I took 2 gm tissue and then added 20ml solvent. After that I collected supernatent and then made up volume 25 ml by about 10 ml hexane. I know absorbance reading and A1% value, I am a bit confused about total volume used amount. Would you someone assist me regarding that "V" value ?
I guess you diluted your supernatant before you take the absorption value, so your V is 25. You need to multiply the absorption-derived carotenoid concentration by 25 to find the total carotenoid quantity and then divide by 2 gm to find the concentration of carotenoids/gm of tissue.
we use the method of Goodwin(1965) in a routine manner to estimate total carotenoids,especially in citrus fruits,using 80% acetone for the extraction and absorbance at 480nm. Please, see the book of Goodwin(1965) for the process of calculation.
I recommend the handbook of RODRIGUEZ-AMAYA for carotenoid analyses.
I follow Wellburn (1994) and attached the pdf file here. This method also measure chlorophyll using various solvents with spectrophotometers of different resolutions. I use 80% (v/v) acetone extractant and incubation of the sample in the extractant overnight in a ratio of 20mg sample to every 20 mL extractant (1 wt of sample:1 volume of extractant) is overnight then read the absorbances at three different wavelengths, with the computed levels all in ppm or mg/L following the formula prescribed in Wellburn. The measurement is straight forward and encountered no difficulty with my rice leaf samples. If there is no component in citrus samples that interferes with the extraction, the extract should be clear and homogeneous, otherwise clarification needs to be done first before taking the absorbance readings.
Spectrofotometry using beta-carotene as standad or fluorescence method for the carotenoids B-carotene, lutein, lutein epoxide and violaxanthin are fluorescent
compounds. See
THE FLUORESCENCE OF CAROTENOIDS
F. T. Wolf and
M. V. Stevens
Article first published online: 2 JAN 2008
DOI: 10.1111/j.1751-1097.1967.tb08761.x
- Badges
- Science topic