I have fresh water micro algae chlorella sp. I want to extract the complete pigment using simpler methods. I tried with 80% of acetone and 90% acetone + DMSO, but unable to extract completely. Means when we are using these methods there is some pigment are remains so the cell of algae is looking green in the pellet when centrifuging.
Mechanical homogenisation improves extraction by up to 20%.
Dimethyl formamide (DMF) proves to be the best extraction solvent.
However, chlorophyll A extraction efficiency is strongly species-specific and further influenced by the growth conditions, i.e. whether cells were in logarithmic or stationary growth phase. To achieve maximum values for chlorophyll A from samples of unknown algal species composition and physiological state it was found necessary to make at least two 24-h extractions in DMF. If the second extraction step contributed >20% to the total chlorophyll A concentration, a third DMF extraction is applied. Using this biomarker assay, up to 313 mg chlorophyll A can be obtained.
For further references you can read this paper :
http://link.springer.com/article/10.2478%2Fs11756-007-0048-x
Poornima's response seems good. I have published an in vitro flourescence method in 1997. This method will possibly not increase your extraction efficiency, but it might still be nice to know that you can do extremely simple extractions by simply injecting 20% of aqueous sample into 80% of Acetone/DMSO (1:1, vol:vol), no centrifugation or filtration. You can read more here:
https://www.researchgate.net/publication/223918536_A_simple_in_vitro_fluorescence_method_for_biomass_measurements_in_algal_growth_inhibition_tests?ev=prf_pub
I would expect pretty good performance with Chlorella, but of course you can extract more if you do sequential extractions as suggested by Poornima. Watch out and keep any light away from the extracts, extracted pigments are very light sensitive.
Article A simple in vitro fluorescence method for biomass measuremen...
Try to freeze-dry the algae wether in pellet or in a filter, grind with a turrax if possible or a glass rod in 100% acetone and ultra-sonicate during 1 min. Leave the extraction medium overnight and centrifuge the sample.
Hi Millor Sabara,
Why should you never use a mixture of solvents? There are several cases in analytical chemistry, where people markedly improved recovery and robustness when using a carefully selected mixture of solvents instead of a pure solvent. You probably have a good reason to make this statement in this context of chlorofyll extractions, please let us know why mixtures of extractors should not be used.
Best regards
Philipp
One thing i want to share with you that when we extract the chlorophyll from the chlorella using acetone method , after the extraction the cells remains green what is the reason ??
Murphy RR Photosynth Res (2006) 89:27–41
Consistent sets of spectrophotometric chlorophyll equations
for acetone, methanol and ethanol solvents
Hi Dear colleagues
I am looking for correlation to find conc, of pigments in extract. Using single solvent different equations are available, but if i go for binary solvent in extraction and want to measure conc. by UV before recovering the used solvent ,then which correlation could i use in case of binary solvent. Please update on this. Thanks
Dear @Amin i think @Philip mayer will be able to answer this question. I will also try to find the proper answer of this question and i will update it as sooon as i get the answer but right now i donot have any answer for this question
5 years later, post from @Alyson Telfer helped me. I just want to put the correct reference for future readers: Ritchie, R.J. (2006). Consistent Sets of Spectrophotometric Chlorophyll Equations for Acetone, Methanol and Ethanol Solvents. Photosynthesis Res., 89, 27-41. Article Consistent Sets of Spectrophotometric Chlorophyll Equations ...
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