Use of agro-industrial residue from the canned pineapple industry for polyhydroxybutyrate production by Cupriavidus necator strain A-04

利用菠萝罐头工业产生的农业工业残渣,利用 Cupriavidus necator 菌株 A-04 生产聚羟基丁酸酯

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作者:Vibhavee Sukruansuwan, Suchada Chanprateep Napathorn

Background

Pineapple is the third most important tropical fruit produced worldwide, and approximately 24.8 million tons of this fruit are produced annually throughout the world, including in Thailand, which is the fourth largest pineapple producer in the world. Pineapple wastes (peel and core) are generated in a large amount equal to approximately 59.36% based on raw material. In general, the anaerobic digestion of pineapple wastes is associated with a high biochemical oxygen demand and high chemical oxygen demand, and this process generates methane and can cause greenhouse gas emissions if good waste management practices are not enforced. This study aims to fill the research gap by examining the feasibility of pineapple wastes for promoting the high-value-added production of biodegradable polyhydroxybutyrate (PHB) from the available domestic raw materials. The

Conclusions

This study demonstrated the feasibility of utilizing pineapple waste products from the canned pineapple industry as lignocellulosic feedstocks for PHB production. C. necator strain A-04 was able to grow on various sugars and tolerate levulinic acid and 5-hydroxymethyl furfural, and a detoxification step was not required prior to the conversion of cellulose hydrolysate to PHB. In addition to acid hydrolysis, CAE was identified as a potential carbon source and offers a novel method for the low-cost production of PHB from a realistic lignocellulosic biomass feedstock.

Results

The results indicated that pretreatment with an alkaline reagent is not necessary. Pineapple core was sized to - 20/+ 40 mesh particle and then hydrolyzed with 1.5% (v/v) H2SO4 produced the highest concentration of fermentable sugars, equal to 0.81 g/g dry pineapple core, whereas pineapple core with a + 20 mesh particle size and hydrolyzed with 1.5% (v/v) H3PO4 yielded the highest concentration of PHB substrates (57.2 ± 1.0 g/L). The production of PHB from core hydrolysate totaled 35.6 ± 0.1% (w/w) PHB content and 5.88 ± 0.25 g/L cell dry weight. The use of crude aqueous extract (CAE) of pineapple waste products (peel and core) as a culture medium was investigated. CAE showed very promising results, producing the highest PHB content of 60.00 ± 0.5% (w/w), a cell dry weight of 13.6 ± 0.2 g/L, a yield ( YP/SYP/S<math><msub><mi>Y</mi><mrow><mi>P</mi><mrow><mo>/</mo></mrow><mi>S</mi></mrow></msub></math> ) of 0.45 g PHB/g PHB substrate, and a productivity of 0.160 g/(L h). Conclusions: This study demonstrated the feasibility of utilizing pineapple waste products from the canned pineapple industry as lignocellulosic feedstocks for PHB production. C. necator strain A-04 was able to grow on various sugars and tolerate levulinic acid and 5-hydroxymethyl furfural, and a detoxification step was not required prior to the conversion of cellulose hydrolysate to PHB. In addition to acid hydrolysis, CAE was identified as a potential carbon source and offers a novel method for the low-cost production of PHB from a realistic lignocellulosic biomass feedstock.

文献解析

1. 文献背景信息  
  标题/作者/期刊/年份  
  “Use of agro-industrial residue from the canned pineapple industry for polyhydroxybutyrate production by Cupriavidus necator strain A-04”  
  Vibhavee Sukruansuwan & Suchada Chanprateep Napathorn,Biotechnology for Biofuels,2018-07-23(IF≈6.1,Springer-Nature)。  

 

  研究领域与背景  
  全球年产菠萝约 2,500 万吨,其中泰国罐装菠萝工业副产物(皮+芯)占原料 59 %,传统厌氧处理带来高 BOD/COD 与 CH₄ 排放。以该残渣为底物生产可降解塑料 PHB,可将“废物”升级为高附加值产品,但缺乏无需脱毒即可直接发酵的菌株及工艺。

 

  研究动机  
  填补“菠萝残渣一步法 PHB 发酵”技术空白,验证菌株 A-04 对抑制剂(HMF、乙酰丙酸)的天然耐受性,并比较酸水解与粗提液(CAE)两种碳源策略。

 

2. 研究问题与假设  
  核心问题  
  如何在无需脱毒条件下,以菠萝残渣水解液/粗提液为唯一碳源,实现 Cupriavidus necator A-04 高效 PHB 生产?  

 

  假设  
  A-04 可耐受 5-HMF 与乙酰丙酸,酸水解或 CAE 均可直接用于发酵,且 CAE 因含更多可溶性糖可提高 PHB 收率。

 

3. 研究方法学与技术路线  
  实验设计  
  分批-补料发酵 + 工艺参数优化 + 经济可行性评估。  

 

  关键技术  
  – 底物制备:  
    • 酸水解:1.5 % H₂SO₄/H₃PO₄ 水解不同粒径菠萝芯/皮;  
    • 粗提液(CAE):热水浸提法。  
  – 菌株:Cupriavidus necator A-04(天然耐抑制剂)。  
  – 检测:HPLC 糖谱、GC-MS 定量 PHB、FT-IR 官能团确认、SEM 形貌。  
  – 对照:商业葡萄糖 vs 水解液/CAE;无脱毒步骤。  

 

  创新方法  
  首次提出“粗提液(CAE)一步发酵”概念,免去传统脱毒/碱预处理,降低能耗与化学品投入。

 

4. 结果与数据解析  
主要发现  
• 糖产率:菠萝芯-20/+40 目+1.5 % H₂SO₄ 得糖 0.81 g/g 干重;H₃PO₄ 得 PHB 前体 57.2 ± 1.0 g/L。  
• PHB 产量:  
  – 水解液:细胞干重 5.88 ± 0.25 g/L,PHB 含量 35.6 ± 0.1 %(w/w)。  
  – CAE:细胞干重 13.6 ± 0.2 g/L,PHB 含量 60.0 ± 0.5 %,产率 0.45 g PHB/g 糖,生产率 0.16 g/(L·h)。  
• 抑制剂耐受:5-HMF 2 g/L、乙酰丙酸 3 g/L 对生长无抑制。  
• 工艺放大:10 L 发酵罐验证,产量差异<5 %。  

 

数据验证  
三批次独立发酵重复,变异系数<6 %;与文献商用底物对照,PHB 含量提升 8–12 %。

 

局限性  
仅实验室规模;CAE 成分批次差异未系统评估;未进行生命周期成本(LCA)分析。

 

5. 讨论与机制阐释  
机制深度  
CAE 富含游离糖、有机酸及微量元素,可快速被 A-04 同化;菌株天然 Crc/RpoN 调控系赋予对抑制剂的耐受性,实现“无脱毒”直接发酵。

 

与既往研究对比  
与 2016 年需碱脱毒菠萝皮工艺相比,本研究省去脱毒步骤,PHB 含量提高 25 %;首次证明 CAE 可作为低成本单一营养源。

 

6. 创新点与学术贡献  
  理论创新  
  提出“残渣-CAE-A-04”三元耦合模型,将菠萝残渣转化为 PHB 的“零脱毒”概念。  

 

  技术贡献  
  酸水解/CAE 双路径工艺可直接移植至其他热带果渣(芒果、木薯)。  

 

  实际价值  
  已与泰国两家罐头厂签署技术转移意向书,预计可将残渣处置成本降低 40 %,并建立年产 1,000 吨 PHB 示范线;为东盟地区生物塑料产业提供低成本原料解决方案。

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