High performance liquid chromatographic method optimized by Box-Behnken design model to determine caffeine in pharmaceutical preparations and urine samples.

A UV-HPLC method optimized by Box-Behnken design model was developed to determine caffeine in pharmaceutical preparations and urine samples. The chromatographic conditions followed were mobile phase: methanol/water/ citrate buffer (pH 4.6) (40:25:35, v/v/v),AcclaimTMDionex C18 column (ODS 100A˚, 5 µm; 4.6 × 250 mm),flow rate (0.9 mL min-1), column temperature (30 °C) and UV-detection wavelength (204 nm). The chromatographic variables: pH (A), % methanol fraction (B), flow rate(C) and column temperature (D) were optimized at 50 µg mL-1caffeine using BBD model. The chromatogram resulted in the asymmetry factor (1.23), theoretical plate 13,786 and retention time (5.79 min). The proposed HPLC method's greenness point was assessed byAnalytical Eco-scale and found to be 78 (as per guidelines, ranked as excellent). The linearity was ranged from2.0 to 70 µg mL-1 with coefficient of correlation (r = 0.999) and detection limit of 0.19 µg mL-1. The proposedmethod was developed successfully and applied for the assay of active caffeine in pharmaceutical preparations and urine samples. The % recovery obtained by both (proposed and reference) methods ranged from 99.98 to 100.05 % followed the compliance (100 ± 2 %) with Canadian Health Protection regulatory guidelines. The performance of the proposed method was compared with published papers and found to be acceptable and superior. The proposed method was quite effective as the reference method, and hence can be used as an alternative method for the assay of active caffeine in pharmaceutical preparations and urine samples.

Antioxidants profile of Momordica charantia fruit extract analyzed using LC-MS-QTOF-based metabolomics.

Momordica charantia fruit is claimed to have healthy benefit. Despite this potential claim, the phytochemical study of this fruit is still lacking. Thus, this study aimed to evaluate the antioxidants profile of Momordica charantia (Cucurbitaceae) fruit. The antioxidant activity of the ethanolic extracts of various polarities was evaluated and the metabolites that are responsible for its activity were identified using metabolomics approach. Six different mixture of ethanol in water that are 0%, 20%, 40%, 60%, 80%, and 100% (v/v) was extracted using dveseeded fruit sample. Liquid chromatography-mass spectrometry-quadrupole time of flight and multivariate data analysis was used to identify the metabolites that were either antioxidants or pro-oxidants. The 80% ethanol extract exhibited the most antioxidant activity when tested in both 2, 2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP) antioxidant assays. This extract showed the most intense LC-MS signals represented to ascorbic acid, margarolic acid, brevifolincarboxylic acid, quercetin 3-O-glycoside, kuguacin H, cucurbitacin E, 3-malonylmomordicin I, and goyaglycoside G correlating to the anti-oxidant activity. This study reports for the first time the existence of brevifolincarboxylic acid in this fruit, and the antioxidant activity of 3-malonylmomordicin I and goyaglycoside G. In addition, the loading plots revealed the unknown compounds possessing the antioxidant activity which are potential to be isolated in the future study.