Evaluation of pigment extraction methods and a recommended protocol for periphyton chlorophyll a determination and chemotaxonomic assessment
Evaluation of pigment extraction methods and a recommended protocol for periphyton chlorophyll a determination and chemotaxonomic assessment
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DOI：
10.1111/j.1529-8817.2006.00257.x
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发表时间：
2006-10-01
影响因子：
2.9
通讯作者：
Mongkronsri, Panne
中科院分区：
文献类型：
--
作者：
Hagerthey, Scot E.;Louda, J. William;Mongkronsri, Panne
人们已经提出了许多提取和定量藻类色素的方法。比较研究发现，由于不同的细胞阻力，不同的溶剂和机械破坏方案的色素提取效率不同，从而导致对色素数据的潜在误解。当藻类的类型或抗性未知时，需要一种有效地从所有分类群中提取色素的方法。通过比较四种溶剂(丙酮、甲醇、甲醇/丙酮、甲醇/丙酮/N，N-二甲基甲酰胺)的萃取率、研磨效果和冷冻干燥效果，建立了一种简单有效的一步提取附生植物色素的方法。用甲醇/丙酮/DMF/水(MAD)提取的冻干样品提取效果最好。与用90%丙酮提取的新鲜/冷冻样品相比，冷冻干燥样品的叶绿素萃取率高出86%。冷冻干燥大大提高了极性和非极性(亲脂/疏水)色素的萃取率，而MAD则增加了极性色素的萃取率，并提高了所有色素的峰分辨率。在提取前新鲜/冷冻或冷冻干燥的样品之间的化学分类评估不同。与新鲜/冷冻相比，冻干材料中蓝藻的相对丰度更高，这是因为蓝藻色素的可萃取性提高。根据这项研究的结果，对于含有蓝藻的藻周样品或当垫子的组成未知时，不推荐使用90%丙酮作为溶剂的传统方法。冷冻干燥和MAD的组合足以从含有丝状蓝藻、绿藻和硅藻的围生藻席中提取色素。
Many methods have been proposed to extract and quantify algal pigments. Comparative studies have found that pigment extraction efficiency varies among solvent and mechanical disruption protocols due to differential cellular resistance, thereby, leading to potential misinterpretation of pigment data. When the type or resistance of algae are unknown, a method is required that efficiently extract pigments from all taxonomic groups. The objective of this study was to develop a simple and efficient one stage periphyton pigment extraction protocol by comparing the extractability of four solvents (acetone, methanol, methanol/acetone, and methanol/acetone/N,N-dimethylformamide), the effects of grinding, and the effects of freeze-drying. The best overall extraction was obtained using freeze-dried samples extracted with methanol/acetone/DMF/water (MAD). Eighty-six percent more chlorophyll was extracted when the sample was freeze-dried relative to fresh/frozen samples extracted with 90% acetone. Freeze-drying greatly improved the extraction of both polar and non-polar (lipophilic/hydrophobic) pigments while MAD increased the extractability of polar pigments and improved peak resolution of all pigments. Chemotaxonomic assessment differed between samples that were fresh/frozen or freeze-dried before extraction. The relative abundance of cyanobacteria was greater for freeze-dried material compared with fresh/frozen due to the improved extractability of cyanobacterial pigments. Based on the results of this study, the traditional approach of 90% acetone as a solvent is not recommended for periphyton samples containing cyanobacteria or when the composition of the mat is unknown. The combination of freeze-drying and MAD was sufficient for the extraction of pigments from a periphyton mat containing filamentous cyanobacteria, green algae, and diatoms.