CN115011643A - 一种利用氨基葡萄糖强化微藻固碳与产油的方法 - Google Patents
一种利用氨基葡萄糖强化微藻固碳与产油的方法 Download PDFInfo
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Abstract
本发明公开了一种利用氨基葡萄糖强化微藻固碳与产油的方法,包括以下步骤:向微藻培养体系中通入CO2并添加氨基葡萄糖以强化CO2气体溶解与传质和胁迫微藻产油;通过投加氨基葡萄糖,首先提高藻液储存二氧化碳的能力,其次加快二氧化碳从空气中溶于藻液的传质过程,有助于提高微藻的光合作用,从而加快微藻生物量的增长;另外,通过氨基葡萄糖对微藻细胞的胁迫,可以促进产油代谢的进行,从而提高细胞含油量。通过生物量和细胞含油量两方面的促进作用,提高微藻油脂的最终产量(油脂产量=生物量×细胞含油率)。
Description
技术领域
本发明涉及微藻生物技术领域,具体涉及一种利用氨基葡萄糖强化微藻固碳与产油的方法。
背景技术
通常情况下,微藻在全营养条件下培养可以获得比较快的生长速率,但是含油量并不高。而大量的研究表明不利的生长环境因子可以提高微藻脂质含量,诸如氮缺乏、硅不足、磷限制、重金属胁迫、高光和高温胁迫、激素作用等等,其中尤以氮缺乏的影响最为显著,并被认为是最有效的刺激油脂含量提高的方法。然而,研究者们发现氮缺乏刺激脂质含量提高的同时,往往伴随着细胞生长的停滞和生物量的减少,这将导致脂质产率无法有效提高。因此,高含油量和高产油率往往难以同时获得。另外,当前微藻产油技术还存在藻液对二氧化碳的储存能力低以及二氧化碳在藻液中的传质速度慢,导致二氧化碳被微藻吸收后,不能得到及时补充的问题。
发明内容
有鉴于此,本发明的目的在于提供一种利用氨基葡萄糖强化微藻固碳与产油的方法,通过往正常培养的微藻中添加一定浓度氨基葡萄糖,同时通入二氧化碳,促进藻液储存二氧化碳的能力并加快二氧化碳从空气中溶于藻液的传质过程,促进微藻的生物量增长,同时增加单个细胞的含油率,最终实现微藻总产油量的增加。
本发明利用氨基葡萄糖强化微藻固碳与产油的方法,包括以下步骤:向微藻培养体系中通入CO2并添加氨基葡萄糖以强化CO2气体溶解与传质和胁迫微藻产油;
进一步,在微藻培养基中添加氨基葡萄糖,在培养过程中通入CO2气体;
进一步,所述氨基葡萄糖浓度为1-5mg/L;
进一步,通入CO2气体体积分数为3-15%,供气速率为10-50mL/min,曝气时间为2-12h;
进一步,所述微藻藻液初始pH为6.5-7.5,初始微藻生物量100-200mg/L;
进一步,所述微藻的培养周期14天,培养温度25-35℃,光照强度3000-6000lux,光暗比12:12;
进一步,所述微藻为海洋微藻、淡水微藻中的一种。
进一步,所述微藻为葡萄藻属、杜氏藻、小球藻属、栅藻属、莱茵衣藻、隐甲藻、筒柱藻、巴夫藻、微绿球藻、等鞭金藻、三角褐指藻、螺旋藻、布朗葡萄藻、微拟球藻中的一种;
进一步,所述微藻培养所用培养基可以为BG11培养基、TAP培养基、SE培养基中的一种。
本发明的有益效果:本发明的利用氨基葡萄糖强化微藻固碳与产油的方法,通过投加氨基葡萄糖,首先促进藻液储存二氧化碳的能力,其次加快二氧化碳从空气中溶于藻液的传质过程,有助于提高微藻的光合作用,从而加快微藻生物量的增长;另外,通过氨基葡萄糖对微藻细胞的胁迫,可以促进产油代谢的进行,从而提高细胞含油量。通过生物量和细胞含油量两方面的促进作用,提高微藻油脂的最终产量(油脂产量=生物量×细胞含油率)。
具体实施方式
为更好的理解本发明,下面的实施例是对本发明的进一步说明,但本发明的内容不仅仅局限于下面的实施例。
实施例中,所使用的实验方法如无特殊说明,均为常规方法,所用的材料、试剂等,如无特殊说明,均可从商业途径得到。
本实施例的利用氨基葡萄糖强化微藻固碳与产油的方法,包括以下步骤:向微藻培养体系中通入CO2并添加氨基葡萄糖以强化CO2气体溶解与传质和胁迫微藻产油;在微藻的培养期中添加一种主要药剂氨基葡萄糖,并通入CO2作为碳源,通过促进碳源供给和改变微藻产油代谢这两方面促进微藻产油,与现有的产油方式不同,且氨基葡萄糖可促进二氧化碳在藻液中的储存及传质,从而大幅提高微藻生物量;氨基葡萄糖可通过胁迫微藻相关代谢,增加微藻的细胞含油率。所采用的氨基葡萄糖成本低、无二次污染,既节约经济成本又环保。CO2气体为可以为工业排放的含有大量CO2的废气,也可以为纯二氧化碳,均可达到发明目的。
本实施例中,在微藻培养基中添加氨基葡萄糖,在培养过程中通入CO2气体。“在微藻培养基中添加氨基葡萄糖”包括两种方式:1、先在未接种微藻的培养基中添加氨基葡萄糖,然后接种微藻;2、在接种了微藻的培养基中加入氨基葡萄糖。上述两种方式均能实现发明目的,而优选采用第一种方式。CO2气体作为碳源供给,促进藻液储存二氧化碳的能力,其次加快二氧化碳从空气中溶于藻液的传质过程。
本实施例中,所述氨基葡萄糖浓度为1-5mg/L;氨基葡萄糖规格可为低粘度或者中粘度。
本实施例中,通入CO2气体体积分数为3-15%,供气速率为10-50mL/min,曝气时间为2-12h;所述微藻藻液初始pH为6.5-7.5,初始微藻生物量100-200mg/L;所述微藻的培养周期14天,培养温度25-35℃,光照强度3000-6000lux,光暗比12:12。
本实施例中,所述微藻为海洋微藻、淡水微藻中的一种,包括葡萄藻属、杜氏藻、小球藻属、栅藻属、莱茵衣藻、隐甲藻、筒柱藻、巴夫藻、微绿球藻、等鞭金藻、三角褐指藻、螺旋藻、布朗葡萄藻、微拟球藻。
本实施例中,所述微藻培养所用培养基可以为BG11培养基、TAP培养基、SE培养基中的一种。
实施例一
本实施例的利用氨基葡萄糖强化微藻固碳与产油的方法,包括以下步骤:在SE培养基中添加氨基葡萄糖1mg/L,然后将处于对数生长期的栅藻按照初始接种浓度约为100mg/L接种于富含营养的SE培养基制得微藻液,微藻液初始pH值为6.5,培养温度为25℃,光照强度为3000Lux,光照周期为12h:12h(光:暗),在微藻液培养过程中通入3%CO2,供气速率为20mL/min,并采用曝气石进行曝气,持续2h,培养周期14天,培养过程中每天测定栅藻生物量。
实施例二
本实施例的利用氨基葡萄糖强化微藻固碳与产油的方法,包括以下步骤:在BG11培养基中添加氨基葡萄糖5mg/L,然后将处于对数生长期的栅藻按照初始接种浓度为100mg/L接种于富含营养的BG11培养基制得微藻液,微藻液初始pH值为7.5,培养温度为35℃,光照强度为4000Lux,光照周期为12h:12h(光:暗),在微藻液培养过程中通入通入15%CO2,供气速率为10mL/min并采用曝气石进行曝气,持续12h,培养周期14天,培养过程中每天测定栅藻生物量。
实施例三
本实施例的利用氨基葡萄糖强化微藻固碳与产油的方法,包括以下步骤:在SE培养基中添加氨基葡萄糖3.5mg/L,然后将处于对数生长期的小球藻按照初始接种浓度为200mg/L接种于富含营养的SE培养基制得微藻液,微藻液初始pH值为7.5,培养温度为35℃,光照强度为6000lux,光照周期为12h:12h(光:暗),在微藻液培养过程中通入15%CO2,供气速率为50mL/min并采用曝气石进行曝气,持续12h,培养周期14天,培养过程中每天测定微小球藻生物量。
实施例四
本实施例的利用氨基葡萄糖强化微藻固碳与产油的方法,包括以下步骤:在TAP培养基中添加氨基葡萄糖2mg/L,然后将处于对数生长期的小球藻按照初始接种浓度为150mg/L接种于富含营养的TAP培养基制得微藻液,微藻液初始pH值为7,培养温度为30℃,光照强度为4000lux,光照周期为12h:12h(光:暗),在微藻液培养过程中通入5%CO2,供气速率为40mL/min,并采用曝气石进行曝气,持续12h,培养周期14天,培养过程中每天测定小球藻生物量。
实施例五
本实施例的利用氨基葡萄糖强化微藻固碳与产油的方法,包括以下步骤:在SE培养基中添加氨基葡萄糖4mg/L,然后将处于对数生长期的莱茵衣藻按照初始接种浓度为120mg/L接种于富含营养的SE培养基制得微藻液,微藻液初始pH值为7,培养温度为28℃,光照强度为5000lux,光照周期为12h:12h(光:暗),在微藻液培养过程中通入10%CO2,供气速率为35mL/min并采用曝气石进行曝气,持续8h,培养周期14天,培养过程中每天测定莱茵衣藻生物量。
实施例六
本实施例的利用氨基葡萄糖强化微藻固碳与产油的方法,包括以下步骤:在SE培养基中添加氨基葡萄糖1mg/L,然后将处于对数生长期的莱茵衣藻按照初始接种浓度为180mg/L接种于富含营养的SE培养基制得微藻液,微藻液初始pH值为7.5,培养温度为30℃,光照强度为5500lux,光照周期为12h:12h(光:暗),在微藻液培养过程中通入2%CO2,供气速率为45mL/min并采用曝气石进行曝气,持续5h,培养周期14天,培养过程中每天测定莱茵衣藻生物量。
上述实施例中,所采用的设备为光生物反应器。
本发明的通过壳聚糖强化微藻产油与固碳的方法,也可以采用二乙醇胺、壳聚糖也能起到一定的强化微藻产油与固碳的目的,但是添加二乙醇胺相较于壳聚糖促进微藻生长存在较大差异,添加二乙醇胺的促进微藻生长远不如壳聚糖,但是在促进细胞油脂含量上二乙醇胺明显优于壳聚糖,而氨基葡萄糖相比于壳聚糖,促进微藻生长明显优于壳聚糖,在促进细胞油脂含量方面略逊于壳聚糖。壳聚糖、氨基葡萄糖、二乙醇胺者三种物质,总体来说,随着投量的增加,促进微藻生长的效果先增加,后减少,加太多的话还会抑制生长但是促进单个细胞含油率方面,是随着剂量的增加而增加的,当然在整个培养过程中,细胞含油率和微藻生长还受其他培养条件因素影响。
对比例1:不添加氨基葡萄糖,其他步骤与实施例一相同。
对比例2:仅将氨基葡萄糖直接替换为壳聚糖,其余与实施例一相同。
对比例3:仅将氨基葡萄糖直接替换为二乙醇胺,其余与实施例一相同。
数据测定
在培养总天数14天后分别对实施例1-6和对比例1-3中微藻的生物量、油含量进行评定:
生物量测定方法:采用干重法测定生物量,通过离心收集藻液中的藻生物质,并将收集的藻生物质置于105℃烘箱中直至烘干,然后放置干燥器中冷却至室温,称重后计算前后重量差即可得到生物量的大小。
脂肪酸甲酯(FAMEs)的量作为含油量:使用简单而快速的FAMEs分析方法。首先脂质经甲酯化后才能变成脂肪酸甲酯。取20mg干藻粉加入体积为10mL的有盖玻璃管,加入新鲜配制的2mL甲酯化试剂(乙酰氯/甲醇,乙酰氯与
甲醇的体积比为1:9),加上盖子并拧紧后,置于80℃的水浴中反应2.5h。待反应完成后,冷却至室温并加入1mL 1%NaCl水溶液和2mL含有苯甲酸甲酯的正己烷(大约0.36mg/mL正己烷),苯甲酸甲酯为内标。经震荡离心后分层,含有FAMEs的上层回收至GC样品管,经无水硫酸钠干燥后测定。FAMEs的总量以及组成使用气相色谱Agilent 6890测定,并使用火焰离子化检测器以及DB-FFAP型号的毛细管柱(30m×0.25mm×0.25μm)。气相色谱运行,条件如下:进样量1μL;分流比1:10;空气流速450mL/min,H 240mL/min,载气(N2)45mL/min;进样口温度250℃;检测器温度300℃;起始炉温为140℃并保持2min,随后以10℃/min的速度升高至240℃,并在240℃时保持2min。通过样品中各脂肪酸甲酯的峰面积和内标峰面积计算FAMEs的含量。
测定数据如下表:
最后说明的是,以上实施例仅用以说明本发明的技术方案而非限制,尽管参照较佳实施例对本发明进行了详细说明,本领域的普通技术人员应当理解,可以对本发明的技术方案进行修改或者等同替换,而不脱离本发明技术方案的宗旨和范围,其均应涵盖在本发明的权利要求范围当中。
Claims (9)
1.一种利用氨基葡萄糖强化微藻固碳与产油的方法,其特征在于:包括以下步骤:向微藻培养体系中通入CO2并添加氨基葡萄糖以强化CO2气体溶解与传质和胁迫微藻产油。
2.根据权利要求1所述的利用氨基葡萄糖强化微藻固碳与产油的方法,其特征在于:在微藻培养基中添加氨基葡萄糖,在培养过程中通入CO2气体。
3.根据权利要求2所述的利用氨基葡萄糖强化微藻固碳与产油的方法,其特征在于:所述氨基葡萄糖浓度为1-5mg/L。
4.根据权利要求3所述的利用氨基葡萄糖强化微藻固碳与产油的方法,其特征在于:通入CO2气体体积分数为3-15%,供气速率为10-50mL/min,曝气时间为2-12h。
5.根据权利要求1所述的利用氨基葡萄糖强化微藻固碳与产油的方法,其特征在于:所述微藻藻液初始pH为6.5-7.5,初始微藻生物量100-200mg/L。
6.根据权利要求5所述的利用氨基葡萄糖强化微藻固碳与产油的方法,其特征在于:所述微藻的培养周期14天,培养温度25-35℃,光照强度3000-6000lux,光暗比12:12。
7.根据权利要求6所述的利用氨基葡萄糖强化微藻固碳与产油的方法,其特征在于:所述微藻为海洋微藻、淡水微藻中的一种。
8.根据权利要求7所述的利用氨基葡萄糖强化微藻固碳与产油的方法,其特征在于:所述微藻为葡萄藻属、杜氏藻、小球藻属、栅藻属、莱茵衣藻、隐甲藻、筒柱藻、巴夫藻、微绿球藻、等鞭金藻、三角褐指藻、螺旋藻、布朗葡萄藻、微拟球藻中的一种。
9.根据权利要求8所述的利用氨基葡萄糖强化微藻固碳与产油的方法,其特征在于:所述微藻培养所用培养基可以为BG11培养基、TAP培养基、SE培养基中的一种。
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