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Reactor design is another relevant issue that determines the process yield
[35, 36, 45]. As mentioned above, for outdoor operation, closed tubular photobiore-
actors are most efficient, not only because of the high sensitivity of Haematococcus
to contamination, but also because of the higher light availability inside this type of
reactor in comparison to other systems [46].
3 The Alternative: The One-Step Strategy
The continuous culture system has recently been proposed as the approach of choice
for unequivocal identification of factors influencing the accumulation of astaxan-
thin in H. pluvialis [25]. To this end, H. pluvialis cultures in photo-chemostats
were maintained in continuous growth regime under steady state conditions. This
approach allows to analyze the influence of a single variable while holding all
other parameters constant, thus avoiding interferences in the interpretation of
results. An in-depth analysis of factors determining astaxanthin accumulation in
Haematococcus has been thus performed under continuous regime. The effective-
ness of such a system, not only for analytical but also for production purposes, has
led to the development of a one-step only strategy [25, 48].
The possibility of maintaining Haematococcus pluvialis cells under conditions
promoting astaxanthin accumulation while maintaining full growth capacity had
not received much attention until now. In the traditional two-step process, astax-
anthin accumulation is induced through severe stress conditions that determine the
transition of vegetative cells to cysts. The one-step system takes advantage of the
capability of growing cells to accumulate astaxanthin. In continuously operated
photoautotrophic cultures, nitrogen supply to the system is controlled to main-
tain a moderate nitrogen limitation that allows astaxanthin accumulation, without
markedly affecting growth [25, 48]. This leads to high astaxanthin productivity,
on the basis of maintaining a cell population that accumulates astaxanthin at a
significant level, keeping besides a high growth rate (0.9 d -1 ).
Performance of the alternative system has been validated for a variety of com-
binations of dilution rate, nitrate concentration in the feed medium, and incident
irradiance ( I o ). Dilution rate and nitrate concentration in the feed medium deter-
mine the nitrogen supply to the cells. Both factors, and especially the dilution rate,
markedly affect population density. The most adequate parameter to quantify the
nitrate supply to the cells is the “specific nitrate input”: SNI
[NO 3 - ] D / C b , where
[NO 3 - ] is the concentration of nitrate in the feed medium (mM); D , the dilution rate
(d -1 ); and C b , the biomass concentration (g dry biomass l -1 ). SNI , therefore, refers
to the amount of nitrate (mmol) made available to cells (g dry biomass) per time
unit (d).
Figure 2 shows, for constant incident irradiance, the influence of SNI on the yield
of both biomass and astaxanthin in continuous culture of H. pluvialis . Lowering the
SNI value up to a certain extent determines a slight reduction in biomass produc-
tivity, while a drastic increase in astaxanthin productivity is recorded under these
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