الفهرس 
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Abstract
This thesis consists of four parts in addition to the references and a summary in Arabic. Each part includes an introduction, literature review, descriptive experimental work for the studied drugs, results and discussion.
Part I: Quantitative Determination of Diacerein in Presence of Its degradate and Impurity in Different Pharmaceutical Formulations
This part includes seven sections.
Section (A): Introduction and Literature Review
This section includes an introduction about the pharmacological action of Diacerein (DIA), its chemical structure, physical properties and summary of the published methods developed for its analysis in binary mixture with its degradation product and active metabolite, Rhein (RH) and in ternary mixtures with RH and its impurity and related substance, Emodin (EMO).
Section (B): Determination of Diacerein and its Degradate by Q-Analysis (Graphical Absorbance Ratio) Method
In this section, Q-Analysis method was developed for simultaneous determination of Diacerein and Rhein. In this section two wavelengths should be carefully selected, one of the two selected wavelengths is an isoabsorptive point and the other is the wavelength of maximum absorption of one of the two components. Using the absorbance values at 365 nm (λ iso) and 257 nm (λ max for DIA) gave the best results regarding selectivity. The developed method has been applied for determination of the studied drugs in different laboratory prepared mixtures. The results obtained by applying the investigated method for determination of DIA in Diacerein ® and Osteocerein ® capsules were statistically compared to those obtained by applying a reported HPLC one and no significant difference was found regarding both accuracy and precision.
Section (C): Determination of Diacerein and its Degradate by Ratio Subtraction and Extended Ratio Subtraction Spectrophotometric Methods
In this section, ratio subtraction (RS) and the recently developed extended ratio subtraction (ERS) methods have been applied for determination of DIA and RH, respectively in their binary mixture using methanol as a solvent. DIA concentrations were determined by measuring the absorbance at its λ max (257 nm) using RS method, while RH concentrations were determined by measuring the absorbance at its λ max (230 nm) using ERS method.
The developed methods have been applied for determination of the studied components in different laboratory prepared mixtures and the results obtained by applying the investigated methods for determination of DIA in Diacerein ® and Osteocerein ® capsules were statistically compared to those obtained by applying a reported HPLC one and no significant difference was found regarding both accuracy and precision.
Section (D): Determination of Diacerein in presence of its Degradate by Spectrofluorimetric Method.
This section deals with the development of simple, sensitive, and specific stability indicating spectrofluorimetric method for the selective determination of DIA in the presence of its degradate, RH, with successive application to spiked human plasma.
In this method, the native fluorescence of DIA solutions in the range of 0.04-0.3 µg mL-1 at λ em = 404 nm upon excitation at λ ex = 255 nm was measured and used for calculation of DIA without interference from its degradate. In addition, DIA was determined in spiked human plasma after drug extraction with methanol and good recoveries were obtained.
The suggested method was used for determination of DIA in Diacerein ® and Osteocerein ® capsules and the results were statistically compared to those obtained by applying a reported HPLC one and no significant difference was found.
Section (E): Determination of Diacerein, its Degradate and its Impurity by Different Spectrophotometric Methods
In this section, different spectrophotometric methods have been investigated for determination of DIA, RH, and EMO in their ternary mixture. The developed spectrophotometric methods include first (1DD) and second (2DD) derivative of ratio spectra methods.
Diacerein was determined by double divisor method using mixture of 10 µg mL-1 each of RH and EMO as a divisor and measuring the amplitudes at 355 nm. On the other hand, RH and EMO were determined by measuring (2DD) and (1DD) amplitudes, respectively at 229 nm using standard spectrum of 10 µg mL-1 of DIA as a divisor. Specificity of the methods has been assessed by their application to different laboratory prepared mixtures. Also double divisor method has been successfully applied to marketed samples containing DIA.
Section (F): Determination of Diacerein, its Degradate, and its Impurity by Chemometric Methods
Multivariate calibration models, such as PCR and PLS, have been successfully applied as selective stability indicating methods for determination of the ternary mixture of DIA, RH, and EMO.
To validate the predictive ability of the developed models, they were applied to predict the concentrations of DIA, RH, and EMO in an external validation set. Statistical analysis of the results obtained by the developed models was compared with a reported HPLC one and no significant difference was found between them.
Section (G): Determination of Diacerein, its Degradate, and its Impurity by TLC-Densitometric Method
In this section, a simple and accurate TLC-Densitometric method has been suggested for simultaneous determination of DIA, RH, and EMO in their ternary mixtures. The suitable mobile phase has been selected to achieve the best separation.
Quantitative determination of the separated bands was carried out at 230 nm upon using hexane: ethyl acetate: acetic acid (60: 40: 0.8, by volume) as mobile phase in the range of 0.5-10 µg band-1 for DIA and RH and 0.5 - 7 µg band 1 for EMO. The suggested TLC-Densitometric method was successfully applied for analysis of the cited drug in pharmaceutical formulations and the results showed good agreement with the labeled amount.
Part II: Quantitative Determination of Metronidazole and Nystatin in Their Binary Mixtures
This part consists of five sections.
Section (A): Introduction and Literature Review
This section includes an introduction about the pharmacological actions of Metronidazole (MET) and Nystatin (NYS), their chemical structures, physical properties and summary of the published methods developed for their analysis in their single form and in their binary mixture.
Section (B): Determination of Metronidazole and Nystatin by Dual Wavelength Spectrophotometric Method
In this section, dual wavelength method has been developed and optimized for simultaneous analysis of MET and NYS using their zero order spectra. The principle for dual wavelength method is that the absorbance difference at two points on the spectra is directly proportional to the concentration of the component of interest, independent of the interfering component.
Absorption values of NYS were the same at 266.5 and 328 nm, so that these wavelengths were selected for the determination of MET. The same as in 291 and 330 nm, the absorbance values of MET were the same and hence those two wavelengths were selected for estimation of NYS
The developed method was successfully applied for quantitation of the studied drugs in laboratory prepared mixtures and in pharmaceutical formulations, Amrizole N® vaginal suppositories and Nystazole® vaginal tablets, the standard addition technique has been applied to verify their validity.
Section (C): Determination of Metronidazole and Nystatin by Isoabsorptive Point Spectrophotometric Method
In this section, the second derivative (2D) amplitudes at 290 nm were used for determination of NYS concentrations, while isosbestic point (ISO) method was applied for determination of the total mixture concentration (MET and NYS) at the isosbestic point (λ iso = 322 nm). Since the concentration of NYS in the mixture can be determined by using 2D spectrophotometric method, therefore MET concentration can be obtained by subtraction.
The developed methods have been validated according to ICH guidelines and were successfully applied for quantitation of the studied drugs in Amrizole N® vaginal suppositories and Nystazole® vaginal tablets.
Section (D): Determination of Metronidazole and Nystatin by TLC-Densitometric Method
This section is concerned with the development of sensitive, economic and specific TLC-Densitometric method for determination of MET and NYS in their bulk powder and pharmaceutical formulations. The studied components were well separated using methanol: hexane: triethylamine (80: 20: 2, by volume) as a developing system and the separated bands were scanned at 305nm.
Linear relationships were obtained between the mean integrated peak area (× 10-4) and the corresponding concentrations of MET and NYS in the concentration range of 1 - 13 and 0.5 - 10 µg band-1, respectively. The developed TLC-Densitometric method has been applied for determination of the two drugs in their pharmaceutical formulations and the results have been compared with those of the reported chemometric method and no significant difference has been found between them.
Section (E): Determination of Metronidazole and Nystatin by RP-HPLC Method
A precise, specific, and accurate RP-HPLC method was proposed for the determination of MET and NYS. In this method, an isocratic elution of the two components was performed at ambient temperature on ODS column using acetonitrile: 0.05 M phosphate buffer (30: 70, v/v) pH= 3.5 as a mobile phase with a flow rate of 1 mL min-1 and UV-detection at 305 nm.
By applying the suggested RP-HPLC method, MET and NYS could be quantified in the range of 3 - 50 and 15 - 50 µg mL-1, respectively. Statistical comparison of the results obtained by the proposed method and a reported PLS chemometric method was carried out. The values of the calculated t and F are less than the tabulated ones which reveals that there is no significant difference between the two methods with respect to accuracy and precision.
Part III: Determination of Aspirin, Caffeine, and Paracetamol in Their Ternary Mixtures
This part comprises four sections.
Section (A): Introduction and Literature Review
This section includes an introduction about the pharmacological actions of Aspirin (ASP), Caffeine (CAF), and Paracetamol (PAR), their chemical structures, physical properties and a summary of the published methods developed for their analysis in their single formulation and in their ternary mixture.
Section (B): Determination of Aspirin, Caffeine, and Paracetamol by Different Spectrophotometric Methods
The developed spectrophotometric methods are the successive ratio-derivative spectra and indirect spectrophotometric methods. Applying successive ratio-derivative spectra method, ASP and PAR could be selectively determined while, CAF could not be determined by this method, and so, it was determined indirectly using mathematical equation.
Aspirin was determined using the amplitudes of the first derivative of the second ratio spectra at 241.2 nm, while PAR was determined using the amplitudes at 228.2 nm. CAF was indirectly determined using mathematical equations computed at 241.2 and 285.6 nm. The proposed methods were used for quantitation of the studied drugs in Aspicure-combi®, Excedrin®, and Markadel® tablets and results of standard addition technique confirmed that tablet additives did not interfere.
Section (C): Simultaneous Determination of Aspirin, Caffeine, and Paracetamol by Successive Mean Centering of Ratio Spectra Spectrophotometric Method
A recent and simple method was developed for the simultaneous determination of ternary mixture without prior separation steps.
In this method, the mean centered second ratio spectra amplitudes peak to peak at 242 to 243, 203 to 204, and 220 to 221 nm were used for quantitation of ASP, CAF and PAR, respectively. Moreover, the suggested method has been applied for determination of the three drugs in their pharmaceutical preparations. Statistical comparison with the reported spectrophotometric method showed no significant difference.
Section (D): Simultaneous Determination of Aspirin, Caffeine, and Paracetamol by TLC-Densitometric Method
In this section, a simple and accurate TLC-Densitometric method has been suggested for the analysis of the ternary mixtures of ASP, CAF, and PAR in bulk powders and in tablets. The suitable mobile phase has been selected to achieve the best separation.
Quantitative determination of the separated bands of ASP, CAF, and PAR was carried out at 230 nm upon using chloroform: methanol: glacial acetic acid: ammonia solution (95: 5: 0.5: 0.2, by volume) as a developing system in the range of 1 – 10 µg band-1 for ASP and PAR and 0.5 - 7 µg band-1 for CAF. The suggested TLC-Densitometric method was successfully applied for analysis of the cited drugs in pharmaceutical formulations.
Part IV: Appendix
This part includes a brief idea about the instruments, solvents and chemicals used in other parts, in addition to the detailed preparation of the solutions used in each part throughout this work and also method of preparation of DIA degradate, RH.