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A high throughput Nile red method for quantitative measurement of neutral lipids in microalgae
Chen, Wei1,2; Zhang, Chengwu1; Song, Lirong2; Sommerfeld, Milton1; Hu, Qiang1; Hu, Q, Arizona State Univ, Dept Appl Biol Sci, 7001 E Williams Field Rd, Mesa, AZ 85212 USA
2009-04-01
Source PublicationJOURNAL OF MICROBIOLOGICAL METHODS
ISSN0167-7012
Volume77Issue:1Pages:41-47
AbstractIsolation of high neutral lipid-containing microalgae is key to the commercial success of microalgae-based biofuel production. The Nile red fluorescence method has been successfully applied to the determination of lipids in certain microalgae, but has been unsuccessful in many others, particularly those with thick, rigid cell walls that prevent the penetration of the fluorescence dye. The conventional "one sample at a time" method was also time-consuming. In this study, the solvent dimethyl sulfoxide (DMSO) was introduced to microalgal samples as the stain carrier at an elevated temperature. The cellular neutral lipids were determined and quantified using a 96-well plate on a fluorescence spectrophotometer with an excitation wavelength of 530 nm and an emission wavelength of 575 run. An optimized procedure yielded a high correlation coefficient (R-2 = 0.998) with the lipid standard triolein and repeated measurements of replicates. Application of the improved method to several green algal strains gave very reproducible results with relative standard errors of 8.5%, 3.9% and 8.6%, 4.5% for repeatability and reproducibility at two concentration levels (2.0 mu g/mL and 20 mu g/mL), respectively. Moreover, the detection and quantification limits of the improved Nile red staining method were 0.8 mu g/mL and 2.0 mu g/mL for the neutral lipid standard triolein, respectively. The modified method and a conventional gravimetric determination method provided similar results on replicate samples. The 96-well plate-based Nile red method can be used as a high throughput technique for rapid screening of a broader spectrum of naturally-occurring and genetically-modified algal strains and mutants for high neutral lipid/oil production. (C) 2009 Published by Elsevier B.V.; Isolation of high neutral lipid-containing microalgae is key to the commercial success of microalgae-based biofuel production. The Nile red fluorescence method has been successfully applied to the determination of lipids in certain microalgae, but has been unsuccessful in many others, particularly those with thick, rigid cell walls that prevent the penetration of the fluorescence dye. The conventional "one sample at a time" method was also time-consuming. In this study, the solvent dimethyl sulfoxide (DMSO) was introduced to microalgal samples as the stain carrier at an elevated temperature. The cellular neutral lipids were determined and quantified using a 96-well plate on a fluorescence spectrophotometer with an excitation wavelength of 530 nm and an emission wavelength of 575 run. An optimized procedure yielded a high correlation coefficient (R(2) = 0.998) with the lipid standard triolein and repeated measurements of replicates. Application of the improved method to several green algal strains gave very reproducible results with relative standard errors of 8.5%, 3.9% and 8.6%, 4.5% for repeatability and reproducibility at two concentration levels (2.0 mu g/mL and 20 mu g/mL), respectively. Moreover, the detection and quantification limits of the improved Nile red staining method were 0.8 mu g/mL and 2.0 mu g/mL for the neutral lipid standard triolein, respectively. The modified method and a conventional gravimetric determination method provided similar results on replicate samples. The 96-well plate-based Nile red method can be used as a high throughput technique for rapid screening of a broader spectrum of naturally-occurring and genetically-modified algal strains and mutants for high neutral lipid/oil production. (C) 2009 Published by Elsevier B.V.
SubtypeArticle
KeywordBiofuel Fluorescence Green Algae Microalgae Nile Red Neutral Lipids
Subject AreaBiochemical Research Methods ; Microbiology
Department[Chen, Wei; Zhang, Chengwu; Sommerfeld, Milton; Hu, Qiang] Arizona State Univ, Dept Appl Biol Sci, Mesa, AZ 85212 USA; [Chen, Wei; Song, Lirong] Chinese Acad Sci, Inst Hydrobiol, Wuhan 430072, Hubei, Peoples R China
DOI10.1016/j.mimet.2009.01.001
WOS HeadingsScience & Technology ; Life Sciences & Biomedicine
Indexed BySCI
Language英语
WOS Research AreaBiochemistry & Molecular Biology ; Microbiology
WOS SubjectBiochemical Research Methods ; Microbiology
WOS KeywordRAPID SCREENING METHOD ; BIOTECHNOLOGY ; CELLS ; DYE
WOS IDWOS:000265314000007
Citation statistics
Cited Times:397[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://ir.ihb.ac.cn/handle/152342/7770
Collection中科院水生所知识产出(2009年前)
Corresponding AuthorHu, Q, Arizona State Univ, Dept Appl Biol Sci, 7001 E Williams Field Rd, Mesa, AZ 85212 USA
Affiliation1.Arizona State Univ, Dept Appl Biol Sci, Mesa, AZ 85212 USA
2.Chinese Acad Sci, Inst Hydrobiol, Wuhan 430072, Hubei, Peoples R China
Recommended Citation
GB/T 7714
Chen, Wei,Zhang, Chengwu,Song, Lirong,et al. A high throughput Nile red method for quantitative measurement of neutral lipids in microalgae[J]. JOURNAL OF MICROBIOLOGICAL METHODS,2009,77(1):41-47.
APA Chen, Wei,Zhang, Chengwu,Song, Lirong,Sommerfeld, Milton,Hu, Qiang,&Hu, Q, Arizona State Univ, Dept Appl Biol Sci, 7001 E Williams Field Rd, Mesa, AZ 85212 USA.(2009).A high throughput Nile red method for quantitative measurement of neutral lipids in microalgae.JOURNAL OF MICROBIOLOGICAL METHODS,77(1),41-47.
MLA Chen, Wei,et al."A high throughput Nile red method for quantitative measurement of neutral lipids in microalgae".JOURNAL OF MICROBIOLOGICAL METHODS 77.1(2009):41-47.
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