HPLC Quantification of Thymoquinone Extracted from Nigella sativa L. (Ranunculaceae) Seeds and Antibacterial Activity of Its Extracts against Bacillus Species

Abstract

The medicinal importance of Nigella sativa seeds for treating various ailments is portrayed by its traditional uses. Owing to its immense pharmacological importance, the thymoquinone phytoconstituent of N. sativa can prove beneficial for the South Asian countries including Pakistan, where this seed is commonly produced and healthcare facilities are limited. In this study, the antibacterial activity of various extracts of N. sativa seeds, extracted thymoquinone, and oil samples have been investigated against Bacillus subtilis and Bacillus licheniformis using well and disc diffusion assay. The inhibition zones ranged between 7 and 44 mm against both the bacterial strains by well diffusion assay, while disc diffusion assay provided inhibition zones in the range of 7–23 mm. Commercial and local Kalonji oil samples were included in the study. Oil samples dissolved in methanol showed increased inhibition of bacteria. However, the extracted thymoquinone showed highest antibacterial activity. Medicine formulated using thymoquinone will prove to be an herbal alternate against the resistant microbiota associated with bacterial infections. Antibacterial activity against some Bacillus species will help signify the effect on normal gut flora when oral therapy is followed. Trying different extraction protocols can help increase extraction efficiency. Study on extraction of thymoquinone in local produce of black seed can be fruitful for conducting the stability studies and can help to gain maximum benefits from the bioactives. The crude extracts from 10 g of these seeds were subjected to preliminary phytochemical investigation. Results showed that although methanol extract had the presence of maximum phytochemicals, hexane extract was the most potent in terms of antibacterial activity. Thymoquinone, a therapeutically important bioactive in N. sativa seed, was extracted employing both solvents. TLC assay and UV spectroscopy were used for its qualitative assessment, while HPLC-UV quantification showed that 250 mg/mL of methanol extract had 368.3 μg/mL thymoquinone, while its successive extraction yielded 32.94 μg/mL thymoquinone.