Two hundred grams of alfalfa (Medicago sativa L.) and Italian ryegrass (Lolium multiflorum Lam.) were macerated with 1l of water using a blender and filtered through cheese cloth. The filtrate was kept anaerobically for 2d with or without 2% sucrose (previously fermented juice; PFJ). Alfalfa PFJ with sucrose and Italian ryegrass PFJ without sucrose were used as silage additives.Both of them were added to alfalfa and Italian ryegrass prior to ensile at 2.5ml/kg as to inoculate 106/g of lactic acid bacteria (LAB). Alfalfa used at the present study was not necessary to add PFJ for enhancing the silage fermentation.But the fermentation quality of Italian ryegrass silage was improved by the addition of alfalfa or Italian ryegrass PFJ and Italian ryegrass PFJ was a little more effective than alfalfa PFJ, suggesting that PFJ from other plant is available but that from the same plant to be ensiled is more effective as silage additive.
Eggshells have high bioavailability and can be used as a source of calcium. The main component is CaCO 3 , which, when heated, is converted to CaO. Seashells are also mainly composed of CaCO 3 and were previously found to exhibit antimicrobial activity after being heated. In this study, heated eggshell powder HESP was found to have antimicrobial activity against bacterial vegetative cells, fungi and bacterial spores. Parameters, such as the minimum inhibitory concentration, were determined with kinetic analysis using an indirect conductimetric assay. Moreover, HESP was able to kill the Bacillus subtilis spores. There were no significant differences in the activity between HESP, heated scallop-shell powder and pure CaO. The MIC values for HESP against bacteria and fungi were 0.29-0.43 and 1.3-1.5 mg/mL, respectively. Against B. subtilis spores, a reduction of two orders of magnitude of viability was confirmed following 20 min of treatment at 10 mg/mL at 60 . The active oxygen generated from the HESP slurry was examined with chemiluminescence. The intensity of this increased with increasing concentrations of the HESP slurry. This suggests that HESP could be used as a natural antimicrobial agent. Although a high pH is the main contributor to this antimicrobial activity, active oxygen species generated from HESP are likely to be the main antimicrobial agents..
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