Introduction
The filamentous cyanobacterium Arthrospira platensis (Spirulina)is an oxygenic photosynthetic organism able to grow in tropical and subtropical environments, and one of only a few microalgal systems that has been successfully commercialized and approved by United States Food and Drug Administration (FDA) as a food supplement (Trabelsi et al., 2009). Arthrospira cultivation and processing yields valuable biochemical components including protein, carbohydrates, fatty acids and pigments such as phycocyanin (PC), which can be used in nutritional, pharmacological, and cosmetic products. Due to these high value applications, as well as relatively easy harvesting and extraction processes, Arthrospira cultivation has been deployed commercially at moderate scale (10 – 100 acre open ponds) for many decades (Lu et al., 2011). It is important to note that Arthrospira is an extremophile, in that it can maintain high productivity under high alkalinity, high pH conditions; this limits predation and competition sufficiently to allow commercial production in open pond systems. Algal cultures are influenced by various abiotic variables such as temperature, irradiance levels, and nutrient availability, all of which play a significant role in regulating photosynthetic activity, biomass composition and overall productivity. Under outdoor cultivation conditions, temperature and light intensity are the two key external factors that determine photosynthetic activity and biomass growth rates. Obviously, both factors are highly variable on a daily and seasonal basis in the natural environment, and spatially within the culture as well (Chaiklahan et al., 2007; Vonshak and Novoplansky, 2008). Typically, Arthrospira is cultivated outdoors for mass production in raceway ponds, where cells encounter fluctuating environments in terms of irradiance, temperature, and nutrient supply. Though the PBR environment tends to be more homogeneous, similar fluctuations are present and temperatures are generally higher due to absorptive heating and the absence of evaporative cooling. Outdoor algal cultures are subjected to high light intensity as well as possible high temperature stress that can negatively impact photosynthetic activity (Torzillo et al., 1991b). These factors can change both the photosynthesis and respiration rates, thereby directly influencing the growth and the chemical composition of the biomass produced (Trabelsi et al., 2009).
Overall, the existing literature is consistent with an optimal temperature range for stable production of roughly 20-35 °C. Our screening studies are consistent with that range and also consistent with an activation energy of about 60 kJ mole-1 (Q10 ~2) under saturating light conditions over that temperature range. It is well-known that productivity is enhanced in semi-continuous operation where the impact of photosaturation effects is lessened. We know of no detailed studies dealing with the effect of temperature and acclimation response on growth and pigment content ofArthrospira in a semi-continuous production mode for extended time scales under tightly controlled (laboratory) conditions. The intention here is to determine what portion of previous learnings translate to semi-continuous operation and the dynamic (light/temperature) conditions experienced outdoors, and examine the responses to abrupt changes in temperature. Therefore, in the present work we will examine temperature effects at a longer time scale, and carry out the experiments in semi-continuous operation mode in PBRs at 20 °C, 30 °C and 35 °C. In subtropical conditions, the outdoor culture temperature in the summer months can be very high in PBRs, reaching up to 35-45 °C for several hours. We have only a limited understanding of temperature impacts on photosynthetic parameters, and pigment accumulation in that outdoor environment. Thus, the scope of this work includes Arthrospira growth under a variety of temperature conditions with a work plan that includes assessment of temperature response and recovery, and quantification of the dynamic change in biomass and pigment content of Arthrospira during the experiments. There is no doubt that high irradiance levels can be a confounding factor both at low and high temperatures. We limit the current study to “average” irradiance conditions in Fort Myers, Florida. The combination of vertical PBR arrays, which dilute the average irradiance levels from about 800 μE/m2-s to about 200 μE/m2-s, and the rapid mixing, which distributes the heat from light absorption more evenly within the PBR volume, lessen the potential for extreme effects due to high irradiance. That is born out by cultivation field observations of Arthrospira growth and laboratory studies of irradiance effects with the same approach used here. Regarding lower temperatures than the 20°C included here, our screening studies do not suggest any issues down to 10°C and field experience in the environment of interest shows that the concerns lie at high temperatures.
The work was performed in three phases (Figure 1a): Phase I employs constant temperature conditions (same for day and night cycles), Phase II shifts the Phase 1 cultures to opposing temperature conditions (low to high, and high to low), and Phase III continues the examination under dynamic summer temperature profiles (hourly variations) in a semi-continuous operation mode. The experimental setup, shown in Figure 1b, involves vertically oriented tubular photobioreactors, designed to be predictive of outdoor performance in large PBR arrays.