Development of Non-Forage Based Incubation System For Culturing Ruminal Lipase-Producing Bacteria In Vitro

Published 12/2013

Volume 3 Issue 4
Pp. 293-303

Keywords: , , , , , , , , ,

Abstract:

Lipids are often used as substrates when measuring growth and lipolytic activity of lipase-producing bacteria, however, the introduction of non-miscible lipid substrates into aqueous in vitro culture systems is problematic for generating accurate and consistent results. The objective of the present study was to develop a digesta-free method for culturing and assaying lipolytic activity of mixed as well as pure populations of known ruminal lipase-producing bacteria. Accordingly, the inclusion of 0, 11, and 21 g of glass beads as a solid support matrix in place of digesta was examined. Results showed a significant increase (P < 0.05) in rate of lipolysis in the incubations containing 11 or 21 g glass beads compared to the non bead incubations. Activity was also increased (P < 0.05) in a separate study when tubes containing beads were incubated horizontally rather than vertically. These results indicate that glass beads are a suitable substitute for rumen digesta when examining lipolytic activity of mixed rumen cultures in vitro. When tested against pure cultures of the ruminal lipase-producing bacteria Anaerovibrio lipolyticus 5s, Butyrivibrio fibrisolvens 49, Propionibacterium avidum and acnes, addition of glass beads did not significantly increase rates of free fatty acid release; however, results showed that there was substantial variation between triplicate sets incubated without glass beads. Standard deviations suggest that the use of glass beads had a tendency to reduce variability within triplicate sets. Thus, inclusion of glass beads provides a clean and consistent incubation system for examining lipase activity in vitro.


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