Nutrient Optimization And Control In Microalgae Cultivation

Microalgae are capable of utilizing sunlight and carbon dioxide to synthesize organicsubstrates such as protein, carbohydrate and lipid, and oxygen is produced at the same time.Microalgae are widely used in food, health care, medicine, environment and bio-energy fields.Considering the problem of high production cost and low productivity in microalgaecultivations, the nutrient salts were chosen to be the main investigating objects. The influenceof the main nutrient on Spirulina platensis and Scenedesmus dimorphus growth andproduction were studied. The relationships among the changes of pH and inorganic carbonsource, the consumption of nitrogen and phosphorus sources and the biomass accumulationwere learned in this study. Based on the relationship between the microalgae growth andnutrient consumption, three kinds of nutrient feeding strategies were proposed and compared.Furthermore, the nutrient feeding strategies were applied for nutrient optimization inSpirulina platensis cultivation and regulation of lipid accumulation in Scenedesmusdimorphus cultivation. As a result, the operating and production cost related to nutrient inmicroalgae cultivations were optimized and controlled by using these nutrient feedingstrategies. Main results were as follows:(1) In Spirulina platensis cultivation, the pH value of culture medium exceeded11.0atthe late growth phase using NaHCO₃as carbon source, which would lead to growth inhibition.Feeding CO₂as carbon source could maintain the pH value of the culture medium at thesuitable level for Spirulina platensis growth (8.0¹0.0). It was found that sufficient carbon,nitrogen and phosphorus sources were benefit for Spirulina platensis growth and high proteinand chlorophyll content could be obtained. But with excess carbon source, nitrogen-limitationor phosphorus-limitation, Spirulina platensis growth could be affected and the accumulationof carbohydrate was promoted. Urea, NH₄NO₃, NH₄Cl and NH₄HCO₃could instead ofNaNO₃as nitrogen source for Spirulina platensis cultivation, but the growth inhibition wouldtake place when the concentration of ammonium was at a relatively high level. The order ofthe inhibitory effect of these nitrogen sources was NH₄HCO₃> NH₄Cl>NH₄NO₃>urea.(2) When the nutrient concentration was relatively low in the culture medium, thespecific consumption rate of nutrient was not only depended on the specific growth rate ofmicroalgae, but also on the nutrient cell quota of microalgae. When the nutrient was relativelysufficient in the culture medium, the cell quota of carbon, nitrogen and phosphorus in themicroalgae cells were relatively stable, and the specific consumption rate of nutrient wasmainly depended on the specific growth rate of microalgae. In the latter situation, the amount of CO₂addition to keep the pH value of culture medium stable, the consumption of nitrogensource and the consumption of phosphorus source were linearly correlated to theaccumulation of biomass. Thus three different nutrient feeding strategies were proposed fornutrient concentrations control in microalgae cultivations. They were the pH and nitrateelectrodes based feedback feeding method, the pH-based feedback feeding method and theabsorbance-based feedback feeding method.(3) Some probable factors affecting the change of nutrients concentration when using theabsorbance-based feedback feeding method and the pH-based feeding method wereinvestigated. The results implied that the fluctuation range of the nutrient concentrations inthe culture medium depended on the absorbance-step, utilization of nutrient and nutrient cellquota using the absorbance-based feedback feeding method. And the fluctuation range ofnutrient concentrations in the culture medium depended on the pH-step, pH value of culturemedium and free CO₂concentration in the culture medium using the pH-based feedingmethod. The proposed nutrient feeding strategies were applied in the Spirulina platensisfed-batch cultivations. It was found that when the pH and nitrate electrodes based feedbackfeeding method was used, the nitrogen and phosphorus concentrations were fluctuant in arelatively wide range due to the potential drift of the nitrate electrode. It implied this feedingmethod was not fit for long-time nutrient monitoring and controlling on-line. Theabsorbance-based feedback feeding method was benefit for the nitrogen and phosphorusconcentration control, while the pH-based feedback feeding method was benefit for the pHand carbon concentration control. Combining the pH-based feedback feeding method with theOD-based feedback feeding method, the carbon, nitrogen and phosphorus concentrationswould be accurately controlled during the microalgae cultivations,which implied they werefit for long-time nutrient control on-line.(4) The nutrient in the Spirulina platensis cultivation was optimized using the feedingstrategies above. Meanwhile, feeding CO₂as the carbon source using the pH-based feedbackfeeding method, the maximum biomass concentration of3.30g/L could be obtained, whichwas92.30%higher than that of using NaHCO₃as the carbon source. Feeding NH₄HCO₃asthe nitrogen source by the absorbance-based feedback feeding method to maintain theammonium concentration in the culture medium at1mmol/L, Spirulina paltensis growthwould not be affected by nitrogen source inhibition or limitation. As a result, the maximumbiomass concentration (2.98g/L), nitrogen-to-cell conversion factor (7.32g/g), and contentsof protein (64.11%) and chlorophyll (13.40mg/g) obtained by using absorbance-basedfeedback feeding method were higher than those of using the constant, variable and pH-based feedback feeding methods.(5) The nutrient optimization and feeding strategies proposed in the thesis were appliedin large-scale cultivations of Spirulina platensis in outdoor open raceway pond. In thetraditional semi-continuous cultivations, the carbon, nitrogen and phosphate sources weresodium bicarbonate, sodium nitrate and dipotassium hydrogen phosphate, respectively andnutrient feeding was based on experience after harvest. When the carbon, nitrogen andphosphate sources were substituted with the carbon dioxide, sodium nitrate and ammoniumcarbonate, and phosphate, respectively, and the feeding method was substituted withabsorbance-based or pH-based feedback feeding method in the outdoor large-scalecultivations, the carbon, nitrogen and phosphorus source concentrations in the culture mediumcould be maintained in the optimum range for Spirulina platensis growth. Meanwhile, thecost of culture medium decreased by about70%and the productivity per area of Spirulinaplatensis improved by more than20%.(6) The results of batch cultivations of Scenedesmus dimorphus showed that theoptimum pH for Scenedesmus dimorphus was7.5. Compared with the situation of usingNaNO₃and urea as the nitrogen source, a faster growth and higher lipid productivity could beobtained using NH₄HCO₃as the nitrogen source. In carbon-sufficient but nitrogen-orphosphorus-limited cultures, the lipid production of Scenedesmus dimorphus could beincreased. Furthermore, the promoting effect of nitrogen limitation on lipid accumulation wasmore than that of phosphorus limitation. Based on the batch cultivation results, CO₂wasadded using the pH-based feedback feeding method and NH₄HCO₃and K2HPO4were addedusing the absorbance-based feedback feeding method in the fed-batch cultivation ofScenedesmus dimorphus. The pH value of the culture medium could be maintained at7.5±0.3.And Scenedesmus dimorphus grew well in the nitrogen and phosphorus sources feeding stage.After the feeding end of nitrogen and phosphorus sources, nitrogen and phosphorus sourcesexhaustion quickly occurred and the accumulation of lipid was promoted at the same time.Compared with the batch cultivations, the constant feeding cultivations with nitrogen feedingrate of0.8mmol/(L·d) and phosphorus feeding rate of9.6μmol/(L·d), and the constantfeeding cultivation with nitrogen feeding rate of0.4mmol/(L·d) and phosphorus feeding rateof4.8μmol/(L·d), the lipid productivity in the fed-batch cultivations using the proposedfeeding method increased by58.2%,53.3%and37.1%, respectively.