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Gas chromatography-mass spectrometry (GC-MS) method for detection of metformin in biological matrices is developed. Distinct derivatization techniques were employed for sample preparation for GC-MS analysis. N-methyl bis (trifluoroacetamide) [MBTFA] was found to be the best derivatization reagent for the GC-MS analysis of metformin. Total ion current (TIC) mode was selected for the monitoring of ions of trifluoroacetyl(TFA) derivative of metformin with m/z ratio of 303. Distinct parameters like specificity, carryover, stability, precision and accuracy were evaluated for validating the identification method. A novel Thin Layer Chromatography (TLC) method is also developed for the detection of metformin as a screening method. The GC-MS method is found to be linear and illustrated within the range 500ng/ml to 2500ng/ml with the value of R2 (coefficient of determination) at 0.9986. Retention time of derivatized metformin was found to be 4 minutes 2 seconds. Precision (%) for the interday and intraday analysis found to be in the respectable spectrum. Accuracy at each concentration level was within the ±15% of the nominal concentration. Henceforth, the proposed method can be employed for determination of metformin in biologicalmatrices. [ABSTRACT FROM AUTHOR]