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中国精品科技期刊2020
朱林清,曾名湧. 聚球藻PCC7002藻蓝蛋白提取纯化及热致褪色机理研究[J]. 食品工业科技,2022,43(16):32−40. doi: 10.13386/j.issn1002-0306.2021110096.
引用本文: 朱林清,曾名湧. 聚球藻PCC7002藻蓝蛋白提取纯化及热致褪色机理研究[J]. 食品工业科技,2022,43(16):32−40. doi: 10.13386/j.issn1002-0306.2021110096.
ZHU Linqing, ZENG Mingyong. Extraction and Purification and Heat-induced Fading Mechanism of Synechococcus sp. PCC7002 Phycocyanin[J]. Science and Technology of Food Industry, 2022, 43(16): 32−40. (in Chinese with English abstract). doi: 10.13386/j.issn1002-0306.2021110096.
Citation: ZHU Linqing, ZENG Mingyong. Extraction and Purification and Heat-induced Fading Mechanism of Synechococcus sp. PCC7002 Phycocyanin[J]. Science and Technology of Food Industry, 2022, 43(16): 32−40. (in Chinese with English abstract). doi: 10.13386/j.issn1002-0306.2021110096.

聚球藻PCC7002藻蓝蛋白提取纯化及热致褪色机理研究

Extraction and Purification and Heat-induced Fading Mechanism of Synechococcus sp. PCC7002 Phycocyanin

  • 摘要: 本文旨在研究聚球藻PCC7002(Synechococcus sp. PCC7002,简称“聚球藻”)藻蓝蛋白(Phycocyanin,PC)的提取纯化及热致褪色机理,通过高压均质、壳聚糖絮凝和硫酸铵盐析提取纯化藻蓝蛋白,并采用色泽指标、UV-Vis光谱、荧光发射光谱、粒径电位、FTIR光谱、SDS-PAGE电泳研究其在热处理(50、60、70、80、90 ℃,30 min)过程中褪色的机理。结果表明:藻蓝蛋白的最适提取条件是将藻粉溶于0.04 mol/L的NaCl溶液中使其浓度达到2 mg/mL并在80 MPa下均质7 min,此时藻蓝蛋白得率为10.5081% ± 0.0936%;经0.15 mg/mL壳聚糖絮凝、50%饱和硫酸铵盐析后,藻蓝蛋白纯度可由0.6950 ± 0.0043提高至1.9084 ± 0.2621。进一步研究藻蓝蛋白的热致褪色机理发现:从60 ℃开始,脱辅基蛋白空间结构的破坏使蛋白骨架维持的藻蓝胆素天然构象发生转变、藻蓝蛋白紫外吸收和特征荧光大幅下降,藻蓝蛋白的蓝色色泽因此大幅消褪;粒径结果表明,藻蓝蛋白在60 ℃时开始聚集,而80 ℃下形成的更大聚集体可能将四吡咯发色团包埋于其内,使得藻蓝蛋白色泽消褪程度加深;此外,FTIR光谱和SDS-PAGE表明热处理过程中藻蓝蛋白β亚基的破坏程度可能远高于α亚基,且被破坏的结构主要是α-螺旋。综上,维持藻蓝蛋白脱辅基蛋白结构或者发色团依托蛋白结构的稳定,是藻蓝蛋白热处理过程中呈色稳定的关键。本研究结果为藻蓝蛋白在热处理过程中护色措施的研究提供了一定的理论依据。

     

    Abstract: This paper aimed to study the extraction and purification of phycocyanin from Synechococcus sp. PCC7002 (hereinafter referred to as “Synechococcus”) and its heat-induced fading mechanism. Phycocyanin was extracted and purified through high-pressure homogenization, chitosan flocculation and ammonium sulfate salting out. And its fading mechanism during heat treatment (50 ℃, 60 ℃, 70 ℃, 80 ℃, 90 ℃, 30 min) was studied by color index, UV-Vis spectrum, fluorescence emission spectrum, particle size and potential, FTIR spectrum and SDS-PAGE electrophoresis. The results showed that optimal extraction conditions of phycocyanin were to dissolve algal power in 0.04 mol/L NaCl solution to make the concentration reach 2 mg/mL and homogenize it at 80 MPa for 7 min, and the resulting yield of phycocyanin was 10.5081%±0.0936%. After flocculation by 0.15 mg/mL chitosan and salting out by 50% saturated ammonium sulfate, phycocyanin purity could be increased from 0.6950 ± 0.0043 to 1.9084±0.2621. Further research on thermal-induced fading mechanism of phycocyanin found: Destruction of spatial structure of the apoprotein started from 60 ℃, which led to transformation of natural conformation of phycocyanobilin chromophore maintained by protein backbone and a significant decrease in ultraviolet absorption and characteristic fluorescence of phycocyanin, therefore blue hue of phycocyanin was greatly destroyed. Particle size results indicated that phycocyanin molecules began to aggregate at 60 ℃, while larger aggregates formed at 80 ℃ may lead tetrapyrrole chromophore to be buried in it, deepening fading degree of phycocyanin color. In addition, FTIR spectroscopy and SDS-PAGE indicated that β subunit of phycocyanin was probably damaged more serious than α subunit during heat treatment, and the damaged structure were mainly α-helix. In summary, maintaining structural stability of apoprotein or protein segment near the phycocyanin chromophore was the key factor of keeping phycocyanin tone stable during the heat treatment. This research would provide certain theoretical basis for the study of phycocyanin color protection technology during the heat treatment process.

     

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