CN108753622A - 一种基于糖蜜废醪液金属离子提高单针藻油脂含量的方法 - Google Patents
一种基于糖蜜废醪液金属离子提高单针藻油脂含量的方法 Download PDFInfo
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Abstract
本发明涉及一种基于糖蜜废醪液金属离子提高单针藻油脂含量的方法,属于生物柴油技术领域。本发明所述方法为将糖蜜废醪液稀释,作为单针藻的诱导培养基,然后加入MgSO4•7H2O,调节pH值在6.8‑7.0之间,分装、灭菌;将利用兼养BG‑11培养好的单针藻种子接种到上述培养基,进行光照摇瓶培养;每天离心收集藻细胞,冻干后,研磨,然后利用有机溶剂提取单针藻内的油脂,制备得到生物柴油。本发明操作简单易行、降低生产成本,能利用糖蜜废醪液提高油脂含量,同时也能减少糖蜜废醪液处理过程中的环境污染。
Description
技术领域
本发明涉及一种基于糖蜜废醪液金属离子提高单针藻油脂含量的方法,属于生物柴油技术领域。
背景技术
生物柴油提炼自动植物油,它是指以油料作物如大豆、油菜等,野生油料植物和工程微藻等水生植物油脂以及动物油脂、餐饮垃圾油等为原料油通过酯交换或热化学工艺制成的可代替石化柴油的再生性柴油燃料。生物柴油是生物质能的一种,其在物理性质上与石化柴油接近,但化学组成不同。
利用微藻制备生物柴油的传统方法,不仅生产成本高,油脂含量和油脂产率低,而且很难应用于大规模生产,本发明在微藻具有光合效率高,生长周期短等优势的基础上改变了传统的微藻培养方式,利用糖蜜废醪液作为基础培养基,既可以解决糖蜜废醪液污水处理的问题,又可以利用糖蜜废醪液中的营养成分培养微藻,同时又能提高微藻中油脂的积累。而在微藻中添加植物激素褪黑素或胺酰酯后,可进一步促进微藻油脂积累。
发明内容
本发明的目的在于提供一种基于糖蜜废醪液金属离子提高单针藻油脂含量的方法,方法过程简单,绿色环保生产成本低,具体包括以下步骤:
(1)制备诱导培养基:将糖蜜废醪液稀释,作为单针藻的诱导培养基,然后加入MgSO4•7H2O,调节pH值在6.8-7.0之间,分装、灭菌;
(2)诱导藻细胞积累油脂:将利用兼养BG-11培养好的单针藻种子接种到上述培养基,进行光照摇瓶培养;
(3)制备生物柴油:每天离心收集藻细胞,冻干后,研磨,然后利用有机溶剂提取单针藻内的油脂,制备得到生物柴油。
优选的,本发明步骤(1)所述培养基中Mg2+的浓度为400~3200μmol/L。
优选的,本发明步骤(1)糖蜜废醪液稀释3200倍。
优选的,本发明所述步骤(2)中培养条件为:温度25±1℃,光照强度3500lux,摇床转速150 r/min。
优选的,本发明所述步骤(3)中制备生物柴油的方法具体为:将藻细胞培养液离心收集,条件为3500 r/min,离心10 min,并以蒸馏水反复洗涤两次后离心冻干;添加冻干藻粉2倍质量石英砂研磨,然后用氯仿与甲醇体积比为2:1的氯仿/甲醇提取油脂,重复提取2次后收集提取液浓缩,提取的油脂添加硫酸-甲醇进行甲酯化制备得到生物柴油,硫酸与甲醇的体积比为3:97。
本发明的有益效果是:
(1)本发明利用Mg2+离子促进了单针藻油脂含量的增加,降低了生产成本;糖蜜废醪液实现零排放,有利于保护环境,将环境污染处理和微藻的培养结合在一起,变废为宝;本发明所述方法与对照组相比,油脂含量达到了50.74%。
(2)本发明中单针藻能很好地在加了MgSO4•7H2O的糖蜜废醪液培养基中生长,不但解决了糖蜜废醪液的处理问题,同时提高了油脂含量,可能存在的作用机制是,镁离子高效促进油脂含量的升高。
附图说明
图1为不同Fe3+、Zn2+、Mg2+浓度条件下油脂含量的变化图。
具体实施方式
结合附图和具体实施例对本发明作进一步详细说明,但本发明保护范围并不限于所述内容。
对比例1
以单针藻Monoraphidium sp. FXY-10为研究对象,利用BG-11培养基,添加葡萄糖(10g/L)培养至稳定期;将糖蜜废醪液稀释3200倍,作为培养单针藻的诱导培养基,调节pH值在6.8-7.0之间;高温高压121 ℃灭菌20 min,接入单针藻种子液,藻细胞初始接种量为1 g/L,培养温度25 ℃,光照强度3500 lux,摇床转速150 r/min,进行光照摇瓶培养。
结果:该对照组藻细胞的最大油脂含量为43.63%。
对比例2
(1)制备诱导培养基:将糖蜜废醪液稀释3200倍,作为单针藻Monoraphidium sp. FXY-10的诱导培养基,然后加入柠檬酸铁铵,调节pH值在6.8-7.0之间,分装、灭菌条件为121℃、20 min;培养基中柠檬酸铁铵添加量分别为4.82 mg/L、10.062 mg/L、20.541 mg/L、41.500mg/L;得到含有Fe3+浓度分别为20μmol/L、40μmol/L、80μmol/L、160μmol/L。
(2)诱导藻细胞积累油脂:将利用兼养BG-11培养好的单针藻种子接种到上述培养基,进行光照摇瓶培养;培养条件为:温度25±1℃,光照强度3500lux,摇床转速150r/min。
(3)制备生物柴油:每天离心收集藻细胞,将藻细胞培养液离心收集,条件为3500r/min,离心10min,并以蒸馏水反复洗涤两次后离心冻干;添加冻干藻粉2倍质量石英砂研磨,然后用氯仿与甲醇体积比为2:1的氯仿/甲醇提取油脂,重复提取2次后收集提取液浓缩,提取的油脂添加硫酸-甲醇进行甲酯化制备得到生物柴油,硫酸与甲醇的体积比为3:97。
表1不同Fe3+浓度条件下油脂含量
对比例3
(1)制备诱导培养基:将糖蜜废醪液稀释3200倍,作为单针藻Monoraphidium sp. FXY-10的诱导培养基,然后加入ZnSO4•7H2O,调节pH值在6.8-7.0之间,分装、灭菌,灭菌条件为121℃,20 min;ZnSO4•7H2O添加量为0.215mg/L、0.502mg/L、1.077 mg/L、2.228mg/L得到含有Zn2+浓度分别为1μmol/L、2μmol/L、4μmol/L、8μmol/L。
(2)诱导藻细胞积累油脂:将利用兼养BG-11培养好的单针藻种子接种到上述培养基,进行光照摇瓶培养;培养条件为:温度25±1℃,光照强度3500 lux,摇床转速150 r/min。
(3)制备生物柴油:每天离心收集藻细胞,将藻细胞培养液离心收集,条件为3500r/min,离心10min,并以蒸馏水反复洗涤两次后离心冻干;添加冻干藻粉2倍质量石英砂研磨,然后用氯仿与甲醇体积比为2:1的氯仿/甲醇提取油脂,重复提取2次后收集提取液浓缩,提取的油脂添加硫酸-甲醇进行甲酯化制备得到生物柴油,硫酸与甲醇的体积比为3:97。
表2不同Zn2+浓度条件下油脂含量
实施例1
(1)制备诱导培养基:将糖蜜废醪液稀释3200倍,作为单针藻Monoraphidium sp. FXY-10的诱导培养基,然后加入MgSO4•7H2O,调节pH值在6.8-7.0之间,分装、灭菌、灭菌条件为121℃,20min; MgSO4•7H2O添加量分别为91mg/L、189.4mg/L、287.8 mg/L、385.2mg/L,得到含有Mg2+浓度分别为400μmol/L、800μmol/L、1600μmol/L、3200μmol/L的诱导培养基。
(2)诱导藻细胞积累油脂:将利用兼养BG-11培养好的单针藻种子接种到上述培养基,进行光照摇瓶培养;培养条件为:温度25±1℃,光照强度3500lux,摇床转速150r/min。
(3)制备生物柴油:每天离心收集藻细胞,将藻细胞培养液离心收集,条件为3500r/min,离心10min,并以蒸馏水反复洗涤两次后离心冻干;添加冻干藻粉2倍质量石英砂研磨,然后用氯仿与甲醇体积比为2:1的氯仿/甲醇提取油脂,重复提取2次后收集提取液浓缩,提取的油脂添加硫酸-甲醇进行甲酯化制备得到生物柴油,硫酸与甲醇的体积比为3:97。
表3不同Mg2+浓度条件下油脂含量
结果表明:以糖蜜废醪液为基础培养基诱导单针藻积累油脂,当Mg2+浓度为1600μmol/L(即MgSO4•7H2O添加量为287.8mg/L)时,单针藻的油脂含量比对照组提高了16.32%;说明在糖蜜废醪液中添加一定量镁离子有助于提高单针藻的油脂积累,对单针藻油脂积累有一定的促进作用。由对比例也可以看出,并不是每一种金属离子都可以提高微藻的生物量、油脂含量及油脂含量。添加Fe3+时,单针藻的油脂含量比对照组差不多,当Zn2+浓度为1μmol/L(即ZnSO4•7H2O添加量为0.215 mg/L)时,单针藻油脂含量没有提高,且下降了13.02%。
Claims (5)
1.一种基于糖蜜废醪液金属离子提高单针藻油脂含量的方法,其特征在于,具体包括以下步骤:
(1)制备诱导培养基:将糖蜜废醪液稀释,作为单针藻的诱导培养基,然后加入MgSO4•7H2O,调节pH值在6.8-7.0之间,分装、灭菌;
(2)诱导藻细胞积累油脂:将利用兼养BG-11培养好的单针藻种子接种到上述培养基,进行光照摇瓶培养;
(3)制备生物柴油:每天离心收集藻细胞,冻干后,研磨,然后利用有机溶剂提取单针藻内的油脂,制备得到生物柴油。
2.根据权利要求1所述基于糖蜜废醪液金属离子提高单针藻油脂含量的方法,其特征在于:步骤(1)所述培养基中Mg2+的浓度为400~3200μmol/L。
3.根据权利要求1所述基于糖蜜废醪液金属离子提高单针藻油脂含量的方法,其特征在于:步骤(1)糖蜜废醪液稀释3200倍。
4.根据权利要求1所述基于糖蜜废醪液金属离子提高单针藻油脂含量的方法,其特征在于:所述步骤(2)中培养条件为:温度25±1℃,光照强度3500lux,摇床转速150 r/min。
5.根据权利要求1所述基于糖蜜废醪液金属离子提高单针藻油脂含量的方法,其特征在于:所述步骤(3)中制备生物柴油的方法具体为:将藻细胞培养液离心收集,条件为3500r/min,离心10 min,并以蒸馏水反复洗涤两次后离心冻干;添加冻干藻粉2倍质量石英砂研磨,然后用氯仿与甲醇体积比为2:1的氯仿/甲醇提取油脂,重复提取2次后收集提取液浓缩,提取的油脂添加硫酸-甲醇进行甲酯化制备得到生物柴油,硫酸与甲醇的体积比为3:97。
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