金纳米颗粒（AuNPs）又称为胶体金，它的粒子尺寸在1~100nm之间，SPR吸收峰在540nm左右，因其独特的物理化学性质在生物医学方面得到广泛应用。为保护环境绿色合成AuNPs，近年来各国学者都在探索藻类合成法合成AuNPs，但是关于藻类合成AuNPs的准确条件研究不够细致，合成效率较低，合成机理也鲜有报道。有研究发现，用金诱变的蓝藻突变体能加强金与蓝藻细胞之间的相互作用。因此本研究试图利用集胞藻6803和小球藻2种藻类作为材料，探索其高效、稳定合成AuNPs的条件，并阐述其机理。

通过用不同浓度的AuCl₃溶液诱变集胞藻6803，获得了耐受金的集胞藻6803耐金突变体。突变体转化金离子实验结果显示，突变体滤液颜色变为紫红色，等离子体共振带（SPR）吸收峰出现在540nm处且比野生型合成量高约5倍，说明集胞藻6803耐金突变体能高效合成AuNPs。为证明是突变体细胞还是其滤液起作用，进行了如下实验。

（1）UV-vis光谱显示集胞藻6803耐金突变体滤液与细胞合成AuNPs的SPR吸收峰均出现在540nm，说明集胞藻6803耐金突变体滤液与细胞均有合成AuNPs的能力，但细胞合成AuNPs后的滤液中SPR吸收值比6803突变体滤液中小，说明细胞合成的AuNPs大部分被吸附在细胞内。

（2）突变体滤液的合成AuNPs受反应体系pH值、金属前驱物浓度的影响，碱性条件可以显著提高滤液合成AuNPs的产量，当pH为12时其产量比中性高约30%，pH（4、7、10、12）越大合成AuNPs的粒径越小，过酸时则无法合成AuNPs；金属前驱物的浓度与滤液合成AuNPs的浓度成正比，当前驱物的浓度低于25µg/mL时，滤液无法合成AuNPs。

（3）在探究滤液合成AuNPs机理实验中，FTIR图谱显示出羰基、胺基、羟基、酰胺等官能团，这是糖类、蛋白质的特征官能团，由此推测滤液中的蛋白质以及多糖对AuNPs的合成起了极其重要的作用。蛋白质与是否合成AuNPs的Spearman等级相关系数的绝对值接近1，说明滤液的蛋白质含量决定AuNPs的合成，滤液蛋白质浓度为15.72-35.42µg/mL时，一般会合成酒红色、紫红色、紫色AuNPs。

（4）在探究细胞合成AuNPs机理实验中，结果显示集胞藻6803耐金突变体吸附AuNPs后，别藻蓝蛋白的含量减少了33.96%，电镜结果显示AuNPs均匀分布在细胞内部的类囊体膜上，由此可以推测细胞合成AuNPs与胞内类囊体薄膜上的藻胆蛋白密切相关。

（5）抗氧化性测定结果：AuNPs的粒径越大其抗氧化能力越强。

除了原核的集胞藻6803外，还对真核的小球藻合成AuNPs进行了研究。其研究方法与集胞藻6803耐金突变体一致，结果表明：（1）小球藻滤液与细胞均有合成AuNPs的能力，碱性条件会提高AuNPs合成效率，合成粒径更小的AuNPs。（2）合成机制研究，滤液中的蛋白质对AuNPs的合成起重要作用。不同之处在于小球藻滤液的蛋白质含量与合成不同颜色的AuNPs有明显规律，即当小球藻滤液的蛋白质含量为29.43-38.48µg/mL、金属前驱物浓度为200µg/mL、pH=7.8时，小球藻滤液合成AuNPs的SPR吸收峰范围在576-586nm之间，为蓝色AuNPs，当蛋白质含量再增大时，小球藻滤液容易合成紫色AuNPs。

综上，无论原核的蓝藻还是真核的绿藻都有合成AuNPs的能力。通过机制研究，证明蓝藻细胞中的藻胆蛋白和滤液中的总蛋白对AuNPs的合成起重要作用，这为进一步提高AuNPs的产量提供根据，也使蓝藻合成的AuNPs在治疗应用方面成为可能。绿藻中没有藻胆蛋白，但有与藻胆蛋白相似的叶绿素蛋白复合体，是否绿藻细胞中AuNPs的合成与复合体有关，需要进一步的研究。

Gold nanoparticles (AuNPs), also known as colloidal gold, have a particle size of 1~100nm and SPR absorption peak of about 540nm. They are widely used in biomedical fields due to their unique physical and chemical properties. In order to protect the environment and synthesize AuNPs green, many countries have been exploring the synthesis of AuNPs by algal synthesis in recent years, but the exact conditions of algal synthesis of AuNPs are not detailed enough, the synthesis efficiency is low, and the synthesis mechanism is rarely reported. Some studies have found that cyanobacteria mutagenesis with gold can enhance the interaction between gold and cyanobacteria cells. Therefore, this study attempted to explore the conditions for the efficient and stable synthesis of AuNPs using two species of alga, Cystis 6803 and Chlorella, as materials, and to elaborate the mechanism.

Gold tolerant mutant of Cystellaria 6803 was obtained by mutagenesis with AuCl3 solution of different concentrations. The results showed that the filtrate color of the mutant changed to purplish red, and the absorption peak of the plasma resonance band (SPR) appeared at 540nm and the synthesis amount was about 5 times higher than that of the wild type, indicating that the gold tolerant mutation of cystophylla 6803 could effectively synthesize AuNPs. The following experiments were carried out to determine whether the mutant cells or their filtrates were responsible.

UV-vis spectra showed the SPR absorption peak of AuNPs synthesized from the filtrate of gold-tolerant mutant and AuNPs synthesized from cells was 540nm, indicating that the filtrate of gold-tolerant mutant and AuNPs synthesized from cells were both capable of synthesizing AUNPs, but the SPR absorption value of AUNPs synthesized from cells was smaller than that of 6803 mutant. It indicated that AuNPs synthesized by cells were mostly adsorbed in cells.

The synthetic AuNPs of the mutant filtrate were affected by the pH value of the reaction system and the concentration of metal precursors. Alkaline conditions significantly increased the yield of AuNPs synthesized by the filtrate. When pH was 12, the yield of AuNPs was about 30% higher than that of neutral. The concentration of metal precursors was proportional to the concentration of AuNPs synthesized from the filtrate, and AuNPs could not be synthesized from the filtrate when the concentration of the precursors was below 25µg/mL.

In the experiment to explore AuNPs synthesis mechanism of filtrate, FTIR spectra showed carbonyl, amine, hydroxyl, amide and other functional groups, which are characteristic functional groups of sugars and proteins. Therefore, it was speculated that proteins and polysaccharides in filtrate played an extremely important role in AuNPs synthesis. The absolute value of Spearman ranking correlation coefficient between protein and AuNPs synthesis was close to 1, indicating that the protein content of the filtrate determined the synthesis of AuNPs. When the protein concentration of the filtrate was 15.72-35.42µg/mL, AuNPs of wine red, purple red and purple were generally synthesized.

In the experiment to explore the mechanism of AuNPs synthesis in cells, the results showed that the content of allophycocyanin decreased by 33.96% after the adsorption of AuNPs by the 6803 gold-tolerant mutant. The electron microscope showed that AuNPs were uniformly distributed on the thylakoid membrane inside cells. It can be inferred that AuNPs synthesis is closely related to phycobilin on intracellular thylakoid membrane.

Antioxidant test results: The larger the size of AuNPs, the stronger the antioxidant capacity.

Synthesis of AuNPs from eukaryotic Chlorella was studied in addition to prokaryotic cystis 6803. The results showed that: (1) Both the filtrate and cells of Chlorella had the ability to synthesize AuNPs, and alkaline conditions improved the synthesis efficiency of AuNPs and synthesized AuNPs with smaller particle size. (2) Study on synthesis mechanism. Protein in filtrate plays an important role in AuNPs synthesis. The difference was that the protein content of Chlorella filtrate was obviously consistent with the synthesis of AuNPs with different colors. That is, when the protein content of Chlorella filtrate was 29.43-38.48µg/mL, the concentration of metal precursors was 200µg/mL, and the pH was 7.8, The SPR absorption peaks of AuNPs synthesized from Chlorella filtrate ranged from 576-586nm, and were blue AuNPs. The purple AuNPs were easily synthesized from Chlorella filtrate when the protein content increased.

In conclusion, both prokaryotic cyanobacteria and eukaryotic green algae were capable of synthesizing AuNPs. Through the mechanism study, it was proved that phycobilin in cyanobacteria cells and total protein in filtrate played an important role in the synthesis of AuNPs, which provided a basis for further increasing the yield of AuNPs and made it possible for the therapeutic application of AuNPs synthesized by cyanobacteria. There is no phycobilin in green algae, but there are chlorophyll-protein complexes similar to phycobilin. Whether the synthesis of AuNPs in green algae cells is related to the complex needs further study.