混合原料沼气厌氧发酵影响因素分析及工艺优化
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摘要
开展农业废弃物多元物料混合厌氧发酵研究是实现农业废弃物资源化利用和减少环境污染的重要途径。开展多种类物料混合厌氧发酵技术研究，可避免单一原料厌氧发酵缺陷，提高沼气发酵产气效率，从而高效率地解决环境污染与能源生产问题。基于该技术的产业价值和科学问题的前沿性，进行此方面的研究具有重要意义。本研究从原料配比、碳氮比（C/N）、底物浓度、接种物比例和温度等方面分析了这些因素对混合原料沼气厌氧发酵效果的影响，并基于这些因素，对混合原料沼气厌氧发酵工艺进行了优化。研究取得以下主要结果：
（1）原料混合发酵CH_4产量较原料单独发酵时有显著提高；3种原料按一定比例混合后沼气发酵效率高于2种原料混合。不同原料混合组，随原料配比的变化，沼气产量和CH_4产量呈先增加后减少的趋势；沼气产量和CH_4产量最大时的原料配比不同，牛鸡麦组为牛粪和鸡粪配比50:50，牛猪麦组为牛粪和猪粪配比75:25，鸡猪麦组为鸡粪和猪粪配比25:75。适宜配比下较高的发酵效率在于其保持了发酵料液中适宜的pH、总铵态氮和游离态NH3含量。混合原料发酵存在协同作用，鸡猪麦混合后CH_4产量提高程度最大，其次为牛猪麦组，牛鸡麦组最小，说明猪粪最适宜与其它粪便混合发酵。
（2）C/N对混合原料沼气发酵效果有显著影响。不同原料混合组，随C/N的增加，沼气产量和CH_4产量呈先增加后减少的趋势。对牛鸡麦组，适宜C/N为30:1，沼气产量和CH_4产量分别为589.6mL g~(-1)VS和262.5mL g~(-1)VS；对牛猪麦组，适宜C/N为25:1，沼气产量和CH_4产量分别为614.0mL g~(-1)VS和286.5mL g~(-1)VS；对鸡猪麦组，适宜C/N为25:1，沼气产量和CH_4产量分别为543.5mL g~(-1)VS和259.7mL g~(-1)VS。低C/N易导致铵态氮积累，降低产气率；高C/N易导致发酵液pH降低，抑制CH_4菌活性而降低产气效率。
（3）混料试验设计结果表明，3种原料混合厌氧发酵的CH_4产量显著高于单一原料及2种原料混合发酵；多原料间在发酵过程中的协同作用提高了CH_4产量；在混合发酵中，粪便对整体CH_4产量的贡献量显著高于秸秆，但不同种类粪便及秸秆间差异不明显。中心组合试验设计结果表明，原料配比和C/N比均影响CH_4产量，且两因素间存在交互作用，说明混合原料发酵体系中不同原料间的协同作用不仅仅在于平衡C/N比，还可能存在微生物协同、营养平衡等方面的作用。混料设计和中心组合设计均得到了优化的原料配比及对应的CH_4产量，两者间差异不明显，但中心组合设计具有更高的精度和更广泛的适用性。
（4）底物浓度对混合原料沼气发酵效果有显著影响。当底物浓度小于20gVS L~(-1)时，累积产气量和CH_4产量都随底物浓度的增加而线性增大，但过高的底物浓度会导致料液酸化，降低发酵效率。同一底物浓度下，牛猪麦组发酵效率最高，其次为牛鸡麦组，鸡猪麦组最差。接种比例（ISR）对混合原料沼气发酵效果有显著影响。当ISR小于2.4时，随接种比例增大，累积产气量和CH_4产量均线性增大；ISR越低，有机酸越容易积累而抑制CH_4菌活性，降低发酵效率。当ISR高于2.4时，进一步提高接种物量，并不能提高发酵效率。各组累积产气量和CH_4产量在同一ISR下表现的大小规律与底物浓度影响相同。
（5）温度对混合原料沼气厌氧发酵有显著影响。随温度升高，料液pH、铵态氮和游离NH3含量都显著增加。20℃下，料液酸化降低了产气效率；中温及高温下，CH_4菌对游离NH3的耐受性增强，较高的游离NH3浓度没有对发酵过程产生明显的抑制作用。不同温度下，C/N对混合原料沼气厌氧发酵的影响存在差异。35℃下，C/N15:1和20:1发生氨氮抑制，而55℃下C/N25:1也产生一定的氨氮抑制。35℃下，最佳C/N为25:1，CH_4产量为272mL g~(-1)VS；55℃下，最佳C/N为30:1，CH_4产量为286mL g~(-1)VS。氨氮是否对发酵产生抑制同时取决于底物的C/N和发酵温度。
（6）响应面法可以用于牛粪（DM）、鸡粪（CM）和麦秆混合厌氧发酵产沼气的工艺参数优化。C/N、DM/CM、底物浓度、ISR对混合厌氧发酵CH_4产量都有显著影响并且C/N和DM/CM、C/N和底物浓度、底物浓度和ISR之间存在的交互作用对CH_4产量也有显著影响。随C/N和DM/CM的增大，CH_4产量呈先升高后下降的趋势，但C/N对CH_4产量的影响大于DM/CM；随C/N和底物浓度的增大，CH_4产量呈先升高后下降的趋势，但C/N对CH_4产量的影响大于底物浓度；随底物浓度和接种比例的增加，CH_4产量呈先增大后减小的趋势，接种比例对CH_4产量的影响大于底物浓度，在底物浓度和ISR都较小时，CH_4产量较低。通过模型预测得到牛粪、鸡粪和麦秆混合厌氧发酵产沼气的最优工艺组合为C/N为26.31，DM/CM为42.96:57.04，底物浓度为15.90g VS L~(-1)，ISR为2.34，预期可得最大CH_4产量为394mL g~(-1)VS。
Developing knowledge in anaerobic co-digestion of various substrates is an importantway to transform agricultural residues into resources and to reduce environmental pollution.Anaerobic co-digestion, for one thing, can replace the digestion of single substrate with thedrawbacks of low efficiency, long retention time and floating, and for another, improve theproduction efficiency and the economic benefit of organic waste treatment process during theindustrialization of methane production. Therefore, based on the industrial value of thistechnology and as the key scientific problem, conducting the research in anaerobicco-digestion is of great significance. This research analyzed the influences of feedingcomposition, C/N, inital substrate loadingand inoculum to substrate ratio (ISR) on theefficiency of anaerobic co-digestion, and based on these factors, optimized this process. Themain results were as followed:
(1) The methane yield in anaerobic co-digestion was increased compared with thedigestion of single substrate. Anaerobic co-digestion of three raw substrates were better thanthat of two raw substrates. With the changes of feeding composition, the biogas yield andmethane potential increased first and then decreased. The feeding composition was differentfor different mixtures in the highest biogas yield and methane potential, with DM (Dairymanure)/CM (Chicken manure)50:50in the mixture of DM, CM and wheat straw (WS),DM/SM (Swine manure)75:25in the mixture of DM, SM and WS, and CM/SM75:25in themixture of CM, SM and WS. The high efficiency in suitable feeding ratios lied in theappropriate pH value and total ammonia and free NH3contents. There were synergetic effectsamong various substrates, with the best effects in the mixture of CM, SM and WS, followedby the mixture of DM, SM and RS, and the least in the mixture of DM, CM and RS,suggesting that SM was the best substrate for anaerobic co-digestion.
(2) C/N had significant effects on the effects of anaerobic co-digestion. For differentmixtures, with the increase of C/N, the biogas yield and methane potential increased first andthen decreased. For the mixture of DM, CM and RS, the biogas yield and methane potentialwere589.6mL g~(-1)VS and262.5mL g~(-1)VS, respectively; for the mixture of DM, SM and WS,the biogas yield and methane potential were614.0mL g~(-1)VS and286.5mL g~(-1)VS,respectively; for the mixture of CM, SM and RS, the biogas yield and methane potential were 543.5mL g~(-1)VS and259.7mL g~(-1)VS, respectively. Lower C/N led to the accumulation ofammonia and then reduced the biogas yield, whereas, higher C/N led the decrease of pH value,inhibiting the activities of methanobacteria and reducing the efficiency.
(3) The results in mixture design (MD) suggested that the methane potential of three rawsubstrates was higher than those of single or two substrates. The synergetic effects amongsubstrates improved the methane potential and the contribution of manure on the improvedmethane potential was higher than straw, but no significant difference was found amongdifferent manures or straws. The results in central composite design (CCD) suggested that thefeeding composition, C/N and their interaction affecting the methane potential, indicating thatthe synergetic effects were not only due to the balanced C/N, but also the synergy ofmicroorganisms and nutritional balance. Optimum feeding compositions were obtainedthrough MD and CCD, but no significant difference was found between these two methods.However, CCD was more accurate and had wider range of applicability.
(4) The initial substrate loading had significant effects on biogas production. When thesubstrate concentration was below20gVS L~(-1), the biogas yield and methane potential linearlyincreased with the increase of substrate concentration, however, higher substrateconcentration easily led to the acidification of liquid and reduce fermentation efficiency.Under the same substrate concentration, the mixture of DM, SM and RS had the highestfermentation efficiency, followed by the mixture of DM, CM and RS, and the mixture of CM,SM and RS is the worst. ISR had a significant impact on biogas production. When the ISRwas less than2.4, with the increase of ISR, the cumulative biogas production and methanepotential linearly increased. The lower the ISR, the more easily the acid accumulated, withhigher inhibition on methanobacteria. When the ISR was higher than2.4, increased ISR wasunable to further improve the efficiency. Under the same ISR, the biogas yield and methanepotential were the same with the treatment under the same substrate concentration.
(5) Temperature significantly influenced the biogas production. With increasedtemperature, pH in liquid, total ammonium and free NH3contents all increased significantly.Under20°C, accumulated liquid acids reduced the efficiency. Under mesophilic andthermophilic temperatures, methanobacteria had higher tolerance on free NH3and the higherfree NH3contents did not inhibit the fermentation process. Under different temperature, theeffects of C/N on anaerobic co-digestion were different. Under35°C, ammonia inhibitionoccurred under C/N15:1and20:1, whereas, under55°C, the same inhibition also occurredunder C/N25:1. Under35°C, the optimum C/N was25:1, with the methane potential of272mL g~(-1)VS, and under55°C, the optimum C/N was30:1, with the methane potential of286mLg~(-1)VS. Whether the ammonia inhibition occurred or not depended both on the C/N and temperature.
(6) Response surface methodology can be used for optimizing the process of anaerobicco-digestion of DM, DM and RS. C/N, DM/CM, initial substrate loading and ISR all hadsignificant effects on methane potential and interctions between C/N and DM/CM, C/N andinitial substrate loading were significant on methane potential, initial substrate loading andISR. With the increase of C/N and DM/CM, the methane potential increased first and thendecreased, but the effects of C/N were higher than DM/CM. With the increase of C/N andsubstrate concentrations, the methane potential increased first and then decreased, but theeffects of C/N were higher than substrate concentrations. With the increase of substrateconcentrations and ISR, the methane potential increased first and then decreased, but theeffects of ISR were higher than substrate concentrations. When the initial substrate loadingand ISR were low, methane potential was also low. The highest methane of394mL g~(-1)VSwas predicted under optimum conditions with C/N of26.31, DM/CM of42.96:57.04, aninitial substrate loading of15.90g VS L~(-1)and ISR of2.34.
（1）原料混合发酵CH_4产量较原料单独发酵时有显著提高；3种原料按一定比例混合后沼气发酵效率高于2种原料混合。不同原料混合组，随原料配比的变化，沼气产量和CH_4产量呈先增加后减少的趋势；沼气产量和CH_4产量最大时的原料配比不同，牛鸡麦组为牛粪和鸡粪配比50:50，牛猪麦组为牛粪和猪粪配比75:25，鸡猪麦组为鸡粪和猪粪配比25:75。适宜配比下较高的发酵效率在于其保持了发酵料液中适宜的pH、总铵态氮和游离态NH3含量。混合原料发酵存在协同作用，鸡猪麦混合后CH_4产量提高程度最大，其次为牛猪麦组，牛鸡麦组最小，说明猪粪最适宜与其它粪便混合发酵。
（2）C/N对混合原料沼气发酵效果有显著影响。不同原料混合组，随C/N的增加，沼气产量和CH_4产量呈先增加后减少的趋势。对牛鸡麦组，适宜C/N为30:1，沼气产量和CH_4产量分别为589.6mL g~(-1)VS和262.5mL g~(-1)VS；对牛猪麦组，适宜C/N为25:1，沼气产量和CH_4产量分别为614.0mL g~(-1)VS和286.5mL g~(-1)VS；对鸡猪麦组，适宜C/N为25:1，沼气产量和CH_4产量分别为543.5mL g~(-1)VS和259.7mL g~(-1)VS。低C/N易导致铵态氮积累，降低产气率；高C/N易导致发酵液pH降低，抑制CH_4菌活性而降低产气效率。
（3）混料试验设计结果表明，3种原料混合厌氧发酵的CH_4产量显著高于单一原料及2种原料混合发酵；多原料间在发酵过程中的协同作用提高了CH_4产量；在混合发酵中，粪便对整体CH_4产量的贡献量显著高于秸秆，但不同种类粪便及秸秆间差异不明显。中心组合试验设计结果表明，原料配比和C/N比均影响CH_4产量，且两因素间存在交互作用，说明混合原料发酵体系中不同原料间的协同作用不仅仅在于平衡C/N比，还可能存在微生物协同、营养平衡等方面的作用。混料设计和中心组合设计均得到了优化的原料配比及对应的CH_4产量，两者间差异不明显，但中心组合设计具有更高的精度和更广泛的适用性。
（4）底物浓度对混合原料沼气发酵效果有显著影响。当底物浓度小于20gVS L~(-1)时，累积产气量和CH_4产量都随底物浓度的增加而线性增大，但过高的底物浓度会导致料液酸化，降低发酵效率。同一底物浓度下，牛猪麦组发酵效率最高，其次为牛鸡麦组，鸡猪麦组最差。接种比例（ISR）对混合原料沼气发酵效果有显著影响。当ISR小于2.4时，随接种比例增大，累积产气量和CH_4产量均线性增大；ISR越低，有机酸越容易积累而抑制CH_4菌活性，降低发酵效率。当ISR高于2.4时，进一步提高接种物量，并不能提高发酵效率。各组累积产气量和CH_4产量在同一ISR下表现的大小规律与底物浓度影响相同。
（5）温度对混合原料沼气厌氧发酵有显著影响。随温度升高，料液pH、铵态氮和游离NH3含量都显著增加。20℃下，料液酸化降低了产气效率；中温及高温下，CH_4菌对游离NH3的耐受性增强，较高的游离NH3浓度没有对发酵过程产生明显的抑制作用。不同温度下，C/N对混合原料沼气厌氧发酵的影响存在差异。35℃下，C/N15:1和20:1发生氨氮抑制，而55℃下C/N25:1也产生一定的氨氮抑制。35℃下，最佳C/N为25:1，CH_4产量为272mL g~(-1)VS；55℃下，最佳C/N为30:1，CH_4产量为286mL g~(-1)VS。氨氮是否对发酵产生抑制同时取决于底物的C/N和发酵温度。
（6）响应面法可以用于牛粪（DM）、鸡粪（CM）和麦秆混合厌氧发酵产沼气的工艺参数优化。C/N、DM/CM、底物浓度、ISR对混合厌氧发酵CH_4产量都有显著影响并且C/N和DM/CM、C/N和底物浓度、底物浓度和ISR之间存在的交互作用对CH_4产量也有显著影响。随C/N和DM/CM的增大，CH_4产量呈先升高后下降的趋势，但C/N对CH_4产量的影响大于DM/CM；随C/N和底物浓度的增大，CH_4产量呈先升高后下降的趋势，但C/N对CH_4产量的影响大于底物浓度；随底物浓度和接种比例的增加，CH_4产量呈先增大后减小的趋势，接种比例对CH_4产量的影响大于底物浓度，在底物浓度和ISR都较小时，CH_4产量较低。通过模型预测得到牛粪、鸡粪和麦秆混合厌氧发酵产沼气的最优工艺组合为C/N为26.31，DM/CM为42.96:57.04，底物浓度为15.90g VS L~(-1)，ISR为2.34，预期可得最大CH_4产量为394mL g~(-1)VS。
Developing knowledge in anaerobic co-digestion of various substrates is an importantway to transform agricultural residues into resources and to reduce environmental pollution.Anaerobic co-digestion, for one thing, can replace the digestion of single substrate with thedrawbacks of low efficiency, long retention time and floating, and for another, improve theproduction efficiency and the economic benefit of organic waste treatment process during theindustrialization of methane production. Therefore, based on the industrial value of thistechnology and as the key scientific problem, conducting the research in anaerobicco-digestion is of great significance. This research analyzed the influences of feedingcomposition, C/N, inital substrate loadingand inoculum to substrate ratio (ISR) on theefficiency of anaerobic co-digestion, and based on these factors, optimized this process. Themain results were as followed:
(1) The methane yield in anaerobic co-digestion was increased compared with thedigestion of single substrate. Anaerobic co-digestion of three raw substrates were better thanthat of two raw substrates. With the changes of feeding composition, the biogas yield andmethane potential increased first and then decreased. The feeding composition was differentfor different mixtures in the highest biogas yield and methane potential, with DM (Dairymanure)/CM (Chicken manure)50:50in the mixture of DM, CM and wheat straw (WS),DM/SM (Swine manure)75:25in the mixture of DM, SM and WS, and CM/SM75:25in themixture of CM, SM and WS. The high efficiency in suitable feeding ratios lied in theappropriate pH value and total ammonia and free NH3contents. There were synergetic effectsamong various substrates, with the best effects in the mixture of CM, SM and WS, followedby the mixture of DM, SM and RS, and the least in the mixture of DM, CM and RS,suggesting that SM was the best substrate for anaerobic co-digestion.
(2) C/N had significant effects on the effects of anaerobic co-digestion. For differentmixtures, with the increase of C/N, the biogas yield and methane potential increased first andthen decreased. For the mixture of DM, CM and RS, the biogas yield and methane potentialwere589.6mL g~(-1)VS and262.5mL g~(-1)VS, respectively; for the mixture of DM, SM and WS,the biogas yield and methane potential were614.0mL g~(-1)VS and286.5mL g~(-1)VS,respectively; for the mixture of CM, SM and RS, the biogas yield and methane potential were 543.5mL g~(-1)VS and259.7mL g~(-1)VS, respectively. Lower C/N led to the accumulation ofammonia and then reduced the biogas yield, whereas, higher C/N led the decrease of pH value,inhibiting the activities of methanobacteria and reducing the efficiency.
(3) The results in mixture design (MD) suggested that the methane potential of three rawsubstrates was higher than those of single or two substrates. The synergetic effects amongsubstrates improved the methane potential and the contribution of manure on the improvedmethane potential was higher than straw, but no significant difference was found amongdifferent manures or straws. The results in central composite design (CCD) suggested that thefeeding composition, C/N and their interaction affecting the methane potential, indicating thatthe synergetic effects were not only due to the balanced C/N, but also the synergy ofmicroorganisms and nutritional balance. Optimum feeding compositions were obtainedthrough MD and CCD, but no significant difference was found between these two methods.However, CCD was more accurate and had wider range of applicability.
(4) The initial substrate loading had significant effects on biogas production. When thesubstrate concentration was below20gVS L~(-1), the biogas yield and methane potential linearlyincreased with the increase of substrate concentration, however, higher substrateconcentration easily led to the acidification of liquid and reduce fermentation efficiency.Under the same substrate concentration, the mixture of DM, SM and RS had the highestfermentation efficiency, followed by the mixture of DM, CM and RS, and the mixture of CM,SM and RS is the worst. ISR had a significant impact on biogas production. When the ISRwas less than2.4, with the increase of ISR, the cumulative biogas production and methanepotential linearly increased. The lower the ISR, the more easily the acid accumulated, withhigher inhibition on methanobacteria. When the ISR was higher than2.4, increased ISR wasunable to further improve the efficiency. Under the same ISR, the biogas yield and methanepotential were the same with the treatment under the same substrate concentration.
(5) Temperature significantly influenced the biogas production. With increasedtemperature, pH in liquid, total ammonium and free NH3contents all increased significantly.Under20°C, accumulated liquid acids reduced the efficiency. Under mesophilic andthermophilic temperatures, methanobacteria had higher tolerance on free NH3and the higherfree NH3contents did not inhibit the fermentation process. Under different temperature, theeffects of C/N on anaerobic co-digestion were different. Under35°C, ammonia inhibitionoccurred under C/N15:1and20:1, whereas, under55°C, the same inhibition also occurredunder C/N25:1. Under35°C, the optimum C/N was25:1, with the methane potential of272mL g~(-1)VS, and under55°C, the optimum C/N was30:1, with the methane potential of286mLg~(-1)VS. Whether the ammonia inhibition occurred or not depended both on the C/N and temperature.
(6) Response surface methodology can be used for optimizing the process of anaerobicco-digestion of DM, DM and RS. C/N, DM/CM, initial substrate loading and ISR all hadsignificant effects on methane potential and interctions between C/N and DM/CM, C/N andinitial substrate loading were significant on methane potential, initial substrate loading andISR. With the increase of C/N and DM/CM, the methane potential increased first and thendecreased, but the effects of C/N were higher than DM/CM. With the increase of C/N andsubstrate concentrations, the methane potential increased first and then decreased, but theeffects of C/N were higher than substrate concentrations. With the increase of substrateconcentrations and ISR, the methane potential increased first and then decreased, but theeffects of ISR were higher than substrate concentrations. When the initial substrate loadingand ISR were low, methane potential was also low. The highest methane of394mL g~(-1)VSwas predicted under optimum conditions with C/N of26.31, DM/CM of42.96:57.04, aninitial substrate loading of15.90g VS L~(-1)and ISR of2.34.
引文
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