at 580 nm. The reaction was extremely rapid at room temperature, and the absorbance value remains unchanged for at least 168 h. The apparent stability constant of the complex was found to be K=1.05×10¹⁰, and the method adheres to Beer's law for 1–20 μg zinc(II) per 25 ml with apparent molar absorptivity of 1.1×10⁴ l mol cm⁻¹. The effect of foreign ions was tested by taking a constant concentration of metal ion and determining its concentration in the presence of large number of foreign ions. The method was applied for determination of zinc(II) in dermal ointments where excellent agreement between reported and obtained results were achieved. The relative standard deviation was better than 2%.     

Blend uniformity end-point determination using near-infrared spectroscopy and multivariate calibration

A multivariate calibration approach using near-infrared (NIR) spectroscopy for determining blend uniformity end-point of a pharmaceutical solid dosage form containing 29.4% (w/w) drug load with three major excipients (crospovidone, lactose, and microcrystalline cellulose) is presented. A set of 21 off-line, static calibration samples were used to develop a multivariate partial least-squares (PLS) calibration model for on-line predictions of the API content during the blending process. The concentrations of the API and the three major excipients were varied randomly to minimize correlations between the components. A micro-electrical-mechanical-system (MEMS) based NIR spectrometer was used for this study. To minimize spectral differences between the static and dynamic measurement modes, the acquired NIR spectra were preprocessed using standard normal variate (SNV) followed by second derivative Savitsky-Golay using 21 points. The performance of the off-line PLS calibration model were evaluated in real-time on 67 production scale (750L bin size) blend experiments conducted over 3 years. The real-time API-NIR (%) predictions of all batches ranged from 93.7% to 104.8% with standard deviation ranging from 0.5% to 1.8%. These results showed the attainment of blend homogeneity and were confirmed with content uniformity by HPLC of respective manufactured tablets values ranging from 95.4% to 101.3% with standard deviation ranging from 0.5% to 2.1%. Furthermore, the performance of the PLS calibration model was evaluated against off-target batches manufactured with high and low amounts of water during the granulation phase of production. This approach affects the particle size and hence blending. All the off-target batches exhibited API-NIR (%) predictions of 94.6% to 103.5% with standard deviation ranging from 0.7% to 1.9%. Using off-target data, a systematic approach was developed to determine blend uniformity end-point. This was confirmed with 3 production scale batches whereby the blend uniformity end-point was determined using the PLS calibration model. Subsequently, the uniformity was also ascertained with conventional thief sampling followed by HPLC analysis and content uniformity by HPLC of the manufactured tablets.     

差示分光光度法测定眼用制剂抑菌剂聚季铵盐-1的含量

目的:建立简便、快捷、准确测定眼用制剂中抑菌剂聚季铵盐-1含量的方法。方法:应用聚季铵盐-1和台盼蓝之间反应发生红移现象,采用差示分光光度法,以台盼蓝溶液作为参比液,在波长681 nm处,测得台盼兰在不同浓度聚季铵盐-1溶液中的差示吸光度。结果:聚季铵盐-1浓度在5.00～15.00μg.mL-1范围内,最大差示吸光度△A与溶液浓度呈良好线性关系(r=0.9996),平均回收率(n=9)为97.8%,RSD=0.87%。结论:该方法操作简便、快捷,成本低,结果准确,适用于测定眼用制剂中聚季铵盐-1的含量。     

Simultaneous spectrofluorimetric determination of amlodipine besylate and valsartan in their combined tablets

Amlodipine, a dihydropyridine calcium channel blocker, and valsartan, an angiotensin II receptor blocker, are co‐formulated in a single‐dose combination for the treatment of hypertension. The combination is used by patients whose blood pressure is not adequately controlled on either component monotherapy. This work describes a simple, sensitive, and reliable spectrofluorimetric method for the simultaneous determination of the two antihypertensive drugs; amlodipine besylate (AML) and valsartan (VAL) in their combined tablets. The method involved measurement of the native fluorescence at 455 nm (λ_Ex 360 nm) and 378 nm (λ_Ex 245 nm) for AML and VAL, respectively. Analytical performance of the proposed spectrofluorimetric procedure was statistically validated with respect to linearity, ranges, precision, accuracy, selectivity, robustness, detection, and quantification limits. Regression analysis showed good correlation between fluorescence intensity and concentration over the concentration ranges 0.2–3.6 and 0.008–0.080 µg mL^?1 for AML and VAL, respectively. The limits of detection were 0.025 and 0.0012 µg mL^?1 for AML and VAL, respectively. The proposed method was successfully applied for the assay of the two drugs in their combined pharmaceutical tablets with recoveries not less than 98.85%. No interference was observed from common pharmaceutical additives. The results were favourably compared with those obtained by a reference spectrophotometric method. Copyright © 2010 John Wiley & Sons, Ltd.