IDR - IIT Kharagpur

Development of Methods for Detoxification and Rapid Detection of Aflatoxin B1 in Red Chilli Powder

Development of Methods for Detoxification and Rapid Detection of Aflatoxin B1 in Red Chilli Powder


The partially purified garlic peroxidase (POD) was used to detoxify AFB1 infected RCP. The enzymatically detoxified RCP was further exposed to long wave (365 nm) UV rays to enhance the detoxification efficiency. The POD with specific activity (246.6 U/mg protein) was extracted from garlic bulbs by 80% ammonium salt precipitation and gel filtration chromatography. The POD was immobilized in 3% κ-carageenan with immobilization efficiency of 95%. The free enzyme retained 37% while entrapped enzyme retained 67% of its initial activity at the end of four months when stored at 37 °C in 0.01 phosphate buffer. The AFB1- POD reaction in phosphate buffer was studied for varied substrate and enzyme concentration, pH, time, temperature and inhibitor sensitivity. The optimum enzymatic reaction occurred in buffered liquid substrate (50 mmol phosphate buffer) at 40 ºC, pH 7, incubation time 50 min using 6U POD/nmol AFB1 resulting in 71.4% detoxification. In RCP the optimum reaction occurred at 12U of POD/nmol AFB1 after 24 h incubation at 37 ºC resulting in 66% detoxification. The km and Vmax for the AFB1-POD reaction was calculated as 0.50 nmol AFB1/min and 100 nmol in liquid system and in RCP as 100 nmol and 0.024 nmol AFB1/min respectively. The Ea and Q10 for AFB1-POD reaction were 55.8 kJ/mol and 3.1 respectively. The 12 U POD treatment for 24 h followed by 30 min UV exposure (365 nm) reduced the AFB1 from 19.8 μg/kg to 5.8 μg/kg while the 13 U POD treatment followed by similar UV treatment reduced the AFB1 from 9.9 μg/kg to 1.6 μg/kg in AFB1 contaminated red chilli powder with minimal quality changes. The detoxification process at these optimum parameters was safe in terms of overcoming toxic potentials of the AFB1 molecule. The FT-NIR spectroscopy in diffuse reflectance mode with sphere macrosample integrating measurement channel was successfully used to detect the AFB1 in RCP. The SLS preprocessing method in wavenumber range of 6900 - 4998 and 4902 - 3999 cm-1 detected the AFB1 in RCP (15-500 μg/kg) with best accuracy. The RMSECV and R2 calculated for this calibration model were 0.65 and 96.7% respectively. Key words: Aflatoxin, Detoxification, FT-NIR, Peroxidase, Toxicity

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