| As people's needs for energy continues to increase,there is extensive attention on the biodiesel with clean energy and low pollution.Microalgae were considered to be extremely valuable due to their multiple advantages,especially the storage of rich lipids.Therefore,they are considered to become a potential material for biodiesel.At present,the lipid content of potential rich lipid microalgae can be improved by screening and many other control measures?such as the limitation of nutrient elements and the regulation of environmental factors and trophic modes?.Shanxi Province is now in the stage of energy restructuring,the development of new renewable energy is imminent,and microalgae are good choices.Based on the above research progress,a native oil-rich algal strain Parachlorella kessleri TY of Shanxi Province was screened in this study.However,there are few reports on this algal species in the literature,especially the related studies on the lipid of algal.A native alga strain from Shanxi Province was taken as the object of this study and explored the nitrogen limiting and trophic modes regulating on its lipid accumulation.The main research results are as follows:?1?In this study,a green alga with single-celled was first screened from the soil of Shanxi Province,then it was identified by morphology and phylogeny,the results showed that they became to P.kessleri,and named P.kessleri TY.At the same time,its lipid-related characteristics were analyzed,the results showed that it's total lipid content reached to 33.27% and the lipid productivity reached to 34.62 mg·L-1d-1,the lipid composition was mainly composed of palmitic acid?C16:0?,linoleic acid?C18:2?and linolenic acid?C18:3?.The total fatty acids of C16 and C18 series were also high,up to 88.15%,and met the relevant standards for biodiesel.So,it was a potential oilproducing alga that could as the raw material for biodiesel,it could be seen that this algae strain has great potential to be used as the raw material of biodiesel.?2?It was found that the growth rate,chloroplast degradation,photosynthetic activity and protein synthesis of P.kessleri TY were decreased under the condition of nitrogen deprivation.However,the content of total carbohydrate content,neutral lipids and total lipids in algal cells were increased,and the content of C16:0,C18:1,C18:2 and C18:3 increased significantly which are suitable for biodiesel.The content of saturated fatty acids?SFA?and monounsaturated fatty acids?MUFA?also increased.The result of transcriptome sequencing showed that the nitrogen deprivation condition was compared with the nitrogen normal condition,the key ratelimiting enzyme related genes?acc C,fab F,fab H,FAD2,SCD?involved in the pathways of fatty acid biosynthesis and fatty acid unsaturated biosynthesis were showed up-regulated significantly.In addition,the key rate-limiting enzyme related genes?ACAA1,ECHS1,MECR?involved in the fatty acid elongation and fatty acid degradation were presented significantly downregulated.Based on physiological,biochemical and transcriptome results,it was not difficult to find that nitrogen deprivation was beneficial to lipid accumulation of P.kessleri TY.?3?P.kessleri TY was cultured under heterotrophic/mixotrophic with glucose as organic carbon source.The result displayed that,the biomass dry weight of the algae strain under the heterotrophic culture was 0.55 g·L-1,the biomass productivity was 78.57 mg·L-1d-1,but the biomass dry weight of the algae strain under the mixotrophic reached 1.53 g·L-1,and its biomass productivity was as high as 218.57 mg·L-1d-1.However,the pigment contents,photosynthetic activity and protein content of P.kessleri TY under the heterotrophic/mixotrophic were significantly lower than the autotrophic culture,indicating that the medium supplemented with glucose was not conducive to the synthesis of pigments,it would seriously inhibit the photosynthetic activity and the protein content.While the algae under the heterotrophic/mixotrophic could accumulate the lipids?total lipid contents were 39.85% and 42.92%,respectively?,especially the neutral lipids.And the algae under the heterotrophic/mixotrophic was suitable for increased in the fatty acid contents as the raw material for biodiesel,especially the content of C18:1 was significantly enhanced.What's more,due to the lower biomass of heterotrophic culture compared with that of mixotrophic culture,the total lipid productivity of heterotrophic culture decreased.On the basis of the above results,we believed that the mixotrophic culture was more conducive to the lipid accumulation of P.kessleri TY.?4?The nitrogen deprivation and mixotrophic culture combined was carried out in order to obtain higher lipid productivity.The results demonstrated that the biomass dry weight of P.kessleri TY under the culture combined with nitrogen deprivation and mixotrophic was 1.15 g·L-1,the biomass productivity was 164.29 mg·L-1d-1,pigment contents and chlorophyll fluorescence were higher than nitrogen deprivation culture and lower than mixotrophic culture,it was indicated that under nitrogen deprivation condition was not conducive to the growth,pigment contents and photosynthetic activity of algae strain.However,its lipid accumulation ability was obviously better than that in nitrogen deprivation culture and mixotrophic culture.The above results indicated that the lipid accumulation of the algal strain in the combined cultivation condition increased significantly due to both nitrogen deprivation culture and mixotrophic culture had positive influences on lipid accumulation.In addition,its carbohydrate content and protein content were significantly lower than those of the other two groups,indicating that the combination of nitrogen deprivation and the addition of carbon sources would greatly reduce its protein content,while most of the newly assimilated carbon sources would be transferred to lipid,thus could facilitate the lipid accumulation.Moreover,the increase in lipid content of P.kessleri TY was much greater than the decrease in biomass,so the lipid productivity was also higher.And fatty acid composition analysis showed that the total fatty acids of C16 and C18,C16:0,C18:1,the contents of SFA and MUFA were also higher in the nitrogen deprivation and mixotrophic combined culture.Therefore,nitrogen deprivation and mixotrophic combined culture can improve the lipid accumulation greatly.In conclusion,the combined cultivation of P.kessleri TY under nitrogen deprivation and mixotrophic culture conditions could maximize the lipid productivity,and get a relatively high content of major fatty acids that could be used in biodiesel production.In the meanwhile,the relatively high content of the main fatty acids was obtained.Therefore,it is of great significance to study microalgae as raw materials of biodiesel. |
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