多釜串联生物反应器系统中自絮凝颗粒酵母乙醇连续发酵的研究
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摘要
模拟现有酒精发酵的工艺流程和发酵罐结构，建立了一套多釜串联反应器系统(TIR)，以脱胚脱皮玉米粉双酶法制备的糖化液为种子培养基和发酵底物，进行了絮凝颗粒酵母SPSC01乙醇连续发酵的实验研究。
在配置种子罐的条件下，由三级发酵罐串联和一个种子培养罐组成的组合反应器系统，种子罐培养基还原糖浓度为100g／L，添加(NH_4)_2HPO_4和KH_2PO_4各2.0g／L，以0.017h~(-1)的恒定稀释速率流加，并溢流至后续发酵系统。发酵底物初始还原糖浓度220g／L，添加(NH_4)_2HPO_4 1.5g／L和KH_2PO_42.5g／L，流加至第一级发酵罐，稀释速率分别控制为0.017、0.025、0.033、0.040和0.050h~(-1)。实验数据表明，种子罐呈均匀悬浮的恒化状态，但自絮凝颗粒酵母在各发酵罐中呈部分固定化状态，在稀释速率0.040h~(-1)条件下，发酵系统呈一定的震荡行为，其他四个稀释速率实验组均达拟稳态。当稀释速率不超过0.033h~(-1)，流出末级发酵罐的发酵液中酒精浓度可以达到12％(v／v)以上，残还原糖和残总糖分别在0.15％和0.35％(w／v)以下，发酵罐的设备生产强度达到2.71g／(L·h)。在不配置种子罐的条件下，使用相同的培养基，平均发酵时间为25h～30h，发酵终点酒精浓度达到13％(v／v)，残还原糖和残总糖分别在0.30％和0.40％(w／v)以下，发酵罐的设备生产强度达到3.47g／(L·h)。两组工艺路线的实验数据都表明，自絮凝颗粒酵母体系完全可以适用于现有酒精发酵装置工艺流程和发酵罐设备结构，可以用于对现有酒精发酵装置进行技术改造，提升其工艺技术水平。
在此基础上，从减轻废糟液处理负荷，实现清洁生产的需要出发，进行了无种子罐条件下酒精蒸馏废液直接全循环使用的自絮凝颗粒酵母酒精连续发酵实验研究，十三次全循环运行得到的实验数据，与无废液循环的实验结果相比，虽然平均发酵时间略增至30h～40h，但发酵终点酒精浓度仍达到12.5％(v／v)以上，残还原糖和残总糖分别在0.70％和0.90％(w／v)以下，说明循环积累物对发酵的影响有限，废糟液直接全循环使用的工艺技术路线可行。
A bioreactor system composed of multi-stage tank fermentors in series was established, in which a self-flocculating yeast strain developed by protoplast fusion technology was applied to ferment corn powder two-stage enzymatic hydrolyzate to produce ethanol.
Firstly, a seed fermentor was coupled into the fermentation system composed of three tanks in series, and the hydrolyzate medium containing reducing sugar 100 g/L, added 2.0g/L (NH4)2HPO4 and KH2PO4, respectively, was fed into the seed fermentor at the dilution rate of 0.017h~' for the self-flocculating yeast seed cultivation. Meanwhile, the hydrolyzate medium containing reducing sugar 220 g/L, added 1.5g/L (NH4)2HPO4 and 2.5g/L KH2PO4) was fed into the first fermentor at the dilution rates of 0.017, 0.025, 0.033, 0.040 and 0.050 h-1, respectively. The experimental results showed that Chemostat states were developed for seed cultivation and ethanol fermentations under the dilution rates of 0.017, 0.025, 0.033 and 0.050 h-1. However, the oscillations of fermented parameters, including residual sugar, ethanol and biomass concentrations, were observed under the dilution rate of 0.040 h-1 for this continuous ethanol fermentation system. Over 12% (v/v) ethanol was achieved, at the same time, residual reducing sugar and total sugar were maintained below 0.15% and 0.35% (w/v) when the dilution rate was controlled no more than 0.033 h-1. The ethanol productivity of the entire fermentation system was calculated to be 2.71g/(L-h).
Secondly, the seed fermentor was removed from the above bioreactor syatem, and the yeast floes grown inside fermentors were used to support ethanol fermentation. Using same ferment medium, over 13% (v/v) ethanol was achieved, at the same time, residual reducing sugar and total sugar were maintained below 0.30% and 0.40% (w/v) when the dilution rate was controlled no more than 0.040 h-1. The ethanol productivity of the entire fermentation system was calculated to be 3.47g/(L-h). These experimental running indicated that this new ethanol fermentation technology using self-flocculating yeast cells can be applied to such a fermentation system composed of tanks in series that was widely used in ethanol fermentation industry in China at present and help improve its efficiency and save production cost.
Finally, the effect of the complete recycling use of distilled effluent on ethanol fermentation was examined in order to save the cost for the treatment of waste distillage and promote cleansing production. Based on the data of 13 circulations, although the average fermentation time needed to be extended to 30~40 hours, over 12.5% (v/v) ethanol was achieved, at the same time, residual reducing sugar and total sugar were maintained below 0.70% and 0.90% (w/v), which indicates that the negative effect of the completely recycling use of waste distillage was limited, and the waste distillage can be completely recycled within the fermentation system.
在配置种子罐的条件下，由三级发酵罐串联和一个种子培养罐组成的组合反应器系统，种子罐培养基还原糖浓度为100g／L，添加(NH_4)_2HPO_4和KH_2PO_4各2.0g／L，以0.017h~(-1)的恒定稀释速率流加，并溢流至后续发酵系统。发酵底物初始还原糖浓度220g／L，添加(NH_4)_2HPO_4 1.5g／L和KH_2PO_42.5g／L，流加至第一级发酵罐，稀释速率分别控制为0.017、0.025、0.033、0.040和0.050h~(-1)。实验数据表明，种子罐呈均匀悬浮的恒化状态，但自絮凝颗粒酵母在各发酵罐中呈部分固定化状态，在稀释速率0.040h~(-1)条件下，发酵系统呈一定的震荡行为，其他四个稀释速率实验组均达拟稳态。当稀释速率不超过0.033h~(-1)，流出末级发酵罐的发酵液中酒精浓度可以达到12％(v／v)以上，残还原糖和残总糖分别在0.15％和0.35％(w／v)以下，发酵罐的设备生产强度达到2.71g／(L·h)。在不配置种子罐的条件下，使用相同的培养基，平均发酵时间为25h～30h，发酵终点酒精浓度达到13％(v／v)，残还原糖和残总糖分别在0.30％和0.40％(w／v)以下，发酵罐的设备生产强度达到3.47g／(L·h)。两组工艺路线的实验数据都表明，自絮凝颗粒酵母体系完全可以适用于现有酒精发酵装置工艺流程和发酵罐设备结构，可以用于对现有酒精发酵装置进行技术改造，提升其工艺技术水平。
在此基础上，从减轻废糟液处理负荷，实现清洁生产的需要出发，进行了无种子罐条件下酒精蒸馏废液直接全循环使用的自絮凝颗粒酵母酒精连续发酵实验研究，十三次全循环运行得到的实验数据，与无废液循环的实验结果相比，虽然平均发酵时间略增至30h～40h，但发酵终点酒精浓度仍达到12.5％(v／v)以上，残还原糖和残总糖分别在0.70％和0.90％(w／v)以下，说明循环积累物对发酵的影响有限，废糟液直接全循环使用的工艺技术路线可行。
A bioreactor system composed of multi-stage tank fermentors in series was established, in which a self-flocculating yeast strain developed by protoplast fusion technology was applied to ferment corn powder two-stage enzymatic hydrolyzate to produce ethanol.
Firstly, a seed fermentor was coupled into the fermentation system composed of three tanks in series, and the hydrolyzate medium containing reducing sugar 100 g/L, added 2.0g/L (NH4)2HPO4 and KH2PO4, respectively, was fed into the seed fermentor at the dilution rate of 0.017h~' for the self-flocculating yeast seed cultivation. Meanwhile, the hydrolyzate medium containing reducing sugar 220 g/L, added 1.5g/L (NH4)2HPO4 and 2.5g/L KH2PO4) was fed into the first fermentor at the dilution rates of 0.017, 0.025, 0.033, 0.040 and 0.050 h-1, respectively. The experimental results showed that Chemostat states were developed for seed cultivation and ethanol fermentations under the dilution rates of 0.017, 0.025, 0.033 and 0.050 h-1. However, the oscillations of fermented parameters, including residual sugar, ethanol and biomass concentrations, were observed under the dilution rate of 0.040 h-1 for this continuous ethanol fermentation system. Over 12% (v/v) ethanol was achieved, at the same time, residual reducing sugar and total sugar were maintained below 0.15% and 0.35% (w/v) when the dilution rate was controlled no more than 0.033 h-1. The ethanol productivity of the entire fermentation system was calculated to be 2.71g/(L-h).
Secondly, the seed fermentor was removed from the above bioreactor syatem, and the yeast floes grown inside fermentors were used to support ethanol fermentation. Using same ferment medium, over 13% (v/v) ethanol was achieved, at the same time, residual reducing sugar and total sugar were maintained below 0.30% and 0.40% (w/v) when the dilution rate was controlled no more than 0.040 h-1. The ethanol productivity of the entire fermentation system was calculated to be 3.47g/(L-h). These experimental running indicated that this new ethanol fermentation technology using self-flocculating yeast cells can be applied to such a fermentation system composed of tanks in series that was widely used in ethanol fermentation industry in China at present and help improve its efficiency and save production cost.
Finally, the effect of the complete recycling use of distilled effluent on ethanol fermentation was examined in order to save the cost for the treatment of waste distillage and promote cleansing production. Based on the data of 13 circulations, although the average fermentation time needed to be extended to 30~40 hours, over 12.5% (v/v) ethanol was achieved, at the same time, residual reducing sugar and total sugar were maintained below 0.70% and 0.90% (w/v), which indicates that the negative effect of the completely recycling use of waste distillage was limited, and the waste distillage can be completely recycled within the fermentation system.
引文
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