Determination of nutrient uptake kinetic parameters: a comparison of methods

Abstract

Three methods which have been used to determine nutrient uptake hnetic parameters were compared using steady-state NH,-limited cultures of the chrysophyte Pseudopedinella pyriforrnis. The first 2 methods involved a multiple flask incubation where different concentrations of substrate were added to each flask. Method 1 used a variable incubation time, while the incubation time of Method 2 was short and constant. The third method, the perturbation method, involved one large addition of the substrate to one culture and hence the nutritional past history varied throughout the experiment. This method was used also with the diatoms Skeletonema costatum and Chaetoceros debilis in Si-Limited chemostats. Results inhcate that, for nutrient-limited cultures, kinetic parameters are best estimated using multiple additions of the substrate and a short constant incubation time (Method 2). It appears that this method determines membrane transport, which is still not completely free of feedback inhibition even when the incubation time is very short (e.g. 2 min). The short incubation time is necessary because the maximal uptake velocity (V,) decreases with increasing incubation time especially for phosphate and ammonium. Method 3 provides valuable information on a third parameter, V,, the approximate assimilation rate of the limiting nutrient, that is not obtained by the other methods. Multlple sequential additions of the limiting nutrient to N-or Si-limited Skeletonema costaturn and Chaetoceros debilis revealed that if the additions were small (e.g. 2 pM NH,), there was no change in subsequently determined nutrient uptake kinetic parameters. If the sequential additions were larger (e.g. 6 PM) then the maximal uptake rate slowed with time.

Variability in Nitrate Uptake Kinetics of Phytoplankton Communities in a Mediterranean Coastal Lagoon

“…Distortions in uptake curves result when the uptake rate is not constant with time during the incubation period (Conway et al, 1976;Collos, 1983;Harrison et al, 1989). This was the case in several instances in the present study, when cumulative nitrate uptake was not related to time in a linear manner (Figure 1).…”

“…Other phytoplankton groups or genera of diatoms are presented in Table 2. Figure 1 shows representative trends in cumulative nitrate uptake as a function of time, and illustrates the three known patterns of uptake for such nutrients (Collos, 1983;Harrison et al, 1989): induced (May and August), constant (November) and surge (February, June and September) uptake [the latter following the terminology of Conway et al (1976)]. This classification has been used because it has implications on the interpretation of the shape of the uptake vs. concentration curves.…”

“…Within 1 h of sampling, the water was dispensed in six 2·5 l polycarbonate bottles and enriched with 15 N-labelled sodium nitrate at the following final concentrations: 1, 5, 10; 20, 50 and 100 M. They were incubated at the surface of the lagoon for 4 h (generally from 1000 to 1400 h). An additional 8 l bottle was used at 100-M enrichment and aliquots were taken every hour over 4 h in order to test the constancy of uptake with time (Harrison et al, 1989). Samples were filtered using GF/F membranes, dried for 24 h at 60 C and stored desiccated.…”

Quantifiying external and internal nitrogen and phosphorus pools, as well as nitrogen and phosphorus supplied through remineralization, in coastal marine plankton by means of a dilution technique

“…Under full nutrient enrichment it is reasonable to assume that the specific growth rate will be constant (and equal to the maximum growth rate possible under the prevailing physical conditions). In the selectively enriched treatments, the assumption of a constant rate is invalidated by the known fact that instantaneous growth rates decline as internal and external nutrient pools become depleted (Goldman & Glibert 1983, Turpin 1988, Harrison et al 1989, and eventually reach zero. Under such conditions , U cannot be expected to be constant either with time or with dilution, and is therefore denoted as p(x,t).…”

The Nutritional Eco-physiology of Chaetomorpha linum and Ulva rigida in Peel Inlet, Western Australia

“…Due to logistical constraints it was not possible to conduct continuous flow experiments. Probyn and Chapman (1982), Gilbert and McCarthy (1984) and Harrison et al (1989) point out that shortterm, batch experiments tend to exaggerate normal uptake rates, particularly of ammonium. Alternatively, it can be argued that short-term experiments do not exaggerate uptake rates, but rather they disclose initial uptake rates which are often higher than rates determined over extended periods.…”