Actually I am trying to estimate the chlorophyll content for bio diesel production and identification of the unknown micro algae.
Hi! We use a combination cell disruption (bead mill) + chlorophyll extraction with acetone. Then you can photometrically estimate chlorophyll content (after Lichtenthaler and Wellburn, 1983). It is maybe easier to determine the microalgae species first, and then to look for the suitable disruption/extraction conditions in literature.
Hi
I think you should follow the protocol of Arnon (1949) using 80% acetone. You take biomass and extract chlorophyll with 80% acetone and take reading 645 nm and 663 nm for both chl-a and b.
@gour i have to take reading every day that means for a particular period of time or only once
For best method of Chlorophyll estimation, you can refer the following link......... C.-H. Su, C.-C. Fu, Y.-C. Chang et al., “Simultaneous estimation
of chlorophyll a and lipid contents in microalgae by three-color
analysis,” Biotechnology and Bioengineering, vol. 99, no. 4, pp.
1034–1039, 2008.
you can follow the porra et.al 1989 protocol, link http://www.chem.ucla.edu/dept/Faculty/merchant/pdf/Porra_1989.pdf
i think this is the best method for extracting chlorophyll from microalgae.
Dr. Gaur Gopal Satpati,
How much volume of 80% acetone and mass of algae we should use for extraction in the method mentioned by you, because the measured absorbance will depend on it.
The best for estimation of chlorophyll content (mg/L) in microalgae as follows;
Two milliliter of culture broth is taken in centrifuge tube, ultrasonicated for 10 min in ice bath with two milliliter of 90% methanol over night. Then the homogenate is centrifuged at 3000 rpm for 5 min. The supernatant is separated and absorbance is read at 665nm and the amount of chlorophyll can be calculated using following formula;
Chlorophyll a (mg L-1) = 13.43 x OD665
I used 100% methanol and maintain the temperature,65C for 1h. Use the the equation described in porra et. al. For the estimation of chrophyll content
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