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Applied and Environmental Microbiology, December 2003, p. 7395-7400, Vol. 69, No. 12
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.12.7395-7400.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Application of a Specific and Sensitive Radiometric Assay for Microbial Lipase Activities in Marine Water Samples from the Lagoon of Nouméa

Nicolas Bourguet,1* Jean-Pascal Torréton,2 Olivier Galy,1 Vincent Arondel,3 and Madeleine Goutx1

Laboratoire de Microbiologie Marine, Université de la Méditerranée, Centre d'Océanologie de Marseille, Campus de Luminy, 13288 Marseille Cedex 09,1 and Laboratoire de Lipolyse Enzymatique, Université de la Méditerranée, Institut de Biologie Structurale et Microbiologie, 13402 Marseille Cedex 20, France,3 Unité de Recherche 103, Institut de Recherche pour le Développement, Centre IRD de Nouméa, 98848 Nouméa Cedex, New Caledonia2

Received 21 February 2003/ Accepted 8 September 2003

Marine microbiologists commonly assay lipase activities by using a synthetic fluorescent analog, 4-methylumbelliferyl (MUF)-oleate. The technique is convenient, but it is considered to be unspecific because of the structure of this analog. This study reports the design of a new specific and sensitive lipase assay based on the use of a radiolabeled triglyceride, [3H]triolein. Free fatty acids (FFA) resulting from its hydrolysis are isolated as a function of time in a one-step liquid-liquid extraction and then radioassayed. MUF-oleate and [3H]triolein techniques were compared by measuring lipase activities at similar substrate concentrations along a trophic gradient in the Southwest Lagoon of New Caledonia, near Nouméa. Hydrolysis rates decreased from the nearshore station to the offshore station and showed similar trends regardless of the technique used. Rates decreased from 5.83 to 0.88 nmol of FFA · liter-1 · h-1 and from 0.76 to 0.23 nmol of 3H-FFA · liter-1 · h-1, respectively. These results appeared to be consistent with bacterial production results, which also decreased similarly (from 0.59 to 0.26 µg of C · liter-1 · h-1). However, the ratio of MUF-oleate activities to [3H]triolein activities, which was constant at the offshore stations (3.8 ± 0.1), gradually increased at the nearshore stations (from 4.1 to 7.6). This result shows that the two assays respond in different ways to changes in environmental conditions and validates the need to set up more specific enzymatic assays.

* Corresponding author. Mailing address: Laboratoire de Microbiologie Marine, UMR 6117, CNRS, Université de la Méditerranée, Centre d'Océanologie de Marseille, Campus de Luminy, Case 907, 163, Avenue de Luminy, 13288 Marseille Cedex 09, France. Phone: 33-4-91-82-90-62. Fax: 33-4-91-82-90-51. E-mail: bourguet{at}com.univ-mrs.fr.

Applied and Environmental Microbiology, December 2003, p. 7395-7400, Vol. 69, No. 12
0099-2240/03/$08.00+0     DOI: 10.1128/AEM.69.12.7395-7400.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.





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