Measurement of lightdependent photosynthetic oxygen production

Numerous studies based on oxygen generation in light and darkened bottles were published in the 50 years between 1935 and 1985. Many of the findings were substantially covered in a thorough synthesis by Harris (1978). Since then, the method has been displaced by more direct and more sensitive techniques. Nevertheless, the experiments based on measurements of photo-synthetic oxygen production in closed bottles suspended at selected depths in the water column yielded consistent generalised results and have bequeathed to plankton science many of the conceptual aspects and quantitative descriptors of productive capacity. The set of sample results illustrated in Fig. 3.3 depicts a typical depth profile of photosynthetic (4-h) exposures of unmodified lake plankton in relation to the underwater light field in an unstratified temperate lake in winter (temperature ~5 oC). The features of general interest include the following:

• The plot of gross photosynthetic oxygen generation is shown as a function of depth in Fig. 3.3a. The curve is fitted by eye. However, it is plain that photosynthesis over the 4 hours peaks a little way beneath the surface, with slower rates being detected at depth. In this instance, as in a large number of other similar experiments, there is an apparent depression in photosyn-thetic rate towards the surface.

• The 'gross photosynthesis' is the measured aggregate oxygen production at a given depth averaged over the exposure period. It is calculated as the observed increase in oxygen concentration in the 'light' bottle (usually a mean of duplicates) plus the observed decrease in concentration in the corresponding 'dark'

Figure 3.3

Specimen depth distributions of (a) total gross photosynthetic rate (NP) and total gross respiration rate (NR); (b) the photosynthetic population (N), in terms of chlorophyll a; (c) chlorophyll-specific photosynthetic rate (P = NP/N) and respiration rate (R = NR/N); (d) underwater irradiance (I) in each of three spectral blocks, expressed as a percentage of the irradiance 1( obtaining immediately beneath the surface. P is replotted against I, either (e) as a percentage of I ( in th green spectral block (peak: 530 nm), or as the reworked estimate of the intensity of the visible light above the water (in |imol photon m-2 s-1) averaged through the exposure period. Original data of the author, redrawn from Reynolds (1984a).

Figure 3.3

Specimen depth distributions of (a) total gross photosynthetic rate (NP) and total gross respiration rate (NR); (b) the photosynthetic population (N), in terms of chlorophyll a; (c) chlorophyll-specific photosynthetic rate (P = NP/N) and respiration rate (R = NR/N); (d) underwater irradiance (I) in each of three spectral blocks, expressed as a percentage of the irradiance 1( obtaining immediately beneath the surface. P is replotted against I, either (e) as a percentage of I ( in th green spectral block (peak: 530 nm), or as the reworked estimate of the intensity of the visible light above the water (in |imol photon m-2 s-1) averaged through the exposure period. Original data of the author, redrawn from Reynolds (1984a).

control, it being supposed that the respiration in the dark applies equally to the similar material in the light. The mean gross photosynthetic rate at the given depth (here, expressed in mg O2m-3h-1) represents NP, the product of a biomass-specific rate of photosynthesis (P, here in mg O2 (mg chla)-1 h-1) and the biomass present (N, shown in Fig 3.3b and expressed as the amount of chlorophyll a in the enclosed plankton, in (mg chla) m-3). At this stage, it is the biomass specific behaviour that is of first interest: P (=NP/N) is plotted in Fig. 3.3c. The shape of the curve of P is scarcely distorted from that of NP in this instance, owing to the uniformity of N with depth. Discontinuities in the depth distribution of N do affect the shape of the NP plot (see below and Fig. 3.4).

It was neither essential nor common to calculate the respiratory uptake, as the depletion of oxygen concentration in the dark controls. All that was necessary was the additional measurement of the initial oxygen concentration at the start of the experiment. The change in the dark bottles over the course of the experiment, normalised to the base period, is the equivalent to the respiration of the enclosed organisms in the dark, NR (here, expressed in mg O2m-3h-1); R (=NR/N) is thus supposed to be the biomass-specific respiration rate (in mg O2 consumed (mg chla)-1 h-1) inserted in Fig. 3.3c. However, the extrapolation must be applied cautiously. Whereas NP, as determined, is attributable to photosynthesis, the separate determination of NR does not exclude respiration by non-photosynthetic organisms, including bacteria and zooplankton. Moreover, it cannot be assumed that even basal respiration rate of phytoplankton is identical in light and darkness: according to Geider and MacIntyre (2002), oxygen consumption in photosynthesising microflagellates is 3- to 20- (mean: 7-) fold greater than dark respiration rate. Accelerated metabolism and excretion of photosynthate in starved or stressed phytoplankton may, conceivably, account for all the materials fixed in photosynthesis: (P-R) is small but dark R need not be large either.

Chlorphyll Depth Profile

Figure 3.4

Some variations in the basic form of depth profiles of gross (NP) and chlorophyll-specific rates (P), explained in the text.

Figure 3.4

Some variations in the basic form of depth profiles of gross (NP) and chlorophyll-specific rates (P), explained in the text.

The decline in biomass-specific P below the subsurface maximum (Pmax) is supposed to be a function of the underwater extinction. In the present example, the underwater light attenuation measured photometrically in each of three spectral bands of visible light (blue, absorption peaking at 430 nm; green, peaking at 530 nm; and red, peaking at 630 nm) is included as Fig. 3.3d. Note that the spectral balance changes with depth (blue light being absorbed faster than red or green light in this case). Replotting the biomass-specific rates of P against Iz, the level of the residual light penetrating to each of the depths of measurement (expressed as a percentage of the surface measurement), gives the new curve shown in Fig. 3.3e. This emphasises the sensitivity of P to Iz, at least at low percentage residual light penetration, and a much more plateau-like feature around Pmax.

Finally, the P vs. I curve is replotted in Fig. 3.3f in terms of the depth-dependent irradiance across the spectrum, approximated from Iz =

(I430 + I530 + Is30)z/3, and taking the immediate subsurface irradiance (I0) as the average over the experiment (in this instance, I0 = 800 |imol photons m-2 s-1).

This last plot, Fig. 3.3f, conforms to the format of the generic P vs. I curve. It has a steeply rising portion, in which P increases in direct proportion to Iz; the slope of this line, generally notated as a (=P/Iz), is a measure of photosyn-thetic efficiency at low light intensities. Expressing P in mg O2 (mg chla)-1 h-1 and Iz in mol photons m-2 h-1, the slope, a, has the dimensions of photosynthetic rate per unit of irradiance, mg O2 (mg chla)-1 (mol photon)-1. As the incident photon flux is increased, P becomes increasingly less light dependent and, so, increasingly saturated by the light available. The irradiance level representing the onset of light saturation is judged to occur at the point of intersection between the extrapolation of the linear light-dependent part of the P - Iz curve with the back projection of Pmax, being the fastest, light-independent rate of photosynthesis measured. This intensity, known as Ik, can be judged by eye or, better, calculated as:

It is usual (because that is how the photon flux is measured) to give light intensities in |imol photons m-2s-1. In terms of being able to explain the shape of the original P vs. depth curve (Fig. 3.3a), it is also useful to be able to define the point in the water column, z(Ik) , where the rate of photosynthesis is directly dependent on the photon flux. The left-hand, light-limited part of the P vs. Iz curve is the most useful for comparing the interexperimental differences in algal performances.

Kirk (1994) described the several attempts that have been made to find a mathematical expression that gives a reasonable fit to the observed relationship of P to I. Jassby and Platt (1976) tested several different expressions then available against their own data. They found the most suitable expressions to be one modified from Smith (1936) and the one they themselves proposed:

(Smith, 1936) P = Pmax tanh(a - Iz/Pmax) (Jassby & Platt, 1976)

In contrast, the significance and mathematical treatment of the right-hand part of the P vs. Iz curve, corresponding to the apparent near-surface depression of photosynthesis in field experiments, was, for a long time, a puzzling feature. It was not necessarily a feature of all P vs. z curves: some of the possible variations are exemplified in Fig. 3.4. Anomalies in the depth distribution of NP, caused by phytoplank-ton abundance (Fig. 3.4a), low water temperature (Fig 3.4c) or by surface scumming of dominant Cyanobacteria (Fig. 3.4d) fail to obscure the incidence of subsurface biomass-specific photosyn-thetic rates. However, they are not seen when dull skies ensure that, even at the water surface, pho-tosynthetic rates are not light-saturated [z(Ik) < 0 m!].

This near-surface depression in measured pho-tosynthertic rate is not, however, reflected in growth and replication. In none of the experiments considered by Gran (1912), nor those carried out half a century later by Lund (1949) or Reynolds (1973a), is there any question that the fastest population growth rates are obtained closest to the water surface. In the late 1970s, some helpful experimental evidence was gathered to show that the surface depression was largely an artefact of the method and the duration of its application. Taking phytoplankton from a mixed water column and holding them steady at supersaturating light intensities for hours on end represents an enforced 'shock' or 'stress'. In these terms, it would not be unreasonable to conclude that the algae would react and to show signs of 'photoinhibition' (see below). Jewson and Wood (1975) showed that continuing to circulate the algae through the light gradient not only spared the algae the symptoms of photinhibition but that the measured Pmax could be sustained. In an analogous experiment, Marra (1978) showed that realistically varying the incident radiation received by phytoplankton avoided the apparent photoinhibition. Harris and Piccinin (1977) determined photosynthetic rates (from oxygen production, measured with electrodes rather than by titration) in bottled suspensions of Oocystis exposed to high light intensities (>1300 |imol photons m-2 s-1) and temperatures (>20 °C) for varying lengths of time. Their results suggested that an elevated photosynthetic rate was maintained for 10 minutes or so but then declined steeply with prolonged exposure. Either the algae were photoinhibited or damaged or they had reacted to prevent such damage (photoprotection) in some way that would enable to retain their vitality to maintain growth (see Section 3.3.4).

For these reasons, determinations of depth-integrated photosynthesis (ENP, in mg O2 m-2 h-1) need no longer need depend on the plani-metric measurement of the area enclosed by the measurements of photosynthetic rates against depth. They are estimable, for instance, from the P vs. Iz curve in Fig. 3.3f, as the area of a trapezium equivalent to Pmax x 2 [(Iq - Ik) + (I0 - Ic), where I0 is the light intensity at the water surface and Ic is the intercept of zero photosynthesis. In terms of P vs. depth, this simplifies to the product Pmax x (the depth from the surface to the point in the water column where the light will half-saturate it); i.e.

Figure 3.5

Hypothetical P vs. I plots to contrast seasonal variations in temperature on photosynthetic behaviour, with special reference to changes in Pmax (tagged) and Ik (arrowed). In either plot, the sequence a^b^c is one of increasing temperature. In the left-hand plot, the dependence upon light is constant; in the right-hand plot, P/I varies to keep Ik constant. Redrawn from Reynolds (1984a).

If the dark respiration rate, NR, is uniform with depth, then the integral is simply the product of the full depth range over which it applies (the height of the full water column, H) depth

Figure 3.5

Hypothetical P vs. I plots to contrast seasonal variations in temperature on photosynthetic behaviour, with special reference to changes in Pmax (tagged) and Ik (arrowed). In either plot, the sequence a^b^c is one of increasing temperature. In the left-hand plot, the dependence upon light is constant; in the right-hand plot, P/I varies to keep Ik constant. Redrawn from Reynolds (1984a).

Based upon the numerous published records, several compendia of the key indices of photosyn-thetic oxygen production of phytoplankton in closed bottles have been assembled (Harris, 1978; Kirk, 1994; Padisak, 2003). Clearly, the gross rate of photosynthesis (NPmax) and the depth integral (XNP) respond to two variables, which are, within limits, highly variable. Other factors notwithstanding, observed Pmax should always be the light-saturated maximum rate at the given temperature. Among the fastest reported examples are 30-32 mg O2 (mg chla)-1 h-1 (noted in warm tropical lakes in Africa, by Talling, 1965; Talling et al., 1973; Ganf, 1975), whereas the specific pho-tosynthetic rates in temperate lakes rarely exceed the 20 mg O2 (mg chla)-1 h-1 found by Bind-loss (1974). Thus, as might be expected, examples of high community rates of oxygen production come from warm lakes supporting large populations of phytoplankton algal chlorophyll and when there are also ample reserves of exploitable CO2: integrals in the range 6-18 g O2 m-3 h-1 have been noted in Lac Tchad, Chad (Leveque et al., 1972), Red Rock Tarn, Australia (Hammer et al., 1973) and Lake George, Uganda (Ganf, 1975). The onset of light saturation, Ik, is also temperature influenced but is generally <300 |imol photons m-2s-1. There are many citations of much lower Ik determinations, 15-50 |imol photons m-2 s-1, generally at temperatures <10 °C (Kirk, 1994). The most consistent values seem likely to relate to the slope, a, which, at low light levels, is less dependent on temperature and more dependent upon light-harvesting efficiency. Taking the plot in Fig. 3.3f, the slope a (i.e. P on I) is calcu lated by rearranging Eq. (3.5):

(|imol photons m-2 s-1 )-1 = 13.2mgO2 (mgchla)-1(molphoton)-1m2

Comparisons among differing P vs. I plots often differentiate patterns of photosynthetic behaviour. In the left-hand box of Fig. 3.5, the slopes (a) show similar light dependence of photosynthesis at low irradiances but the sequence of increasing Pmax values could be the response of the same alga to increasing temperatures. In the right-hand plot, data for three different algae are shown, all saturating at similar levels but with differing photosynthetic efficiencies. A higher a enables a faster rate of photosynthesis to be maintained when light intensity is low.

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