Different methods for the determination of gestodene, and cyproterone acetate in raw material and dosage forms

Four new precise accurate and selective methods have been developed for the determination of gestodene (I) and cyproterone acetate (II). The first method (A) depends on reaction of (I) and (II) with isoniazide in an acid medium and the colored products were measured at 378 and 400 nm, respectively. The second method (B) depends on the reaction of (I) and (II) with tetrazolium blue in an alkaline medium and the colored products were measured quantitatively at 515 and 520 nm, respectively. The optimum conditions for the analysis were studied. Both methods determined gestodene (I) in concentration range from 4 to 24 microg ml(-1) with mean percentage recoveries 99.54%+/-1.20 and 99.63%+/-1.89 for method A and B, respectively. For cyproterone acetate, the concentration ranges were 4-36 and 8-40 microg ml(-1) with mean percentage recoveries 99.94%+/-1.19 and 99.23%+/-2.00 for methods A and B, respectively. The third method (C) depends on the quantitative evaluation of (I) and (II) densitometrically using dichloroethane:methanol:water (95:5:0.2) as mobile phase and the chromatogram were scanned at 247 and 281 nm, respectively. Method (C) determines (I) and (II) in concentration ranges from 0.2 to 1.6 and 0.1-0.7 microg microl(-1) using Hamilton syringe 10 microl, with mean percentage recoveries 99.94%+/-1.19, and 99.82%+/-1.75, respectively. The fourth method (D) is a first derivative one depends on measuring the D(1) value at 303 nm for (II) only in concentration range 10-20 microg ml(-1) with mean percentage recoveries 99.95%+/-1.49.     

Stability indicating methods for the determination of diloxanide furoate

Five new selective, precise and accurate methods are described for the determination of diloxanide furoate (DI) in presence of its degradation products. Method A utilizes the first and second derivative spectrophotometry at 270 and 280 nm, respectively. Method B is a RSD(1) spectrophotometric method based on the simultaneous use of the first derivative of ratio spectra and measurement at 270 nm. Method C is a pH-induced difference spectrophotometry using UV measurement at 295 nm. Method D is a densitometric one, after separation on silica gel plate using chloroform: methanol as mobile phase and the spots were scanned at 258 nm. Method E is reversed phase high performance liquid chromatography using methanol: water (80:20% v/v) as mobile phase at a flow rate of 1 ml/min and UV detection at 258 nm. Regression analysis showed good correlation in the concentration ranges 5-30, 5-25, 10-40 microg/ml, 100-500 ng/spot, 2-50 microg/ml with percentage recoveries of 99.92+/-0.56 and 99.79+/-0.47, 99.23+/-0.38, 99.96+/-0.06, 99.03+/-0.51, 98.81+/-0.68 for methods A, B, C, D and E, respectively. These methods are suitable as stability indicating methods for the determination of DI in presence of its degradation products either in bulk powder or in pharmaceutical formulations.     

Stability-indicating methods for the determination of doxazosin mezylate and celecoxib

Two stability-indicating methods were developed for the determination of doxazosin mesylate (I) and celecoxib (II) in the presence of their degradation products. The first method depends on the use of first derivative spectrophotometry (D(1)) at 256, 269 nm for (I) and (II), respectively. This method determines (I) and (II) in concentration ranges of 0.8-12 and 1-20 microg ml(-1) with mean percentage accuracies of 99.21+/-0.88 and 99.59+/-1.67% for (I) and (II), respectively. The second method depends on the quantitative densitometric evaluation of thin-layer chromatography of (I) and (II) in the presence of their degradation products without any interference. Methylisobutyl ketone-glacial acetic acid-water (20:10:10) was used as a mobile phase for (I) and cyclohexane-dichloromethane-diethyleamine (50:40:10) for (II). The chromatograms were scanned at 248 and 253 nm for (I) and (II), respectively. This method determines (I) and (II) in concentration ranges of l-4 microg per spot for both drugs with mean percentage accuracies of 100.19+/-0.95 and 99.91+/-1.95% for (I) and (II), respectively. The suggested methods were used to determine doxazosin mesylate and celecoxib in bulk powder, laboratory-prepared mixtures and pharmaceutical dosage forms (cardura tablet and celebrex capsule). The results obtained by applying the proposed methods were statistically analysed and compared with those obtained by the reported methods.     

Quantitative determination of piroxicam in a new formulation (piroxicam-beta-cyclodextrin) by derivative UV spectrophotometric method and HPLC

A derivative ultraviolet (UV) spectrophotometric method for the determination of piroxicam in piroxicam--beta-cyclodextrin tablets was developed. Phosphate buffer (pH 7.8, 0.1 M) and ethanol were used as a solvent system throughout the study. In this study, determination of piroxicam was conducted by using first order derivative amplitudes at 261.4 nm (n=4). Standards for the calibration graph ranging from 2.40 to 20.0 microg/ml were prepared from working standard. The proposed method is accurate with 99.70%+/-0.50 recovery value and precise with coefficient of variation (CV) of 1.29. The results were compared with those obtained using a high-performance liquid chromatography (HPLC) procedure. A reversed-phase C(18) column with aqueous phosphate buffer:methanol, 60:40, v/v, mobile phase was used. UV detector was set at 254 nm. Calibration solutions used in HPLC were ranging from 5 to 20 microg/ml. Results obtained in HPLC were comparable to those obtained by derivative UV spectrophotometric method.     

First-derivative spectrophotometric and LC determination of cefuroxime and cefadroxil in urine

Two methods are presented for the determination of cefuroxime and cefadroxil in human urine using first (1D) derivative spectrophotometry and high-performance liquid chromatography. Cefuroxime and cefadroxil were determined by measurement of their first-derivative amplitude in 0.1 N sodium hydroxide at 292.5 and 267.3 nm, respectively in the concentration range of 2-10 microg ml(-1) for each drug. The HPLC method depends upon using a LiChrospher 100 RP-18 (5 microm) column at ambient temperature for cefuroxime and 35 degrees C for cefadroxil with mobile phases consisting of water-acetonitrile-acetic acid (85:15:0.1 v/v) at a flow rate of 1.5 ml min(-1) for cefuroxime; and 0.02 M potassium dihydrogen phosphate-acetonitrile (95:5 v/v) containing 0.003% (w/v) hexanesulphonic acid sodium salt and adjusted to apparent pH 3 with phosphoric acid at a flow rate of 2 ml min(-1) for cefadroxil. Quantitation was achieved with UV detection at 275 and 260 nm for cefuroxime and cefadroxil, respectively, based on peak area with linear calibration curves at the concentration ranges of 2-10 microg ml(-1) for cefuroxime and 5-20 microg ml(-1) for cefadroxil. The proposed methods were applied to the determination of dissolution rate for tablets and capsules containing each drug. The urinary excretion patterns as the cumulative amounts excreted have been calculated for each drug using the proposed methods.     

Simultaneous determination of terbinafine HCL and triamcinolone acetonide by UV derivative spectrophotometry and spectrodensitometry

A binary mixture of terbinafine hydrochloride and triamcinolone acetonide was determined by three different methods. The first one concerned with determination of both drugs using first derivative (D(1)) spectrophotometric technique at 297 and 274 nm over concentration ranges of 5-30 and 4-24 microg ml(-1), with mean percentage accuracies 99.90+/-0.67 and 100.25+/-0.49, respectively. The second method depends on ratio-spectra 1st derivative (RSD(1)) spectrophotometry at 298 and 248 nm over the same concentration ranges with mean percentage accuracies 100.22+/-0.51 and 99.93+/-0.56, respectively. The spectrodensitometric analysis provides a rapid and precise method for the separation and quantitation of both terbinafine hydrochloride and triamcinolone acetonide. The method depends on the quantitative densitometric evaluation of thin layer chromatogram of terbinafine hydrochloride and triamcinolone acetonide at 283 and 238 nm over concentration ranges of 5-25 and 2.5-22.5 microg spot(-1), with mean percentage accuracies 100.66+/-0.51 and 100.27+/-0.73, respectively. The suggested procedures were checked using laboratory prepared mixtures and were successfully applied for the analysis of their pharmaceutical preparations. The three methods retained their accuracy and precision when applying the standard addition technique. The results obtained by applying the proposed methods were statistically analysed and compared with those obtained by a reported method.     

Determination of theophylline and ephedrine HCL in tablets by ratio-spectra derivative spectrophotometry and LC

Two methods are described for the determination of theophylline (THP) and ephedrine hydrochloride (EPH) in combined pharmaceutical tablet forms. The first method depends on the use of the first derivative of the ratio-spectra obtained by dividing the absorption spectrum of binary mixtures by a standard spectrum of one of the compounds. The first derivative amplitudes at 231.8 and 250.3 nm were selected for the assay of THP and EPH, respectively. Calibration graphs were established for 20-180 microg ml(-1) for THP and 10-50 microg ml(-1) for EPH. The second method is based on high-performance liquid chromatography on a reversed-phase column using a mobile phase of methanol-water (40+60,v/v) (pH 3) with detection at 217 nm. Linearity was obtained in the concentration range of 5-150 microg ml(-1) for THP and 15-75 microg ml(-1) for EPH. The detection limits for THP and EPH were 0.73 and 0.92 microg ml(-1) by ratio-spectra derivative spectrophotometry and 0.59 and 0.86 microg ml(-1) by HPLC, respectively. The proposed methods were successfully applied to the determination of these drugs in laboratory-prepared mixtures and in tablets. The relative standard deviations were found to be less than 1.5%, indicating reasonable repeatibility of both methods.     

Stability-indicating spectrophotometric and spectrofluorimetric methods for determination of alfuzosin hydrochloride in the presence of its degradation products

Validated stability-indicating spectrophotometric and spectrofluorimetric assays (SIAMs) were developed for the determination of alfuzosin hydrochloride (ALF) in the presence of its oxidative, acid, and alkaline degradation products. Three spectrophotometric methods were suggested for the determination of ALF in the presence of its oxidative degradation product; these included the use of zero order (0D), first order (1D), and third order (3D) spectra. The absorbance was measured at 330.8 nm for (0D) method, while the amplitude of first derivative (1D) method and that of third derivative (3D) method were measured at 354.0 and 241.2 nm, respectively. The linearity ranges were 1.0-40.0 microg/ml for (0D) and (1D) methods, and 1.0-10.0 microg/ml for (3D) method. Two spectrofluorimetric methods were developed, one for determination of ALF in the presence of its oxidative degradation product and the other for its determination in the presence of its acid or alkaline degradation products. The first method was based on measuring the native fluorescence of ALF in deionized water using lamda(excitation) 325.0 nm and lamda(emission) 390.0 nm. The linearity range was 50.0-750.0 ng/ml. This method was also used to determine ALF in human plasma with the aid of a suggested solid phase extraction method. The second method was used for determination of ALF via its acid degradation product. The method was based on the reaction of fluorescamine with the primary aliphatic amine group produced on the degradation product moiety. The reaction product was determined spectrofluorimetrically using lamda(excitation) 380.0 nm and lamda(emission) 465.0 nm. The linearity range was 100.0-900.0 ng/ml. All methods were validated according to the International Conference on Harmonization (ICH) guidelines, and applied to bulk powder and pharmaceutical formulations.     

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.     

Determination of zolpidem hemitartrate by quantitative HPTLC and LC

Two methods are described for the determination of zolpidem hemitartrate in presence of its degradation product. The first method was a TLC-UV densitometric one in which the mobile phase methanol: water (20:80) was used for developing the TLC plates. The R(f) of zolpidem hemitartrate was found to be 0.29+/-0.01 and that of its degradation product was 0.59+/-0.01. Linearity range was 0.5-4 microg/spot with mean recovery percentage (99.98+/-0.988)%. The second method was an HPLC method. HPLC was performed on a Bondapack C(18) column. The mobile phase was composed of a mixture of acetonitrile-0.01 M KH(2)PO(4) (40:60). The pH was adjusted to 3.5+/-0.1. Flow rate was 1.2 ml/min. Calibration graphs were linear in the range of 0.5-5 microg/ml with UV detection at 245 nm. Both methods have been successfully applied to pharmaceutical formulations. The results obtained were statistically compared with those obtained by applying the reported methods.     

Application of first derivative UV-spectrophotometry and ratio derivative spectrophotometry for the simultaneous determination of candesartan cilexetil and hydrochlorothiazide

Two-component mixtures of candesartan cilexetil (CAN) and hydrochlorothiazide (HYD) were assayed by first derivative and ratio derivative spectrophotometry. The first method depends on zero-crossing and peak to base measurement. The first derivative amplitudes at 270.1 and 255.5 nm were selected for the assay of (CAN) and (HYD), respectively. The second method depends on first derivative of the ratio spectra by division of the absorption spectrum of the binary mixture by a normalized spectrum of one of the components and then calculating the first derivative of the ratio spectrum. The first derivative of the ratio amplitudes at 236, 250, 232, 267 and 280 nm were selected for the determination of (CAN) and (HYD), respectively. Calibration curves were established for 6.0-38.0 microg x ml(-1) for (CAN) and 4.0-28.0 microg x ml(-1) for (HYD) in binary mixtures. Good linearity, precision and selectivity were found, and the two methods were successfully applied to the pharmaceutical dosage form containing the above-mentioned drug combination without any interference by the excipients.     

Stability indicating methods for the determination of aceclofenac

Five new selective, precise and accurate methods for the determination of aceclofenac in the presence of its degradation product; diclofenac are described. Method A utilizes third derivative spectrophotometry at 242 nm. Method B is RSD(1) spectrophotometric method based on the simultaneous use of the first derivative of ratio spectra and measurement at 245 nm. Method C is a pH-induced difference (deltaA) spectrophotometry using UV measurement at 273 nm. Method D is a spectrodensitometric one, which depends on the quantitative densitometric evaluation of thin layer chromatogram of aceclofenac at 275 nm. Method E is RP-HPLC that depends on using methanol: water (60:40 v/v) as mobile phase at a flow rate of 1 ml/min and UV detection at 275 nm. Regression analysis of a beer's plot showed good correlation in the concentration ranges 5-40, 10-40, 15-50, 50-200, 1-50 microg/ml for methods A, B, C, D and E, respectively. These methods are suitable as stability indicating methods for the determination of aceclofenac in presence of its main degradation product, diclofenac. The proposed methods were applied for the analysis of the drug in its pharmaceutical formulation and the results obtained were compared with those obtained with the official B.P. method.     

Determination of cisapride in pharmaceutical preparations using derivative spectrophotometry

Derivative spectrophotometric and high performance liquid chromatographic methods (HPLC) were described for the determination of cisapride in pharmaceutical preparations. Spectrophotometrically, cisapride was determined by measuring the 1D-values at 264, 300 nm and 2D-values at 276, 290 and 276-290 nm. Beer's Law was obeyed in the range 2-12 microg ml(-1). The HPLC method depends upon using micropack-Si-10 column at ambient temperature with a mobile phase consisting of methanol-concentrated ammonia (99.25:0.75) at a flow rate of 1 ml min(-1). Quantitation was achieved by UV detection at 272 nm using quinine as internal standard. Calibration curve was linear over the concentration range 2-10 microg ml(-1). Both derivative spectrophotometry and HPLC methods showed good linearity, precision and reproducibility. No interference was found from tablet or suspension matrices at the selected derivative wavelengths and chromatographic conditions. The proposed methods were successfully applied to the assay of commercial tablets and suspension. The procedures were rapid, simple and suitable for quality control applications.     

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.     

Determination of some antihistaminic drugs by atomic absorption spectrometry and colorimetric methods

Atomic absorption spectrometry (AAS) and colourimetric methods have been developed for the determination of pizotifen (I), ketotifen (II) and loratadine (III). The first method depends on the reaction of the three drugs (I); (II) and (III) with cobalt thiocyanate reagent at pH 2 to give ternary complexes. These complexes are readily extracted with organic solvent and estimated by indirect atomic absorption method via the determination of the cobalt content in the formed complex after extraction in 0.1 M hydrochloric acid. It was found that the three drugs can be determined in the concentration ranges from 10 to 74, 12 to 95 and 10 to 93 microg ml(-1) with mean percentage recovery of 99.71+/-0.87, 99.70+/-0.79 and 99.62+/-0.75%, respectively. The second method is based on the formation of orange red ion pairs as a result of the reaction between (I); (II) and (III) and molybdenum thiocyanate with maximum absorption at 469.5 nm in dichloromethane. Appropriate conditions were established for the colour reaction. Under the proposed conditions linearity was obeyed in the concentration ranges 3.5-25, 5-37.5 and 2.5-22.5 microg ml(-1) with mean percentage recovery of 99.60+/-0.41, 100.11+/-0.43 and 99.31+/-0.47% for (I): (II) and (III), respectively. The third method depends on the formation of radical ion using 2,3-dichloro-5,6-dicyano-p-benzoquinone (DDQ). The colour formed was measured at 588 nm for the three drugs (I); (II) and (III), respectively. The method is valid in concentration range 10-80 microg ml(-1) with mean percentage recovery 99.75+/-0.44, 99.94+/-0.72 and 99.17+/-0.36% for (I); (II) and (III), respectively. The proposed methods were applied to the analysis of pharmaceutical preparations. The results obtained were statistically analysed and compared with those obtained by applying the official and reference methods.     

Determination of Lansoprazole in pharmaceutical dosage forms by two different spectroscopic methods

Two different ultraviolet (UV) spectroscopic methods were developed for determination of Lansoprazole in pharmaceutical dosage forms. The solutions of the standard and the sample were prepared in 0.1 M NaOH and phosphate buffer pH 6.6. Both UV spectrophotometric and derivative spectroscopic techniques were applied. Second-order derivative spectra were generated between 200 and 400 nm at N = 9, deltalambda = 31.5. The linear range for the UV spectrophotometric method was 3.0-25.0 microg ml(-1) and that for the derivative spectroscopic method was 0.5-25.0 microg ml(-1). The developed methods were applied to three different pharmaceutical preparations. The percentage recovery was 100.2%.     

Determination of aceclofenac in bulk and pharmaceutical formulations.

Three sensitive and reproducible methods for quantitative determination of aceclofenac (AC) in pure form and in pharmaceutical formulation are presented. The first method is based on the reaction between the drug via its secondary aromatic amino group and p-dimethylaminocinnamaldehyde (PDAC) in acidified methanol to give a stable coloured complex after heating at 75 degrees C for 20 min. Absorption measurements were carried out at 665.5 nm. Beer's law is obeyed over concentration range 20-100 microg ml(-1) with mean recovery 100.33 +/- 0.84. The other two methods are high performance liquid chromatography (HPLC) and densitometric methods by which the drug was determined in the presence of its degradation products over concentration range of 20-70 microg ml(-1) and 1-10 microg per spot and mean recoveries are 99.59 +/- 0.90 and 99.45 +/- 1.09, respectively.     

Simultaneous determination of moexipril hydrochloride and hydrochlorothiazide in tablets by derivative spectrophotometric and high-performance liquid chromatographic methods

Two new simple and selective assay methods have been presented for the binary mixtures of moexipril hydrochloride (MOEX) and hydrochlorothiazide (HCTZ) in pharmaceutical formulations. The first method depends on second-derivative ultraviolet spectrophotometry with zero-crossing measurements at 215 and 234 nm for MOEX and HCTZ, respectively. The assay was linear over the concentration ranges 1.0-11.0 microg ml(-1) for MOEX and 0.5-9.0 microg ml(-1) for HCTZ. The determination limits for MOEX and HCTZ were found to be 1.0 and 0.5 microg ml(-1), respectively; while the detection limits were 0.2 microg ml(-1) for MOEX and 0.1 microg ml(-1) for HCTZ. The second method was based on isocratic reversed-phase liquid chromatography by using a mobile phase acetonitrile-20 mM phosphate buffer (pH 4.0) (50:50, v/v). Lisinopril was used as an internal standard (IS) and the substances were detected at 212 nm. The linearity range for both drugs was 0.5-12.0 microg ml(-1). The determination and detection limits were found to be 0.100 and 0.010 microg ml(-1) for MOEX and 0.025 and 0.005 microg ml(-1) for HCTZ, respectively. The proposed methods were successfully applied to the determination of these drugs in synthetic mixtures and commercially available tablets with a high percentage recovery, good accuracy and precision.     

Stability indicating methods for the determination of clozapine

Five new selective, precise and accurate methods are described for the determination of clozapine in the presence of its main degradation product. Method A utilizes the second and third derivative spectrophotometry at 315 and 305 nm, respectively. Method B is RSD(1) spectrophotometric method based on the simultaneous use of the first derivative of ratio spectra and measurement at 295 nm. Method C is a pH-induced difference (delta A) spectrophotometry using UV measurement at 325 nm. Method D is a densitometric one, after separation on silica gel plate using methanol: water as mobile phase, and the spot was scanned at 295 nm. Method E is RP-HPLC using acetonitrile: water (40:60 v/v) as mobile phase at a flow rate of 1 ml/min and UV detection was at 295 nm. Regression analysis showed good correlation in the concentration ranges 3-10, 4-10, 10-25 micro g/ml, 200-1000 ng/spot, 5-100 micro g/ml with percentage recoveries of 99.4+/-0.28, 99.8+/-0.20, 100.05+/-0.11, 99.41+/-0.34, 100.11+/-0.07 and 100.07+/-0.05% for methods A, B, C, D and E, respectively. These methods are suitable as stability indicating methods for the determination of clozapine in the presence of its main degradation product either in bulk powder or in pharmaceutical formulations.     

Spectrophotometric resolution of metronidazole and miconazole nitrate in ovules using ratio spectra derivative spectrophotometry and RP-LC

Metronidazole and miconazole nitrate in ovules was determined by ratio spectra derivative spectrophotometry and by high-performance liquid chromatography (HPLC). The first method depends on ratio spectra first derivative spectrophotometry, by utilizing the linear relationship between substances concentration and ratio spectra first derivative peak amplitude. The ratio first derivative amplitudes at 242.6 [(1)DD(242.6)], 274.2 [(1)DD(274.2))] 261.8 [(1)DD(261.8))] 273.5 [(1)DD(273.5))]and 281.5 [(1)DD(281.5)] nm were selected for the assay of metronidazole and miconazole nitrate, respectively. The second method is based on high-performance liquid chromatography on a reversed-phase column using a mobile phase of methanol-water-phosphoric acid (30:70:0.20 v/v) (pH 2.8) with programmable detection at 220.0 nm. The minimum concentration detectable by HPLC was 0.9 microg ml(-1) for metronidazole and 0.3 microg ml(-1) for miconazole nitrate and by ratio derivative spectrophotometry 4.0 microg ml(-1) for metronidazole and 0.5 microg ml(-1) for miconazole nitrate. The proposed procedures were successfully applied to the simultaneous determination of metronidazole and miconazole nitrate in ovules with a high percentage of recovery, good accuracy and precision.