Browsing by Author "Saleh S.S."
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Item Advanced approaches for the treatment and amplification of weak spectral signals produced by critical concentrations in white multicomponent systems(Elsevier B.V., 2020) Lotfy H.M.; Saleh S.S.; El-Maraghy C.M.; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini Street; Cairo; 11562; Egypt; Pharmaceutical Chemistry Department; Faculty of Pharmaceutical Sciences & Pharmaceutical Industries; Future University in Egypt; Cairo; 12311; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); 11787 6th October; EgyptAn analytical investigation was carried out to study the treatment and amplification of the spectral signals produced by critical concentrations with high accuracy and precision using two advanced approaches. The factorized-spectrum approach was applied through two novel methods which were: absorptivity centering technique via both: factorized zero order absorption spectrum (ACT-FSD0 ?A) and factorized ratio spectrum (ACT-FSR?P). The proposed methods were found to be linear in the ranges of (15�100 ?g/mL) and (3�40 ?g/mL) for ASP and MTO, respectively. Those methods were compared to the methods following the geometrical standard addition approach: ratio H-point standard addition method (RHPSAM) and geometrical induced amplitude modulation (GIAM). The approaches were applied for the determination of the minor component metoclopramide in its mixture with the major component aspirin in the challengeable ratio of (1,90) respectively in a white multicomponent system. The results obtained from the proposed approaches were statistically compared with each other. The methods were validated according to ICH guidelines where the results were found to be within the acceptable limits. The methods were found to be accurate and reliable for the determination of metoclopramide critical concentration besides aspirin concentration. The results of single factor ANOVA analysis indicated that there is no significant difference among the developed methods. These methods provided simple resolution of this binary combination from synthetic mixtures and pharmaceutical preparation and can be conveniently adopted for routine quality control analysis. � 2019 Elsevier B.V.Item Column performance study of different variants of liquid chromatographic technique: An application on pharmaceutical ternary mixtures containing tetryzoline(Oxford University Press, 2015) Salem H.; Hassan N.Y.; Lotfy H.M.; Saleh S.S.; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); 6th October City; 11787; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini Street; Cairo; 11562; EgyptHigh-performance liquid chromatography (HPLC), ultra-performance liquid chromatography (UPLC) and rapid resolution liquid chromatographic (RRLC) methods have been developed and validated for the separation and quantitation of both or either of two ternary mixtures present in ophthalmic solutions. The first mixture contains chloramphenicol, dexamethasone sodium phosphate and tetryzoline HCl (TZH); while the second one contains ofloxacin, prednisolone acetate and TZH. Both preparations contain benzalkonium chloride as a preservative. The columns used were a HPLC column (C18 5 ?m particle size), a RRLC column (C18 2.6 ?m particle size) and a UPLC column (C18 1.7 ?m particle size). A comparative study was conducted to illustrate the effect of the change in column particle size and dimensions on the other chromatographic conditions, backpressure and the separation of both ternary mixtures. The methods were validated as per ICH guidelines where accuracy, repeatability, interday precision and robustness were found to be within the acceptable limits. The RRLC column provided shorter run time and better resolution than HPLC, while the UPLC column gave the shortest run time for all columns. The RRLC column resulted in minimum backpressure, so it could be used with any HPLC instrument, which makes the method more practical and economic. The results obtained from the proposed methods were statistically compared with official ones where no significant difference was observed. � The Author 2014. Published by Oxford University Press.Item A comparative study of different aspects of manipulating ratio spectra applied for ternary mixtures: Derivative spectrophotometry versus wavelet transform(Elsevier, 2015) Salem H.; Lotfy H.M.; Hassan N.Y.; El-Zeiny M.B.; Saleh S.S.; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); 11787 6th October City; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini Street; 11562 Cairo; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; Modern University for Technology and Information (MTI); 12582 Al Hadaba Al Wosta; Cairo; EgyptThis work represents a comparative study of different aspects of manipulating ratio spectra, which are: double divisor ratio spectra derivative (DR-DD), area under curve of derivative ratio (DR-AUC) and its novel approach, namely area under the curve correction method (AUCCM) applied for overlapped spectra; successive derivative of ratio spectra (SDR) and continuous wavelet transform (CWT) methods. The proposed methods represent different aspects of manipulating ratio spectra of the ternary mixture of Ofloxacin (OFX), Prednisolone acetate (PA) and Tetryzoline HCl (TZH) combined in eye drops in the presence of benzalkonium chloride as a preservative. The proposed methods were checked using laboratory-prepared mixtures and were successfully applied for the analysis of pharmaceutical formulation containing the cited drugs. The proposed methods were validated according to the ICH guidelines. A comparative study was conducted between those methods regarding simplicity, limitation and sensitivity. The obtained results were statistically compared with those obtained from the reported HPLC method, showing no significant difference with respect to accuracy and precision.� 2014 Elsevier B.V. All rights reserved.Item Comparative study of new spectrophotometric methods; An application on pharmaceutical binary mixture of ciprofloxacin hydrochloride and hydrocortisone(Sociedad Chilena de Quimica, 2013) Lotfy H.M.; Hassan N.; Elgizawy S.M.; Saleh S.S.; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini Street; 11562 Cairo; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; Assuit University; 71515 Assuit; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); EgyptSimple, specific, accurate and precise spectrophotometric methods were developed and validated for simultaneous estimation of Ciprofloxacin hydrochloride (CIP) and Hydrocortisone (HYD) in their pure form and pharmaceutical dosage form. Two new spectrophotometric methods were applied: extended ratio subtraction (EXRSM) and ratio difference (RDSM) methods. The results were compared to three well-established methods: Mean centering of ratio spectra method (MCR), Isoabsorpative point spectrophotometric method (IsoM) and Absorbance ratio method (ARM). The linearity range for the spectrophotometric methods was found to be (2-14 ?g/mL) and (1-14 ?g/mL) for CIP and HYD respectively. The selectivity of the developed methods was investigated by analyzing laboratory prepared mixtures of the two drugs and their combined dosage form. The results obtained from the proposed methods were statistically compared using one-way analysis of variance (ANOVA) where no significant difference was observed between the new (EXRSM and RDSM) and the well-established methods which prove their validity for the analysis of this binary mixture. The methods were validated as per ICH guidelines regarding accuracy, precision, repeatability and robustness; which were found to be within the acceptable limits. The new methods were simple, sensitive and don't need a special program, so they could be easily applied as alternative methods to LC methods in quality control laboratories lacking the required facilities for those techniques.Item A comparative study of novel spectrophotometric methods based on isosbestic points; Application on a pharmaceutical ternary mixture(2014) Lotfy H.M.; Saleh S.S.; Hassan N.Y.; Salem H.; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini Street; 11562 Cairo; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); 11787 6th October City; EgyptThis work represents the application of the isosbestic points present in different absorption spectra. Three novel spectrophotometric methods were developed, the first method is the absorption subtraction method (AS) utilizing the isosbestic point in zero-order absorption spectra; the second method is the amplitude modulation method (AM) utilizing the isosbestic point in ratio spectra; and third method is the amplitude summation method (A-Sum) utilizing the isosbestic point in derivative spectra. The three methods were applied for the analysis of the ternary mixture of chloramphenicol (CHL), dexamethasone sodium phosphate (DXM) and tetryzoline hydrochloride (TZH) in eye drops in the presence of benzalkonium chloride as a preservative. The components at the isosbestic point were determined using the corresponding unified regression equation at this point with no need for a complementary method. The obtained results were statistically compared to each other and to that of the developed PLS model. The specificity of the developed methods was investigated by analyzing laboratory prepared mixtures and the combined dosage form. The methods were validated as per ICH guidelines where accuracy, repeatability, inter-day precision and robustness were found to be within the acceptable limits. The results obtained from the proposed methods were statistically compared with official ones where no significant difference was observed. � 2014 Elsevier B.V. All rights reserved.Item A comparative study of progressive versus successive spectrophotometric resolution techniques applied for pharmaceutical ternary mixtures(Elsevier, 2014) Saleh S.S.; Lotfy H.M.; Hassan N.Y.; Salem H.; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); 11787 6th October City; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini Street; 11562 Cairo; EgyptThis work represents a comparative study of a novel progressive spectrophotometric resolution technique namely, amplitude center method (ACM), versus the well-established successive spectrophotometric resolution techniques namely; successive derivative subtraction (SDS); successive derivative of ratio spectra (SDR) and mean centering of ratio spectra (MCR). All the proposed spectrophotometric techniques consist of several consecutive steps utilizing ratio and/or derivative spectra. The novel amplitude center method (ACM) can be used for the determination of ternary mixtures using single divisor where the concentrations of the components are determined through progressive manipulation performed on the same ratio spectrum. Those methods were applied for the analysis of the ternary mixture of chloramphenicol (CHL), dexamethasone sodium phosphate (DXM) and tetryzoline hydrochloride (TZH) in eye drops in the presence of benzalkonium chloride as a preservative. The proposed methods were checked using laboratory-prepared mixtures and were successfully applied for the analysis of pharmaceutical formulation containing the cited drugs. The proposed methods were validated according to the ICH guidelines. A comparative study was conducted between those methods regarding simplicity, limitation and sensitivity. The obtained results were statistically compared with those obtained from the official BP methods, showing no significant difference with respect to accuracy and precision. � 2014 Elsevier B.V. All rights reserved.Item A comparative study of validated spectrophotometric and TLC- spectrodensitometric methods for the determination of sodium cromoglicate and fluorometholone in ophthalmic solution(2013) Saleh S.S.; Lotfy H.M.; Hassan N.Y.; Elgizawy S.M.; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); 11787 6th October City; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini Street; 11562 Cairo; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; Assuit University; 71515 Assuit; EgyptThe determination of sodium cromoglicate (SCG) and fluorometholone (FLU) in ophthalmic solution was developed by simple, sensitive and precise methods. Three spectrophotometric methods were applied: absorptivity factor (a-Factor method), absorption factor (AFM) and mean centering of ratio spectra (MCR). The linearity ranges of SCG were found to be (2.5-35?g/mL) for (a-Factor method) and (MCR); while for (AFM), it was found to be (7.5-50?g/mL). The linearity ranges of FLU were found to be (4-16?g/mL) for (a-Factor method) and (AFM); while for (MCR), it was found to be (2-16?g/mL). The mean percentage recoveries/RSD for SCG were found to be 100.31/0.90, 100.23/0.57 and 100.43/1.21; while for FLU, they were found to be 100.11/0.56, 99.97/0.35 and 99.94/0.88 using (a-Factor method), (AFM) and (MCR), respectively. A TLC-spectrodensitometric method was developed by separation of SCG and FLU on silica gel 60 F254 using chloroform:methanol:toluene:triethylamine in the ratio of (5:2:4:1v/v/v/v) as developing system, followed by spectrodensitometric measurement of the bands at 241nm. The linearity ranges and the mean percentage recoveries/RSD were found to be (0.4-4.4?g/band), 100.24/1.44 and (0.2-1.6?g/band), 99.95/1.50 for SCG and FLU, respectively. A comparative study was conducted between the proposed methods to discuss the advantage of each method. The suggested methods were validated in compliance with the ICH guidelines and were successfully applied for the determination of SCG and FLU in their laboratory prepared mixtures and commercial ophthalmic solution in the presence of benzalkonium chloride as a preservative. These methods could be an alternative to different HPLC techniques in quality control laboratories lacking the required facilities for those expensive techniques. 2012.Item Computation of geometric representation of novel spectrophotometric methods used for the analysis of minor components in pharmaceutical preparations(Elsevier, 2015) Lotfy H.M.; Saleh S.S.; Hassan N.Y.; Salem H.; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini; Cairo; 11562; Egypt; Pharmaceutical Chemistry Department; Faculty of Pharmaceutical Sciences and Pharmaceutical Industries; Future University in Egypt (FUE); Cairo; 12311; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); 6th October; 11787; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; Deraya University; Minia; EgyptNovel spectrophotometric methods were applied for the determination of the minor component tetryzoline HCl (TZH) in its ternary mixture with ofloxacin (OFX) and prednisolone acetate (PA) in the ratio of (1:5:7.5), and in its binary mixture with sodium cromoglicate (SCG) in the ratio of (1:80). The novel spectrophotometric methods determined the minor component (TZH) successfully in the two selected mixtures by computing the geometrical relationship of either standard addition or subtraction. The novel spectrophotometric methods are: geometrical amplitude modulation (GAM), geometrical induced amplitude modulation (GIAM), ratio H-point standard addition method (RHPSAM) and compensated area under the curve (CAUC). The proposed methods were successfully applied for the determination of the minor component TZH below its concentration range. The methods were validated as per ICH guidelines where accuracy, repeatability, inter-day precision and robustness were found to be within the acceptable limits. The results obtained from the proposed methods were statistically compared with official ones where no significant difference was observed. No difference was observed between the obtained results when compared to the reported HPLC method, which proved that the developed methods could be alternative to HPLC techniques in quality control laboratories. � 2015 Elsevier B.V. All rights reserved.Item Design, optimization, and validation of thin-layer chromatography-densitometry and chemometry-assisted spectrophotometry: A comparative study applied on quaternary mixture(Akademiai Kiado Rt., 2015) Salem H.; Hassan N.Y.; Lotfy H.M.; Saleh S.S.; Department of Analytical Chemistry; Faculty of Pharmacy; Deraya University; Minia; Egypt; Department of Analytical Chemistry; Faculty of Pharmacy; Cairo University; Kasr-El Aini Street; Cairo; 11562; Egypt; Department of Pharmaceutical Chemistry; Faculty of Pharmaceutical Sciences and Pharmaceutical Industries; Future University in Egypt; Cairo; 12311; Egypt; Department of Analytical Chemistry; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); October City; 11787 6th; EgyptThis work presents a comparative study on the development and validation of two analytical techniques applied for the simultaneous determination of hydrocortisone acetate (HCA), fusidic acid (FSA), methyl paraben (MPB), and propyl paraben (PPB) formulated as a topical cream. The first technique was thin-layer chromatography (TLC)-densitometric method, which was developed by separating the four components on silica gel 60 F254 using methylene chloride-methanol-benzene in the ratio of 10:2:5, v/v, as the developing system, followed by densitometric measurement of the bands at 240 nm. The second technique was the chemometric method using two models: principle component regression model (PCR) and partial least squares (PLS). The suggested techniques were validated in compliance with the International Conference on Harmonization (ICH) guidelines and were successfully applied for the determination of the quaternary mixtures as prepared synthetically in laboratory and in the commercial topical pharmaceutical formulation. Akadmiai Kiad, Budapest.Item Development and validation of impurity-profiling UPLC method for the determination of sodium cromoglicate and tetryzoline hydrochloride: Application on rabbit aqueous humor(Elsevier, 2015) Lotfy H.M.; Saleh S.S.; Hassan N.Y.; Salem H.; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini; Cairo; 11562; Egypt; Pharmaceutical Chemistry department; Faculty of Pharmaceutical sciences and Pharmaceutical industries; Future University in Egypt (FUE); Cairo; 12311; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); 6th October; 11787; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; Deraya University; Minia; EgyptSodium cromoglicate (SCG), antihistaminic agent, and tetryzoline hydrochloride (TZH), a sympathomimetic agent, are formulated together as an ophthalmic preparation. An ultra-performance liquid chromatographic method with UV detection (UPLC-UV) was developed and validated for the quantitative determination of SCG and TZH in rabbit aqueous humor. Due to the instability of both SCG and TZH under alkaline conditions, the UPLC method was applied for their determination in the presence of their possible degradation impurities. The separation was performed using C18 column (1.7?m particle size) and isocratic elution system with methanol: 1% o-phosphoric acid (65: 35, v/v). The optimum flow rate was 0.5ml/min and the detection was done at 230nm. The suggested method was validated in compliance with the ICH guidelines and was successfully applied for determination of sodium cromoglicate (SCG) and tetryzoline HCl (TZH) as prepared synthetically in laboratory mixtures, and in the presence of their alkali-induced degradation impurities. The suggested method was effectively applied the determination of spiked rabbit aqueous humor samples as well as commercial pharmaceutical formulation. � 2015 Elsevier B.V.Item Development of membrane electrodes for the specific determination of tetryzoline hydrochloride in presence of its degradation product in pharmaceutical formulations and biological fluids(Center of Excellence in Electrochemistry, Univ. of Tehran, 2015) Hassan N.Y.; Lotfy H.M.; Saleh S.S.; Salem H.; Analytical Chemistry Department; Cairo University; Kasr�El Aini Street; Cairo; 11562; Egypt; Analytical Chemistry Department; Future University (FUE); New Cairo; 11835; Egypt; Analytical Chemistry Department; October University for Modern Sciences and Arts (MSA); 6th October City; 11787; Egypt; Analytical Chemistry Department; Deraya University; Minia; EgyptMembrane selective electrodes were used to determine tetryzoline hydrochloride (TZH) in pure form, pharmaceutical preparations and in biological fluids. The membrane selective electrodes include construction of water insoluble ion-association complexes. The TZH ion exchangers were formed using tetraphenyl borate (TZH-TPB), phosphomolybdic acid (TZH-PMA) and phosphotungstic acid (TZH-PTA), in a plasticized PVC (polyvinyl chloride) matrix, using dibutyl phthalate (DBP) or dioctylphthalate (DOP) as a plasticizer. The performance characteristics of the developed sensors were evaluated according to IUPAC recommendations. The developed sensors showed good responses but the best electrochemical characteristics and selectivity coefficients were achieved with TZH-TPB sensor using DBP as a plasticizer, where the linear responses of TZH was found within the concentration ranges of 10?6 to 10?2 mol/L and Nernstian slope was calculated to be of 56.8 mV/ decade at 25 �C, over the pH range of 5�9. The suggested method was used to determine TZH in synthetic mixtures, pharmaceutical formulations and in presence of its alkali degradation product. The proposed sensors displayed useful analytical characteristics for the determination of TZH in biological fluids such as rabbit aqueous humor and human plasma. The later application can be used to detect oral TZH poisoning in children. The obtained results were statistically compared with the official method, showing no significant difference with respect to accuracy and precision. � 2015 The Authors.Item Investigating advanced approaches based on iso-absorptivity coefficient in unresolved spectral signals of binary mixtures(Hindawi Limited, 2019) Lotfy H.M.; Saleh S.S.; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini Street; Cairo; 11562; Egypt; Pharmaceutical Chemistry Department; Faculty of Pharmaceutical Sciences and Pharmaceutical Industries; Future University in Egypt; Cairo; 12311; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); 11787 6th of October; EgyptSeveral spectrophotometric approaches utilize different functions of the iso-absorptivity coefficient in zero-order absorption signals and its manipulated spectra. This work introduced an investigation concerning the efficiency power of recent methods based on iso-absorptivity coefficient in different spectral signals. These methods were as follows: absorptivity centering method (a-Centering), absorbance subtraction method (AS), amplitude modulation method (AM,) and amplitude summation method (A-Sum). These methods were applied to determine the binary mixture of ofloxacin (OFX) and dexamethasone (DXM). Linearity of the proposed methods was investigated in the range of 1.0-10.0 ?g/ml for both drugs. The proposed methods were validated as per ICH guidelines and were successfully applied for the simultaneous determination of OFX and DXM in their pharmaceutical preparation without interference from additives. Statistical analysis of the results obtained by the proposed spectrophotometric methods compared with a reported method revealed no significant difference between the proposed and reported methods, confirming accuracy and precision at 95% confidence limit. � 2019 Hayam M. Lotfy and Sarah S. Saleh.Item Novel two wavelength spectrophotometric methods for simultaneous determination of binary mixtures with severely overlapping spectra(Molecules, 2015) Lotfy H.M.; Saleh S.S.; Hassan N.Y.; Salem H.; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini Street; Cairo; 11562; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); October City; 11787 6[th]; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; Deraya University; Minia; EgyptThis work presents the application of different spectrophotometric techniques based on two wavelengths for the determination of severely overlapped spectral components in a binary mixture without prior separation. Four novel spectrophotometric methods were developed namely: induced dual wavelength method (IDW), dual wavelength resolution technique (DWRT), advanced amplitude modulation method (AAM) and induced amplitude modulation method (IAM). The results of the novel methods were compared to that of three well-established methods which were: dual wavelength method (DW), Vierordt's method (VD) and bivariate method (BV). The developed methods were applied for the analysis of the binary mixture of hydrocortisone acetate (HCA) and fusidic acid (FSA) formulated as topical cream accompanied by the determination of methyl paraben and propyl paraben present as preservatives. The specificity of the novel methods was investigated by analyzing laboratory prepared mixtures and the combined dosage form. The methods were validated as per ICH guidelines where accuracy, repeatability, inter-day precision and robustness were found to be within the acceptable limits. The results obtained from the proposed methods were statistically compared with official ones where no significant difference was observed. No difference was observed between the obtained results when compared to the reported HPLC method, which proved that the developed methods could be alternative to HPLC techniques in quality control laboratories. � 2014 Elsevier B.V.Item Recent development in ultraviolet spectrophotometry through the last decade (2006-2016): A review(Innovare Academics Sciences Pvt. Ltd, 2016) Lotfy H.M.; Saleh S.S.; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini; Cairo; 11562; Egypt; Pharmaceutical Chemistry Department; Faculty of Pharmaceutical Sciences and Pharmaceutical Industries; Future University in Egypt (FUE)Cairo 12311; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); 6th October; 11787; EgyptSeveral techniques have been proposed for treatment of spectrophotometric data, with the objective of extracting the largest amount of analytical information from spectra composed of unresolved bands. The instrumental development and analytical applications of UV regions absorption spectrophotometry produced in the last 10 y (since 2006) are reviewed. The methods were classified according to the spectra data used, including zero order absorption, derivative and ratio amplitudes. The proposed methods were applied to solve different complex pharmaceutical mixtures and the obtained results were accepted when compared to other reported methods. � 2016 The Authors.Item Testing the purity of spectral profiles: Finger-print resolution of complex matrices and extraction of absorbance signals(Elsevier B.V., 2018) Lotfy H.M.; Saleh S.S.; Analytical Chemistry Department; Faculty of Pharmacy; Cairo University; Kasr-El Aini Street; Cairo; 11562; Egypt; Pharmaceutical Chemistry Department; Faculty of Pharmaceutical Sciences & Pharmaceutical Industries; Future University in Egypt; Cairo; 12311; Egypt; Analytical Chemistry Department; Faculty of Pharmacy; October University for Modern Sciences and Arts (MSA); 6th October; 11787; EgyptThe application of spectrophotometric techniques has shown a tremendous development over the past few years, where it is possible to determine the concentrations of several components in complex matrix. A new feature will be introduced in this work where the application of spectrophotometric techniques will be enhanced to resolution and checking the purity of signals. The finger-print resolution �ratio subtraction method� (RSM) was coupled with the novel complementary method �unified constant subtraction� (UCS); in addition to the methods: extended ratio subtraction method (EXRSM), constant multiplication (CM) or �spectrum subtraction� (SS). These techniques were applied for the determination of the complex matrix of the binary mixture of chloramphenicol and dexamethasone. By applying official spectrophotometric methods, direct determination of the components was allowed with no need for validation procedures. The spectrophotometric techniques were successfully applied to the laboratory prepared mixtures and the combined dosage form where the purity of the extracted signals were tested by calculating the spectral contrast angle (?) and the spectral ratio factor (SRF) where the results were compared to show the capability to recover pure spectral profiles and detect the presence of impurities. The proposed methods proved that spectrophotometric techniques can be used for identification and separation of signals, similar to chromatographic techniques. � 2018 Elsevier B.V.