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KMID : 1011620180340020016
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Korean Journal of Food and Cookey Science
2018 Volume.34 No. 2 p.16 ~ p.142
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Application of Physicochemical Degradation to Increase the Detection Rate of Leaf-Associated Bacteria on Spinach
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Kim Min-Su
Park Eun-Jin
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Abstract
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Purpose: To increase the detection accuracy of resident bacteria on the surfaces of spinach leaves, we combined enzymatic degradation of leaf tissues with physical separation of bacterial cells from the leaf surface of spinach.
Methods: The total numbers of bacterial cells using stomaching only and stomaching with enzyme treatment were compared. We used two enzymes (pectinase and cellulase) to degrade the plant cell wall. Treatment time for stomaching was determined by comparing the numbers of bacterial cells at different treatment times. Then, the treated samples were incubated with each enzyme at 37, 38, 39, and 40°C for 2 hours. The treatment time for enzymatic degradation was determined by monitoring the number of bacterial cells at intervals of 30 minutes.
Results: We observed an average 10-fold higher bacterial cell separation in stomaching than in sonication; however, no significant differences were determined among different treatment times. The optimum conditions for enzymatic degradation were determined to be 165 U/g of pectinase for 1.5 hours, 10 U/g of cellulase for 1 hour, and 5 U/g of cellulase for 1.5 hours at 40°C.
Conclusion: Combining stomaching and enzymatic degradation, the detection rate of bacterial cells was elevated by 49% with pectinase and 42% with cellulase. The method developed in this study can be used for detecting naturally internalized bacteria into spinach leaves, and these results indicate that leaf surface-associated bacteria may be underestimated rather than previously reported.
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KEYWORD
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spinach, enzymatic digestion, bacterial detection, pectinase, cellulase
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