National Academy of Agricultural Sciences (NAAS)
|
PRINT ISSN : 2319-7692
Online ISSN : 2319-7706 Issues : 12 per year Publisher : Excellent Publishers Email : editorijcmas@gmail.com submit@ijcmas.com Editor-in-chief: Dr.M.Prakash Index Copernicus ICV 2018: 95.39 NAAS RATING 2020: 5.38 |
Antimicrobial resistance and consumer demand for clean-label food products are driving urgent interest in naturally derived food bio-preservatives. Antimicrobial peptides from food-grade plant sources represent particularly promising candidates, combining potent broad-spectrum antimicrobial activity with biodegradability, food-safety regulatory compatibility, and renewable sourcing. This study aimed to evaluate the protein extraction efficiency, AMP characterisation, and antimicrobial activity of eleven food-grade plant materials — Cinnamon, Nutmeg, Clove, Black pepper, Sorghum, Kidney beans, Peas, Pearl millet, Fig, and Guava — relevant to Indian food systems, against food spoilage bacteria isolated from spoiled Indian food samples. Proximate analysis (Kjeldahl protein; Soxhlet fat; Anthrone carbohydrate) was performed on all eleven samples. Protein extraction was conducted by cold acetone precipitation from Tris-HCl buffer homogenates. Total extractable protein was quantified by Bradford assay with BSA standard curve validation (R² = 0.993). Protein purity was assessed by NanoDrop UV-Visible spectral scanning (A260/A280 ratio). Protein and peptide profiling was performed by RP-HPLC (Waters XBridge C18; 0.1% TFA/acetonitrile gradient; 214 nm and 280 nm detection). Antimicrobial activity of aqueous extracts was evaluated by agar well diffusion (zone of inhibition) and broth microdilution (minimum inhibitory concentration) against six food spoilage bacteria: E. coli ATCC 25922, K. pneumoniae NCIM 2719, S. Typhi (clinical), S. aureus NCIM 2079, Enterococcus sp., and Bacillus sp. Priority AMP fractions from Black pepper (SP4) and Guava (G1) were purified by DEAE-Sepharose ion exchange chromatography and Sephadex G-50 gel filtration, and subjected to LC-MS/MS peptide sequencing with database annotation against APD3. Bradford extractable protein ranged from 19.7 µg/mL (Pearl millet; S8) to 86.4 µg/mL (Black pepper; S4). NanoDrop A260/A280 ratios (0.60–0.78) confirmed acceptable to moderate protein purity in all eleven samples. RP-HPLC identified AMP- candidate fractions (RT 20–35 min; 32–55% acetonitrile) in all samples; Guava (S11) showed the highest AMP-candidate zone peak area (846.9 mAU·min), followed by Clove (S3; 656.2 mAU·min) and Black pepper (S4; 634.8 mAU·min). Antimicrobial activity testing showed Peas producing the broadest-spectrum activity (mean ZOI 15.6 mm across six organisms), followed by Nutmeg (13.5 mm) and Sorghum (13.4 mm). LC-MS/MS identified nsLTP1-class Pepin-1 from Black pepper (78% sequence identity, APD3) and nsLTP2 from Guava (83% identity) as the primary active peptide components in purified fractions, with Pearson correlation r = +0.79 (p < 0.01) between Bradford protein concentration and RP-HPLC AMP-zone peak area across the sample panel. Black pepper, Peas, and Guava emerged as first-priority AMP bio- preservative candidates based on convergent evidence from five independent analytical methods. The integrated platform of acetone precipitation, Bradford and NanoDrop characterisation, RP-HPLC profiling, SDS-PAGE, and LC-MS/MS identification provides a validated and reproducible analytical pipeline for AMP discovery from food- grade plant sources. The nsLTP2 identification from Guava represents the first LC- MS/MS-confirmed AMP from Psidium guajava reported in the food science literature. These findings support the translational development of plant-derived AMP bio- preservatives as alternatives to synthetic chemical food preservatives for Indian and global food industry applications.
Arima, H., and Danno, G. (2002). Isolation of antimicrobial compounds from guava (Psidium guajava L.) and their structural elucidation. Bioscience, Biotechnology, and Biochemistry, 66(8), 1727–1730.
Atanda, O. O., Ikenebomeh, M. J., and Asagbra, A. (2007). Inhibitory activity of Clove extract on food spoilage fungi. African Journal of Microbiology Research, 1(5), 40–44.
Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72(1–2), 248–254.
Broekaert, W. F., Terras, F. R., Cammue, B. P., and Osborn, R. W. (1995). Plant defensins: novel antimicrobial peptides as components of the host defense system. Plant Physiology, 108(4), 1353–1358.
Cholewa-Waclaw, J. et al., (2020). Lipid transfer protein-based edible coatings for fresh-cut fruit preservation. Food Chemistry, 318, 126481.
Cotter, P. D., Hill, C., and Ross, R. P. (2013). Bacteriocins: developing innate immunity for food. Nature Reviews Microbiology, 11(2), 95–105.
Dang, L., Van Damme, E. J. M. (2014). Antimicrobial peptides from Cinnamomum species: characterization and function. Journal of Antimicrobial Chemotherapy, 69, 3223–3232.
Delves-Broughton, J., Blackburn, P., Evans, R. J., and Hugenholtz, J. (1996). Applications of the bacteriocin, nisin. Antonie van Leeuwenhoek, 69(2), 193–202.
Dykes, L., and Rooney, L. W. (2006). Sorghum and millet phenols and antioxidants. Journal of Cereal Science, 44(3), 236–251.
FAO (2019). The State of Food and Agriculture: Moving Forward on Food Loss and Waste Reduction. FAO, Rome.
Glasel, J. A. (1995). Validity of nucleic acid purities monitored by 260/280 absorbance ratios. BioTechniques, 18(1), 62–63.
Gyawali, R., and Ibrahim, S. A. (2014). Natural products as antimicrobial agents. Food Control, 46, 412–429.
Hancock, R. E. W., and Sahl, H. G. (2006). Antimicrobial and host-defense peptides as new anti- infective therapeutic strategies. Nature Biotechnology, 24(12), 1551–1557.
Lam, S. K., and Ng, T. B. (2010). A protein with antiproliferative, antifungal and antibacterial activities from mung bean seeds. Journal of Biosciences, 35(4), 569–574.
Naik, J. R., Kumar, A., and Prakash, V. (2017). Bioactive protein hydrolysates from Pearl millet grain. Food Chemistry, 223, 86–93.
O'Neill, J. (2016). Tackling Drug-Resistant Infections Globally: Final Report and Recommendations. Review on Antimicrobial Resistance. HM Government / Wellcome Trust, London.
Padovan, L., Scocchi, M., and Tossi, A. (2010). Structural aspects of plant antimicrobial peptides. Current Protein and Peptide Science, 11(3), 210–219.
Schagger, H. (2006). Tricine-SDS-PAGE. Nature Protocols, 1(1), 16–22.
Solomon, A., Golubowicz, S., Yablowicz, Z., Grossman, S., Bergman, M., Gottlieb, H. E., et al., (2006). Antioxidant activities and anthocyanin content of fresh fruits of common Fig (Ficus carica L.). Journal of Agricultural and Food Chemistry, 54(20), 7717–7723.
Sunila, E. S., and Kuttan, G. (2004). Immunomodulatory and antitumor activities of Piper longum Linn. Journal of Ethnopharmacology, 90(2–3), 339–346.
Tam, J. P., Wang, S., Wong, K. H., and Tan, W. L. (2015). Antimicrobial peptides from plants. Pharmaceuticals, 8(4), 711–757.
Thevissen, K., et al., (2012). Defensin-based food preservation: antifungal activity against Aspergillus parasiticus in bread. Food Microbiology, 30(1), 43–49.
Wang, G., Li, X., and Wang, Z. (2016). APD3: The antimicrobial peptide database as a tool for research and education. Nucleic Acids Research, 44(D1), D1087–D1093.
WHO (2015). WHO Estimates of the Global Burden of Foodborne Diseases. WHO, Geneva.
Ye, X. Y., and Ng, T. B. (2002). A new antifungal protein from kidney beans (Phaseolus vulgaris). Biochemical and Biophysical Research Communications, 291(3), 717–722.
Yemm, E. W., and Willis, A. J. (1954). The estimation of carbohydrates in plant extracts by anthrone. Biochemical Journal, 57(3), 508–514.
Zasloff, M. (1987). Magainins, a class of antimicrobial peptides from Xenopus skin: isolation, characterization of two active forms, and partial cDNA sequence of a precursor. Proceedings of the National Academy of Sciences USA, 84(15), 5449–5453.
Zasloff, M. (2002). Antimicrobial peptides of multicellular organisms. Nature, 415(6870), 389–395.
Citations
|
|
|
