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PHARMACEUTICAL SCIENCES
http://doi.org/10.5281/zenodo.1064353
Please cite this article in press as Tukaram B Sawant and Dhananjay V Mane., To Develop Hplc Method for the
Assay of Memantine Hydrochloride Tablets Using Refractive Index (RI) Detector, Indo Am. J. P. Sci, 2017; 4(11).
EXPERIMENTAL:
Materials and reagents:
Memantine hydrochloride pure drug (100.0% w/w),
tablets with label claim 30 mg of Memantine
hydrochloride per tablet were provided by Wockhardt
Ltd Aurangabad, Maharashtra India.
Fig. 1: Structure of Memantine Hydrochloride.
MEM is used as option in a new class of AD HPLC grade methanol was purchased from Merck
treatment and showed very good results in terms of Chemicals, Mumbai, India. Ultrapure water was
efficacy and safety for patients with moderate to generated from Milli-Q water purifier. Diethylamine
severe symptoms. It has been recently approved by was purchased from Spectrochem, Mumbai, India.
European Union and Australia for the treatment of Orthophosphoric acid and hydrochloric acid were
purchased from Merck Ltd, Mumbai, India. Sodium
moderately severe to severe AD [5].
chloride was purchased from Merck Chemicals,
The drugs which are used in treatment of AD need to
Mumbai, India.
be given in appropriate dosage regimen. Hence it is
very important to estimate the amount of drug present
in the dosage forms. For determination of amount of Instruments and Methods:
drug (assay) in different formulations like tablets, Instrumentation:
capsules etc. a simple, rapid and economical Waters High performance liquid chromatography
analytical method is required. (HPLC) system with Refractive Index Detector.
The mobile phase (blank), placebo solution, The placebo was accurately weighed about 120mg
individual impurities (10g/ml) and standard drug for all level. For 80% (80g/ml) level of recovery
solution (100g/ml) were injected in sequence for studies, 40.0 mg of MEM standard was spiked along
evaluation of specificity of proposed method. The with 120mg placebo and dissolved in 50ml of mobile
chromatograms were monitored for any peak eluted phase. Further diluted 5.0 ml of this solution to 50 ml
at the retention time of drug. with mobile phase. The resulting solution was filtered
through 0.45 syringe filter. In same manner for
Precision 100% and 120% recovery studies 50mg (100g/ml)
Precision express the measure of how close the and 60mg (120.0g/ml) of MEM was spiked with
analytical results are to each other from a set of 120mg of placebo respectively and prepared the
measurements under controlled analytical conditions. solutions. All the above solutions were prepared in
Precision proves random errors of the measurement. triplicate and were analyzed using proposed
Precision is a measure of the degree of repeatability chromatographic condition. The recovery at each
(Intra-day), intermediate precision and level was calculated by using the theoretical value
reproducibility (inter-day) of the analytical method from exact weight taken for spiking. The % recovery
under normal operating circumstances. was calculated with respect to amount added. The
Precision is usually measured as the relative standard acceptance criteria for % recovery are in the range of
deviation (RSD) of analytical results acquired from 98 - 102%.
independently prepared quality control standards.
Method precision was evaluated by six sample Linearity
preparations of same homogeneous test sample of The linearity of an analytical method is its ability to
MEM (30mg/tablet) and calculated % assay for each elicit test results that are directly, or by means of
sample preparation. The % RSD for set of six well-defined mathematical transformations,
preparations was calculated. proportional to the concentration of analytes in the
samples within a given range.
The linearity plot was constructed for MEM in the to the system considering as an initial at 0 hr as
range of 50 to 150g/ml. The primary stock solution baseline. A solution stability of MEM was carried out
of 1000g/ml of MEM was prepared in mobile phase. for sample solution (100.0g/ml) in a tightly capped
From the primary stock solution, appropriate volumetric flask at ambient temperature for 72 hr.
dilutions were made to get concentration of 50.0, The results were calculated against freshly injected
70.0, 100.0, 120.0 and 150.0. The calibration curve standard.
was plotted as concentration of the respective drug
solutions versus the peak area at each level. The System Suitability
results were statistically evaluated and correlation The rationale of the system suitability assessment is
coefficient determination (r2), slope and y-intercept to make sure that the complete testing system
values were calculated. (including instrument, reagents, columns, analysts) is
appropriate for the intended application.
Robustness System suitability tests (SST) are vital part of liquid
Robustness is the capacity of a method to remain chromatographic methods. They are used to verify
unaffected by small deliberate variations in method the reproducibility of the chromatographic
parameters. One consequence of evaluation of parameters and system is satisfactory for the analysis
robustness is that a series of system suitability to be done. SST is support on the concept that the
parameters is established to ensure that the analytical equipment, electronics, analytical operations and
procedure is maintained whenever used. In the samples to be analyzed comprise an integral system
present study the working concentration 100.0g/ml that can be evaluated as such.
of MEM was used for the determination of the The system suitability test was performed in
robustness of the method. The following parameters accordance with USP [18].
were considered for the robustness of the proposed
chromatographic method. Application of chromatographic method for
Effect of pH in the mobile phase (0.2) determination of assay of MEM tablets:
Effect of organic modifier in mobile phase Assay test of MEM tablets was performed using
composition ( 2%) developed method.
Effect of flow rate (0.1)
Column oven and detector temperature (2C) RESULTS:
Validation of chromatographic method
Solution stability of sample and standard in Specificity:
mobile phase The overlay chromatogram (Fig. 2) of diluent,
The solution stability of MEM sample was performed placebo, known impurities and standard solution
to understand stability which will be helpful to were revealed that there is no interference at the
understand sample handling in proposed retention of MEM. The developed chromatographic
chromatographic method. The sample and standard method was found to be highly specific for
solution after preparation were injected immediately determination of assay for MEM tablets and capsules.
Fig. 2: Chromatogram of Memantine HCl along with diluents, placebo and known impurities.
Solution stability of sample and standard and accurate over the range. The validation of the
The MEM found to be stable up to 72h in dissolution method is done accordingly to ICH Q2 (R1) and USP
medium at ambient temperature. The difference 38. All the parameters are meeting the acceptance
between the initial results and after 72h was found to criteria. Since method is simple and time saving
be 0.5%. hence it can be used conveniently by laboratories to
determine the assay of different formulations of
Application of developed method: MEM.
The proposed validated method was used for
determination of assay in MEM tablets and the ACKNOWLEDGEMENTS:
results were found within the specification (98.5%). The authors thank The Principal, Shri. Chhatrapati
Shivaji College, Omerga, Dr. Babasaheb Ambedkar
DISCUSSION: Marathwada University and Wockhardt Research
MEM lacs chromophore, due to this limitation, MEM Centre (Global Technical Services), Aurangabad
cannot be readily assayed by HPLC-UV techniques (431210), MS, India for providing support and
and hence refractive index detector was selected for facilities during research work respectively.
detection. During development of this method,
method optimization was carried out by using REFERENCES:
different HPLC columns, by changing concentration 1.Reisberg, B., Doody, R., Stffler, A., Schmitt, F.,
of organic modifier (methanol) and also different pH Ferris, S., Mbius, H.J. Memantine in moderate-to-
to achieve the separation between impurity and severe Alzheimer's disease. New England Journal of
MEM. The main peak of MEM in the proposed Medicine 2003; 348(14): 1333-1341.
method was found at 7.0 min. (Fig. 3). The sensitivity 2.Alzheimers Disease International: World
of standard and sample at concentration 100 g/mL Alzheimer Report 2015.
of MEM was found good. https://www.alz.co.uk/research/WorldAlzheimerRepo
The method was found economical, simple and rt2015.pdf (accessed May 4, 2016).
robust. The system suitability was found to be with 3.Fan, L.Y., Chiu, M.J., Combotherapy and current
acceptance criteria. The proposed method is more concepts as well as future strategies for the treatment
accurate and time saving than the current published of Alzheimers disease. Neuropsychiatric Disease
methods. It can be used by laboratories to determine and Treatment 2014; 10: 439-451.
the assay of memantine hydrochloride tablets of 4.Ballard, C., Gauthier, S., Corbett, A., Brayne, C.,
different strengths. The refractive index detector is a Aarsland, D., Jones, E. Alzheimer's disease. The
universal detector and easily available. Analyst need Lancet 2011; 377(9770): 1019-1031.
not to do derivatization of drug which is complex and 5.Tariot, P.N., Farlow, M.R., Grossberg, G.T.,
time consuming. Graham, S.M., McDonald, S., Gergel, I. Memantine
treatment in patients with moderate to severe
CONCLUSION: Alzheimer disease already receiving donepezil: a
The validation of the method proven that the method randomized controlled trial. The Journal of the
is linear in the range of 50150 g/ml, to be precise
American Medical Association 2004; 291(3): 317- 12.Hassan, M.G., Emara, K.M., Mohamed, H.A.,
324. Abdel -Wadood, H.M., Ikeda, R., Wada, M., Kuroda,
6.Yanamadala, G. and Praveen Srikumar, P. A pre- N. and Nakashima, K. Determination of memantine
column derivatization technique for the development in rat plasma by HPLC-fluorescence method and its
and validation of a stability indicating HPLC-UV application to study of the pharmacokinetic
method for the determination of Memantine interaction between memantine and methazolamide.
Hydrochloride in bulk and formulations by using (2- Biomedical Chromatography 2012; 26(2): 214-219.
napthoxy) acetyl chloride. Der Pharma Chemica 13.Toker, S.E., Sarl, O., etin, S.M. and nal, A.
2014; 6(4):169-180. A new HPLC method with fluorescence detection for
7.Jalalizadeh, H., Raei, M., Tafti, R.F., Farsam, H., the determination of memantine in human plasma.
Kebriaeezadeh, A., Souri, E. A Stability-indicating Journal of Separation Science 2011; 34(19): 2645-
HPLC method for the determination of memantine 2649.
Hydrochloride in dosage forms through derivatization 14.Xie, M.F., Zhou, W., Tong, X.Y., Chen, Y.L., Cai,
with 1-fluoro-2, 4-dinitrobenzene. Scientia Y., Li, Y. and Duan, G.L. High performance liquid
Pharmaceutica 2014; 82(2): 265-279. chromatographic determination of memantine
8.Narola, B., Singh, A.S., Santhakumar, P.R. and hydrochloride in rat plasma using sensitive
Chandrashekhar, T.G. A validated stability-indicating fluorometric derivatization. Journal of Separation
reverse phase HPLC assay method for the Science 2011; 34(3): 241-246.
determination of memantine hydrochloride drug 15.International Conference on Harmonization
substance with UV-detection using precolumn Harmonized Tripartite Guideline. (2005) Validation
derivatization technique. Analytical chemistry of analytical procedures: text and methodology Q2
insights 2010; 5: 37-45. (R1).http://www.ich.org/products/guidelines/quality/
9.Rystov, L., Chadwick, R., Krock, K. and Wang, T. quality-single/article/validation-of-analytical-
Simultaneous determination of Maillard reaction procedures-text-and-methodology.html (accessed on
impurities in memantine tablets using HPLC with January 10, 2016).
charged aerosol detector. Journal of Pharmaceutical 16.United States of Pharmacopoeia 38 NF 33, (2015),
and Biomedical Analysis 2011; 56(5): 887-894. United States Pharmacopeial Convention General
10.Puente, B., Garcia, M.A., Hernandez, E., chapter <1225>, Validation of Compendial
Bregante, M.A., Perez, S., Pablo, L. and Prieto, E. Procedures, pp.1445-1450.
Determination of memantine in plasma and vitreous 17.United States of Pharmacopoeia 38 NF 33, (2015),
humour by HPLC with precolumn derivatization and United States Pharmacopeial Convention General
fluorescence detection. Journal of Chromatographic chapter <1092>, The Dissolution Procedure:
Science 2011; 49(10): 745-752. Development and Validation, pp. 1090-1097.
11.Zarghi, A., Shafaati, A., Foroutan, S.M., 18.United States of Pharmacopoeia 38 NF 33, (2015),
Khoddam, A. and Madadian, B. Sensitive and rapid United States Pharmacopeial Convention General
HPLC method for determination of memantine in chapter <621>, Chromatography, pp. 424-434.
human plasma using OPA derivatization and 19.Waters 2414 Refractive Index Detector Operators
fluorescence detection: application to Guide. (2008) .
pharmacokinetic studies. Scientia Pharmaceutic http://www.waters.com/webassets/cms/support/docs/
2010; 78(4): 847-856. 71500241402rb.pdf (accessed December 11, 2015).