الفهرس 
Valaciclovir HydrochlorideOnly 14 pages are availabe for public view
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Abstract
This thesis is concerned with the development of different simple, sensitive and
selective methods for determination of anti-viral drugs; namely valaciclovir HCl,
oseltamivir phosphate and zanamivir either alone or in presence of their acid and/or
alkaline degradates. Furthermore, the work aimed to determine aciclovir, the active
metabolite of valaciclovir HCl, in human plasma which could be used for bioequivalence
studies.
The establishment of a method that quantify the pure drug in presence of its different
degradates is of great pharmaceutical value, because any degradation occurring at normal
storage conditions can be determined.
This thesis includes five parts:
Part one: Introduction
This part is divided into two sections
Section A: General Introduction
This section comprises a brief data about anti-viral drugs, their classes, mode of action
and drug stability.
Section B: Literature Review
In this section, the structures, properties and review of the published analytical
procedures developed for their analysis, are stated.
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Part two: chromatographic Methods
This part is divided into nine sections
Section A: Introduction to Ultra High Pressure Liquid chromatography.
Section B: Stability Indicating Rapid Resolution Liquid chromatography (RRLC)
Method for Determination of Valaciclovir HCl:
This section describes a new stability indicating RRLC method for determination of
valaciclovir HCl in presence of its alkaline- and acid- degradates . This method based on
the separation of intact drug using ZORBAX SB phenyl 50.0 mm L x 4.6 mm id x 1.8
μm particle size column as stationary phase , Binary mixture of 0.01M of n- tetra butyl
ammonium hydrogen sulfate and methanol in the ratio of (95:5, v/v) as mobile phase,
with flow rate of 0.3 ml min -1and UV detection at 254 nm .Linear correlation was
obtained in the range of( 0.40-40.0 μg ml-1) without interference from the alkaline and
acid degradates.
The proposed method was applied for the analysis of valaciclovir HCl in pure form,
laboratory prepared mixtures containing different percentages of its degradates and in
pharmaceutical preparation. The validity was assessed by applying the standard addition
technique. The obtained results were statistically compared with those of the reference
method. Satisfactory results were obtained upon applying the suggested method in
pharmaceutical formulation which encourages the use of the proposed method in quality
control laboratories.
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Section C: Bio-analytical RRLC Method for Determination of Aciclovir, the Active
Metabolite of Valaciclovir, in Spiked Human Plasma:
This section is an application of proposed RRLC method describes in section B for
determination of aciclovir, the active metabolite of valaciclovir HCl, in human plasma.
Section D: Stability Indicating Reversed Phase Rapid Resolution Liquid
Chromatography (RP-RRLC) Method for Determination of Oseltamivir Phosphate:
This section describes a new stability indicating RP- RRLC method for determination
of oseltamivir phosphate in presence of its alkaline-degradate. This method based on the
separation of the intact drug using Zorbax XDB C18 50.0 mm L x 4.6 mm id x 1.8μm
particle size column as stationary phase ,water (pH 3.0): methanol (50:50,v/v) as mobile
phase with flow rate of 1.0 ml min-1 and UV detection at 215 nm. Linear correlations
obtained in the range of (4.0-240.0 μg ml-1) without interference of the alkaline
degradate.
The proposed method was successively applied for the analysis of oseltamivir
phosphate in pure form, laboratory prepared mixtures containing different percentages of
its alkaline-degradate and in pharmaceutical preparation. The validity was assessed by
applying the standard addition technique. The results obtained were statistically
compared with those obtained with reference method and no significant difference was
observed regarding accuracy and precision. The results indicate that the proposed method
could be used for routine analysis of oseltamivir phosphate in presence of its alkalinedegradate.
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Section E: Stability Indicating Reversed Phase High Performance Liquid
Chromatography (RP-HPLC) Method for Determination of Valaciclovir HCl :
This section describes a new stability indicating RP- HPLC method for determination
of valaciclovir HCl in presence of its alkaline- and acid- degradates, based on the
separation of the intact drug using Hypersil BDS C18 250.0 mm L x 4.6 mm id x 5.0 μm
particle size column as stationary phase, Phosphate buffer (pH 4.6): Methanol (90:10,v/v)
as mobile phase with flow rate of 1.0 ml min-1 and UV detection at 252 nm. Linear
correlations obtained in the range of (0.5-20.0 μg ml-1) without interference of the
alkaline- and acid -degradates .
The suggested method was applied for the analysis of valaciclovir HCl in pure form,
laboratory prepared mixtures containing different ratios of its degradates and in
pharmaceutical preparation. The validity was assessed by applying the standard addition
technique. Statistical analysis of the results showed no significant difference when
compared with those obtained with reference method regarding accuracy and precision.
Section F: Stability Indicating Reversed Phase High Performance Liquid
Chromatography (RP-HPLC) Method for Determination of Oseltamivir Phosphate:
This section describes a new stability indicating RP- HPLC method for determination
of oseltamivir phosphate in presence of its alkaline-degradate. This method based on the
separation of the intact drug using Symmetry C8 150.0 mm L x 4.6 mm id x 5.0μm
particle size column as stationary phase ,phosphate buffer (pH 3.0): methanol (60:40,v/v)
as mobile phase with flow rate of 1.5 ml min-1 and UV detection at 215 nm. Linear
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correlations obtained in the range of (1.0-100.0 μg ml-1) without interference of the
alkaline -degradate.
The proposed method was applied for the analysis of oseltamivir phosphate in pure
form, laboratory prepared mixtures containing different ratios of its alkaline-degradate
and also in pharmaceutical preparation. The validity was assessed by applying the
standard addition technique. The obtained results were statistically compared with those
of the reference method. Satisfactory results were obtained upon applying the suggested
method in pharmaceutical formulation so it can be used in quality control laboratories.
Section G: Stability Indicating Reversed Phase High Performance Liquid
Chromatography (RP-HPLC) Method for Determination of Zanamivir:
This section describes a new stability indicating RP- HPLC method for determination
of zanamivir in presence of its acid-degradate. This method based on the separation of
the intact drug using Xterra C18 250.0 mm L x 4.6 mm id x 5.0μm particle size column
as stationary phase ,0.01M octane sulfonic acid sodium salt( pH 6.0) as mobile phase
with flow rate of 1.0 ml min-1 and UV detection at 235 nm. Linear correlations obtained
in the range of (2.0-100.0 μg ml-1) without interference of the acid -degradate.
Laboratory prepared mixtures containing zanamivir and different ratios of the aciddegradate
were analyzed by the proposed method and satisfactory results were obtained.
The proposed method was also used for determination of zanamivir in pure form and in
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pharmaceutical preparation and good results were obtained. The validity of the suggested
method was assessed by applying standard addition technique.
The obtained results were statistically compared with those obtained by the reference
method. The calculated t and F values were less than the theoretical ones, indicating that
both methods are equally accurate and precise.
Section H: Stability Indicating Spectrodensitometric Thin Layer chromatography
(TLC) Method for Determination of Valaciclovir HCl:
This section describes a new stability indicating densitometric TLC for determination
of valaciclovir HCl in the presence of its alkaline-degradate . This method based on
spotting methanolic solution of valaciclovir HCl on a silica gel F254 plate, using methanol:
ethyl acetate: ammonia in ratio of (50:50:10, by volume) as mobile phase. Scanning of
the chromatogram was carried out at 254 nm by reflection mode. Linear correlation was
obtained in the range of (0.4-4.0μg/spot).
Application of the proposed method to the analysis of valaciclovir HCl in pure form,
laboratory prepared mixtures containing different ratios of its alkaline- degradate and also
in pharmaceutical preparation showed that neither the excipients usually formulated in
the market preparation nor the alkaline –degradate interfere with the determination . The
validity was assessed by applying the standard addition technique. The reliability of the
proposed method was determined through statistical comparison with the reference
method. The calculated t and F values were found to be less than the theoretical ones,
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indicating that there is no significant difference between the two methods in respect to
accuracy and precision which encourages the use of the proposed method in quality
control laboratories.
Section I: Stability Indicating Spectrodensitometric Thin Layer chromatography
(TLC) Method for Determination of Oseltamivir Phosphate:
This section describes a new stability indicating densitometric TLC for determination
of oseltamivir phosphate in the presence of its alkaline-degradate . This method based on
spotting methanolic solution of oseltamivir on a silica gel F254 plate, using methanol:
ethyl acetate: diethyl ether: ammonia (80:60:20:5, by volume) as mobile phase. Scanning
of the chromatogram was carried out at 215 nm by reflection mode. Linear correlation
was obtained in the range of (5.0-50.0μg/spot).
The proposed method was applied for the analysis of oseltamivir phosphate in pure
form, laboratory prepared mixtures containing different ratios of its alkaline- degradate
and also in pharmaceutical preparation. The validity was assessed by applying the
standard addition technique. Statistical comparison of the results obtained by the
proposed densitometric method and the reference method was done and no significant
difference was observed regarding accuracy and precision.
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Part three: Spectrophotometric methods
This part is divided into two sections
Section A: Stability Indicating First Derivative of Ratio Spectra Spectrophotometric
Method for determination of Oseltamivir Phosphate in Presence of its Alkaline-
Degradate:
This section describes stability indicating method used for the determination of
oseltamivir phosphate in the presence of its alkaline-degradates. This method based on
the use of first derivative ratio spectrophotometric method (1DD). Oseltamivir could be
determined in presence of its alkaline-degradates by dividing the spectra of the intact
drug by the spectrum of its alkaline-degradate (10.0 μg ml-1) as divisor .The peak
amplitudes of the first derivative ratio spectra were measured at 223nm .A linear
relationship was obtained in concentration range of (5.0-45.0μg ml-1)for determination of
oseltamivir phosphate without interference from the alkaline-degradate.
The proposed derivative ratio method was used for the determination of oseltamivir
phosphate in laboratory prepared mixtures and in the available pharmaceutical dosage
form where good results were obtained. The validity of the suggested procedure was
proved by applying the standard addition technique. Statistical comparison of the results
obtained by applying the proposed method and the reference method permits one to
conclude with 95% of confidence, that there is no significant difference between the
proposed method and the reference method.
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The proposed analytical procedure is simple, rapid, specific and could be applied for
quality and stability monitoring of oseltamivir phosphate.
Section B: Stability Indicating First Derivative of Ratio Spectra Spectrophotometric
Method for determination of Zanamivir in Presence of its acidic- Degradate:
This section describes stability indicating method used for the determination of
zanamivir in the presence of its acid-degradate. This based on the use of first derivative
ratio spectrophotometric method (1DD). Zanamivir could be determined in presence of its
acid-degradates by dividing the spectra of the intact drug by the spectrum of its aciddegradate
(12.0 μg ml-1) as divisor .The peak amplitudes of the first derivative ratio
spectra were measured at 222 nm .A linear relationship was obtained in concentration
range of (4.0-36.0μg ml-1) for determination of zanamivir without interference from the
acid-degradate.
The proposed method was applied for the analysis of zanamivir in pure form,
laboratory prepared mixtures containing different ratios of its acid-degradate and also in
pharmaceutical preparation. The validity was assessed by applying the standard addition
technique. The obtained results were statistically compared with those of the reference
method. Satisfactory results were obtained upon applying the suggested method in
pharmaceutical formulation which encourages the use of the proposed method in quality
control laboratories.
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Part four: General Discussion
This part shows a short summary of the developed methods. A comparison between the
studied methods, choosing the most sensitive method for each drug, is also stated.
Part five: References
This thesis refers to 115 references, contains 64tables and 24 figures and ends with an
Arabic summary.