Abstract
BACKGROUND: Purpose of the study was to establish the extinction coefficients of chlorophyll a and b in N,N-dimethylformamide(DMF) and Dimethylsulfoxide(DMSO) solvents and to find out the conditions of optimal extraction temperature and time in citrus leaves. METHODS AND RESULTS: Chlorophyll a and b standards were dissolved in DMF and DMSO. Extinction coefficients of chlorophyll pigments were determined and their contents were quantified using spectrophotometer. Chlorophyllous pigments of citrus(Citrus unshiu Marc. cv. Okitsu wase) leaves were extracted at 25, 40, 60 and $80^{\circ}C$ for 4, 6, 8, 24 and 48 hours to determine the optimal extraction condition. CONCLUSION: The extinction coefficients of chlorophyll a(Chl a) and chlorophyll b(Chl b) of DMF extracts for high extinction wavelength were 663.8 and 647.2 nm. Similarly, the high extinction wavelength of DMSO extracts were 665.8 and 649.0 nm for chl a and b respectively. Chl a, Chl b and total chlorophyll content of DMF extracts were Chl a = $12.10A_{663.8}-2.74A_{647.2}$, Chl b = $21.94A_{647.2}-5.06A_{663.8}$ and total $Chl=19.193A_{647.2}+7.04A_{663.8}$. Similarly, Chl a, Chl b and total Chl of DMSO extracts were Chl a = $14.53A_{665.8}-5.40A_{649.0}$, Chl b = $26.98A_{649.0}-7.11A_{665.8}$ and total $Chl=21.58A_{649.0}+7.43A_{665.8}$. The chlorophyll extracts of DMF and DMSO were very stable in dark. High chlorophyll contents of citrus leaves were found at $80^{\circ}C$ extraction for 6 hours in DMF and at $80^{\circ}C$ extraction for 24 hours in DMSO. However, the chlorophyll content was decreased significantly after 8 hours in DMF extraction while it was remained up to 30 hours in DMSO extraction.
