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. Ectoenzymatic activity, prokaryotic heterotrophicabundances and production were determined in the Mediterranean Sea. Samplingwas carried out in the sub-surface, the deep chlorophyll maximum layer(DCM), the core of the Levantine intermediate waters and in the deeper partof the mesopelagic layers. Michaelis–Menten kinetics were assessed using alarge range of concentrations of fluorogenic substrates (0.025 to 50  µ M). As a consequence, Km (Michaelis–Menten half-saturation constant) and Vm (maximum hydrolysis velocity) parameters were determined for both low- andhigh-affinity enzymes for alkaline phosphatase, aminopeptidase (LAP) and β -glucosidase ( β GLU). Based on the constant derived from thehigh-LAP-affinity enzyme (0.025–1  µ M substrate concentration range),in situ hydrolysis of N proteins contributed 48 %  ±  30 % to theheterotrophic bacterial nitrogen demand within the epipelagic layers and180 %  ±  154 % in the Levantine intermediate waters and the upperpart of the mesopelagic layers. The LAP hydrolysis rate was higher thanbacterial N demand only within the deeper layer and only when consideringthe high-affinity enzyme. Based on a 10 % bacterial growth efficiency, thecumulative hydrolysis rates of C proteins and C polysaccharides contributedon average 2.5 %  ±  1.3  % to the heterotrophic bacterial carbondemand in the epipelagic layers sampled (sub-surface and DCM). This studyclearly reveals potential biases in current and past interpretations of thekinetic parameters for the three enzymes tested based on the fluorogenic-substrate concentration used. In particular, the LAP  / β GLU enzymaticratios and some of the depth-related trends differed between the use ofhigh and low concentrations of fluorogenic substrates.

biogeosciences

Spatial patterns of ectoenzymatic kinetics in relation to biogeochemical properties in the Mediterranean Sea and the concentration of the fluorogenic substrate used