Changhong Yao

Enhancing starch production of a marine green microalga Tetraselmis subcordiformis through nutrient limitation

Microalgal starch is a potential feedstock for biofuel production. The effects of KNO(3) and MgSO(4) concentrations and light intensity on biomass and starch production by the marine microalga, Tetraselmis subcordiformis, were investigated. Under 200 μmol m(-2) s(-1) irradiance and sulfur-deprived conditions, a starch productivity of 0.62 g L(-1) d(-1) and a starch content of 62.1% based on dry weight (DW) was achieved. A starch content of 54.3% was achieved under low irradiance and nitrogen starvation, which was 6.5% higher than that under nutrient- and light-sufficient conditions. Photosynthetic activity was indispensable for starch accumulation. It is difficult to reach high starch productivity and starch concentration simultaneously. Proper nutrient concentrations are necessary to achieve high starch productivity or starch concentration based on the target. The high starch productivity and starch content suggest that T. subcordiformis is a promising microalgal starch producer.

Application of Fourier transform infrared (FT-IR) spectroscopy in determination of microalgal compositions

Fourier transform infrared spectroscopy (FT-IR) was applied in algal strain screening and monitoring cell composition dynamics in a marine microalga Isochrysis zhangjiangensis during algal cultivation. The content of lipid, carbohydrate and protein of samples determined by traditional methods had validated the accuracy of FT-IR method. For algal screening, the band absorption ratios of lipid/amide I and carbo/amide I from FT-IR measurements allowed for the selection of Isochrysis sp. and Tetraselmis subcordiformis as the most potential lipid and carbohydrate producers, respectively. The cell composition dynamics of I. zhangjiangensis measured by FT-IR revealed the diversion of carbon allocation from protein to carbohydrate and neutral lipid when nitrogen-replete cells were subjected to nitrogen limitation. The carbo/amide I band absorption ratio had also been demonstrated to depict physiological status under nutrient stress in T. subcordiformis. FT-IR serves as a tool for the simultaneous measurement of lipid, carbohydrate, and protein content in cell.

Salinity manipulation as an effective method for enhanced starch production in the marine microalga Tetraselmis subcordiformis.

Microalgal starch is considered a promising feedstock for bioethanol production. The biomass and starch accumulation in the marine microalga Tetraselmis subcordiformis were characterized under different salinities in response to nitrogen repletion (+N) or depletion (-N) at high irradiance (HI) or low irradiance (LI). Under favorable nutritional conditions (HI+N), biomass accumulation was seldom affected under 20% normal salinity, though starch accumulation were somewhat reduced. Increased salinity impaired overall biomass and starch accumulation, though it led to a temporary starch accumulation at initial cultivation phase. Under nitrogen deprivation, decreased salinity strengthened biomass and starch accumulation regardless of irradiance. The highest starch content of 58.2% dry weight and starch productivity of 0.62 g L(-1) d(-1) were obtained under HI-N with 20% normal salinity. Decreased salinity combined with -N generated moderate stress to facilitate starch accumulation. Salinity manipulation can be effectively applied for enhanced starch production in marine microalgae.

Identification of fatty acid biomarkers for quantification of neutral lipids in marine microalgae Isochrysis zhangjiangensis

Fatty acid biomarkers of the marine microalga Isochrysis zhangjiangensis in response to nitrogen depletion were identified by multivariate statistical analysis. Seven fatty acids (C16:0, C18:1n9, C18:4n3, C18:3n3, C18:5n3, C22:6n3, and C16:1n7) were selected and identified as potential biomarkers, among which C18:1n9 and C18:4n3 were enriched in the neutral lipids and glycolipids, respectively. Due to significant changes and specific distributions in different types of lipids, C18:1n9 and C18:4n3 were identified as biomarkers to quantify neutral lipid content. C18:1n9 was positively related with neutral lipid content, while C18:4n3 was negatively related with neutral lipid content. The correlation coefficients of C18:1n9 and C18:4n3 content in total fatty acids and neutral lipid content were 0.9960 and 0.9787, respectively. The good agreement between the neutral lipids content and fatty acid biomarkers content in total fatty acids indicates that neutral lipids can be quantified by specific fatty acids. Using fatty acid biomarkers to quantify neutral lipids requires much less sample analyzed by gas chromatography, which is a universal method, plus the simplicity of operation compared with conventional methods. The striking feature of the approach is individual fatty acid to represent the type of lipids. Therefore, applying fatty acid biomarkers to identify specific type of lipids promises to maximize lipid-related information and can be used to make further discoveries related to lipid metabolism.

Identification of carbohydrates as the major carbon sink of the marine microalga Isochrysis zhangjiangensis (Haptophyta) and optimization of its productivity by nitrogen manipulation.

Microalgae represent a potential feedstock for biofuel production. During cultivation under nitrogen-depleted conditions, carbohydrates, rather than neutral lipids, were the major carbon sink of the marine microalga Isochrysis zhangjiangensis (Haptophyta). Carbohydrates reached maximum levels of 21.2 pg cell(-1) on day 5, which was an increase of more than 7-fold from day 1, while neutral lipids simultaneously increased 1.9-fold from 4.0 to 7.6 pg cell(-1) during the ten-day nitrogen-depleted cultivation. The carbohydrate productivity of I. zhangjiangensis was improved by optimization of the nitrate supply mode. The maximum carbohydrate concentration was 0.95 g L(-1) under batch cultivation, with an initial nitrogen concentration of 31.0 mg L(-1), which was 2.4-fold greater than that achieved under nitrogen-depleted conditions. High performance liquid chromatography (HPLC) analysis showed that the accumulated carbohydrate in I. zhangjiangensis was composed of glucose. These results show that I. zhangjiangensis represents an ideal carbohydrate-enriched bioresource for biofuel production.

Evaluation of the integrated hydrothermal carbonization-algal cultivation process for enhanced nitrogen utilization in Arthrospira platensis production

Sustainable microalgal cultivation at commercial scale requires nitrogen recycling. This study applied hydrothermal carbonization to recover N of hot-water extracted Arthrospira platensis biomass residue into aqueous phase (AP) under different operation conditions and evaluated the N utilization, biomass yield and quality of A. platensis cultures using AP as the sole N source. With the increase of temperature at 190-210°C or reaction time of 2-3h, the N recovery rate decreased under nitrogen-repletion (+N) cultivation, while contrarily increased under nitrogen-limitation (-N) cultivation. Under +N biomass accumulation in the cultures with AP under 190°C was enhanced by 41-67% compared with that in NaNO3, and the highest protein content of 51.5%DW achieved under 200°C-2h was also 22% higher. Carbohydrate content of 71.4%DW under -N cultivation achieved under 210°C-3h was 14% higher than that in NaNO3. HTC-algal cultivation strategy under -N mode could save 60% of conventional N.

Utilization of recovered nitrogen from hydrothermal carbonization process by Arthrospira platensis.

In the context of sustainable cultivation of microalgae, the present study focused on the use of nitrogen from the hot-water extracted biomass residue of Arthrospira platensis by hydrothermal carbonization (HTC) and the sequential cultivation of the same alga with the HTC aqueous phase (AP). Nearly 90% of the nitrogen recovered from HTC into AP was in the organic form. Under nitrogen-limited condition with HTCAP as nitrogen source the yield and content of carbohydrate were enhanced by 21% and 15% respectively compared with that under the same nitrogen level provided by NaNO3, which entitled HTCAP for the substitution of conventional nitrate. In the same way pilot-scale cultivation of A. platensis in raceway ponds outdoors demonstrated that carbohydrate content of 43.8% DW and productivity of 10.3g/m(2)/d was achieved. Notably 54% of organic nitrogen in the HTCAP could be recycled by cultivation of pre-nitrogen starved A. platensis as seeds under nitrogen limitation.

Protein–protein interaction network of the marine microalga Tetraselmis subcordiformis: prediction and application for starch metabolism analysis

Under stressful conditions, the non-model marine microalga Tetraselmis subcordiformis can accumulate a substantial amount of starch, making it a potential feedstock for the production of fuel ethanol. Investigating the interactions of the enzymes and the regulatory factors involved in starch metabolism will provide potential genetic manipulation targets for optimising the starch productivity of T. subcordiformis. For this reason, the proteome of T. subcordiformis was utilised to predict the first protein–protein interaction (PPI) network for this marine alga based on orthologous interactions, mainly from the general PPI repositories. Different methods were introduced to evaluate the credibility of the predicted interactome, including the confidence value of each PPI pair and Pfam-based and subcellular location-based enrichment analysis. Functional subnetworks analysis suggested that the two enzymes involved in starch metabolism, starch phosphorylase and trehalose-phosphate synthase may be the potential ideal genetic engineering targets.

Coordinated regulation of nitrogen supply mode and initial cell density for energy storage compounds production with economized nitrogen utilization in a marine microalga Isochrysis zhangjiangensis

Lipids and carbohydrates are main energy storage compounds (ESC) of microalgae under stressed conditions and they are potential feedstock for biofuel production. Yet, the sustainable and commercially successful production of ESC in microalgae needs to consider nitrogen utilization efficiency. Here the impact of different initial cell densities (ICDs) on ESC accumulation in Isochrysis zhangjiangensis under two nitrogen supply modes (an initially equal concentration of nitrogen per-cell in the medium (N1) and an equal total concentration of nitrogen in the culture system (N2)) were investigated. The results demonstrated that the highest ESC yield (1.36gL(-1)) at N1, which included a maximal nitrogen supply in the cultivation system, and the highest ESC content (66.5%) and ESC productivity per mass of nitrogen (3.28gg(-1) (N) day(-1)) at N2, were all obtained under a high ICD of 8.0×10(6)cellsmL(-1). Therefore I. zhangjiangensis qualifies for ESC-enriched biomass production with economized nitrogen utilization.

Investigating Cellular Responses During Photohydrogen Production by the Marine Microalga Tetraselmis subcordiformis by Quantitative Proteome Analysis

The marine microalga Tetraselmis subcordiformis could photoproduce hydrogen under the regulation of carbonyl cyanide m-chlorophenylhydrazone (CCCP), and a hydrogen production process kinetic analysis was characterized by two peaks, suggesting that two distinct mechanisms might exist in this alga. Therefore, 2D nanoliquid chromatography−tandem mass spectrometry (LC-MS/MS) was introduced to analyze the proteome of samples from different time points. A total of 912 proteins were identified, providing a global view of the cellular responses at the proteomic level. These proteins can be divided into multiple functional groups including stress responses, energy metabolism and redox homeostasis. The quantitative proteomic data provided more details on the electron donors for hydrogen production. During the first stage, photosystem II produced electrons for hydrogen production; during the second stage, metabolites were the major electron donors via nonphotochemical plastoquinone reduction by NADH dehydrogenase.