Successful cultivation requires continuous monitoring of physicochemical parameters, that is, pH, temperature, oxygen concentration, and nutrient status. The basic bio logical method used is microscopic examination to detect morphological changes and contamination by other microalgae and protozoa. Nutrient status can be followed by monitoring the concentration of nitrogen, using it as a measure for adding the proportional amounts of other nutrients. In mass cultivation ofmicroalgae, monocultures are usually required for biomass exploitation. The appearance of 'contaminants' (other microalgae as well as protozoa, bacteria, or fungi) might indicate that the cultivated culture has come under stress. Contaminants often represent one of the major limitations to large scale production in microalgal cultures, particularly with strains that cannot be grown in a selective medium out doors. For the cultivation of some microalgae (e.g., Haematococcus), the use of a closed system becomes mandatory.
A sufficient mixing of the microalgal suspension is necessary to ensure nutrient diffusion and a homogeneous light supply to the cells, as well as to prevent the accu mulation of oxygen in the culture, particularly when they are grown in a closed system. Indeed, excessive oxygen accumulation in a culture can promote photoinhibition of photosynthesis and a decline in growth. On the other hand, excessive mixing can cause hydrodynamic or sheer stress to the cells, and consequently a similar reduc tion of productivity.
Biophysical and biochemical monitoring methods gen erally reflect the status of the cells' photosynthetic apparatus and are used to adjust the appropriate cultiva tion conditions for the production of biomass or certain compounds. The concentration of dissolved oxygen mea sured by an oxygen electrode is considered as a reliable and sensitive indicator of photosynthetic activity in microalgal cultures.
Recently, chlorophyll fluorescence has become one of the most common and useful approaches used for monitoring the physiological status of cultures. Its non invasiveness, sensitivity, ease of use, as well as its promptness make it a convenient and suitable technique in microalgal biotechnology. The ratio Fv/Fm (variable to maximum fluorescence yield) is considered to be a convenient measure of the performance of photochemi cal processes in photosystem II (the PSII photochemical yield): it relates the utilization of absorbed light energy to primary production. A decline in the Fv/Fm ratio can be considered as a reliable warning signal of culture stress.
Culture growth might be estimated as changes of optical density (OD) at 750 nm, biomass dry weight, or the number of cells. The specific growth rate is usually estimated as p (h J) = (ln X2 — ln Xj)/(t2 — t1), where X is cell number or dry weight at various times. Biomass productiv ity can be expressed as the areal or volumetric yield per unit time, that is, in gm 2 day 1 or in gl 1 day 1, respectively.
Basically, two cultivation regimes are used for the growth of microalgal cultures. In the batch regime, the culture is inoculated and at a certain point of growth it is harvested. In the continuous regime, the culture is har vested continuously according to its growth rate and fresh medium is added to replace nutrients. In practice, semi continuous or semibatch regimes are usually adopted, that is, where a part of the culture is harvested at regular intervals.
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