Simultaneous Determination of Lamivudine and Zidovudine in Pharmaceutical Formulations by High Performance Thin Layer Chromatography (HPTLC)-Densitometric Method
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Date
2008-01
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Addis Ababa Universty
Abstract
A high performance thin layer chromatographic (HPTLC) method for the simultaneous
determination of lamivudine and zidovudine in binary mixture has been developed.
The method developed was based on HPTLC separation of the two drugs followed by
densitometric measurements of spots at 276 and 271 nm for lamivudine and zidovudine
respectively. Separation was carried out on Merck HPTLC silica gel 60 F254 plates, using
toluene:chloroform:methanol (1:6:3 v:v) as mobile phase.
Validation of the method was performed based on ICH guide line in terms of linearity,
accuracy, precision, limit of detection, limit of quantification and robustness.
Second order polynomial equation was obtained for the regression line in the range of 250–
1400 and 250–1700 ng/spot for lamivudine and zdovudine, respectively. Correlation
coefficient (r) values were 0.9998 for both analytes. The method provides sufficient accuracy
indicated by % recoveries ranged from 98.68 to 100.73 for lamivudine and from 98.56 to
100.61 for zidovudine. For system precision study the low coefficient of variation values (<
2%) for both lamivudine and zidovudine insured reproducible performance of the instrument.
In the method precision study coefficient of variation < 2% was obtained which showed that
the proposed method provides acceptable intra-day and inter-day variation. The detection and
quantification limits for lamivudine and zidovudine were found to be 3.06 and 9.28 and 3.34
and 10.13 ng/spot, respectively. Mobile phase composition, amount of mobile phase, time
from spotting to development and time from development to scanning parameters were
employed while testing for robustness of the method and the standard deviation of peak areas
was calculated for each parameter. The low coefficient of variation value (< 2%) indicated the
robustness of the method. Statistical manipulation did not show any significant effect of one
parameter over the others on the robustness of the method.
No chromatographic interference from the tablet excipients was observed. The described
method can be readily utilized for analysis of both drugs in pharmaceutical formulations.Statistical evaluations were performed to correlate the accuracy and dosage form analysis
results of lamivudine and zidovudine determined by means of the proposed HPTLC and the
reported HPLC method. The calculated t-test and F ratio value did not exceed the theoretical
values at the 95% confidence level, indicating no significant difference between the
performance of the two methods regarding the mean values and standard deviations.
No degradation product was found in the sample analyzed by this method. The time and cost
effectiveness of the developed HPTLC method was also compared with the reported HPLC
method. It was found that the HPTLC method is both time and cost effective for the
determination of lamivudine and zidovudine mixtures in pharmaceutical formulations
compared to the reported HPLC method.
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Keywords
Simultaneous determination of lamivudine and zidovudine