| Mortierella alpina has the characteristics of high lipid accumulation and with abundantPUFAs, especially ARA, and has been used for industrialization production of ARA. Nitrogenlimitation has been a common strategy to initiate lipid accumulation. At present, the researchconcerning effect of nitrogen sources on the lipid accumulation in M. alpina mainly focusedon the screening of nitrogen sources, but little on its regulation mechanisms. What’s more, thescreening of nitrogen sources was carried out in shake flask, and neglected the effect of pHchange on lipid synthesis in culture. This work mainly studied the regulation of lipidaccumulation by pH and nitrogen sources of the growth media, and determined the keyenzymes’ activities during lipid accumulation process of the fungus. And the possiblemechanism of this regulation was analyzed.Firstly, when M. alpina was cultivated at different nitrogen sources at the sameconcentration in shake flasks for5days, the total fatty acid content and the changes of pH inmedium were analyzed, the results showed that the higher pH environment caused bypotassium nitrate or soy peptone as the nitrogen source could promote the lipid accumulation,while the lower pH environment by using ammonium tartrate or ammonium sulfate asnitrogen source in culture inhibited the lipid accumulation. Therefore, it is suggested that pHmay be the main reason for the difference of lipid production in M. alpina cultivated atdifferent nitrogen sources in shake flask. Based on this result, ammonium tartrate andpotassium nitrate as examples of low pH and high pH nitrogen sources were used to study theeffect of pH lipid accumulation in M. alpina.Secondly, M. alpina was cultivated at different pH condition in fermenter withammonium tartrate and potassium nitrate as nitrogen source, respectively. Lipid analysis ofthe culture showed that the total fatty acid content of the fungus with ammonium tartrate andpotassium nitrate as nitrogen source was42.19%and34.93%, respectively, under the constantpH value of6.2. When the pH of the media was adjusted to5.4with both ammonium tartrateand potassium nitrate as nitrogen sources after the nitrogen sources depleted, the total fattyacid content was27.47%and30.91%, respectively; While the pH adjusted to4.2, the totalfatty acid content was14.84%and25.63%, respectively. This suggested that the total fattyacid content accumulated by M. alpina would decreased when the pH of the media wasreduced. Therefore, the pH in the culture medium had significant influence on lipidaccumulation in M. alpina, and the higher pH could promote lipid accumulation, and the pHof the media may counteract the impact of nitrogen on lipid accumulation in the fungus.Finally, activity assays of the key enzymes related to lipid synthesis of M. alpina inshake flask and fermenter showed that the changes of total fatty acid content and enzymeactivity of G-6-PDH, ACL, FAS showed a good correlation, this indicated that G-6-PDH,ACL and FAS are the key enzymes involved lipid accumulation in M. alpina. Furthermore,compared to ammonium tartrate, potassium nitrate upregulated the activities of G-6-PDH andACL during the lipid accumulation; while the pH of the media increased could upregulate theactivities of G-6-PDH and FAS during lipid accumulation. Therefore, when M. alpina cultivated in shake flask, the lipid synthesis was regulated by the combination of nitrogensource and pH. Similarly in fermenter, when M. alpina was cultivated at the appropriate pH,nitrogen source was the main regulating factor for lipid synthesis. Therefore, in addition to theregulation by nitrogen source, the pH was firstly proposed as the regulating factor for lipidaccumulation in M. alpina. |
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