Some Problems in the Estimation of Chlorophyll‐a and Phaeopigments from Pre‐ and Post‐acidification Spectrophotometrie Measurements

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Abstract
The rate of conversion of chlorophyll‐a to phaeophytin‐a in dilute acid organic solvents is markedly pH‐dependent. The widely used spectrophotometric measurement before and after acidification to discriminate between chlorophyll‐a and phaeopigments is applicable only between pH 2.6—2.8. At higher pH the reaction proceeds slowly and occasionally may be incomplete. At lower pH complicating reactions may occur, such as breakdown of certain carotenoid pigments causing (time‐dependent) increase of background absorption, and formation of di‐cations of phaeophytin with spectral characteristics different from the usually obtained mono‐cations of phaeophytin. Consequently, it is stressed to control carefully the acid concentration, the water content of the solvent, and the amount of MgCO3 used in the filtration procedure. In general, for the purpose of spectrophotometric acidification experiments, acidification by dilute hydrochloric acid added to organic solvents containing 10—20% water, and omission of MgCO3 would be preferable.On the basis of observed complications in the red part of the absorption spectrum of the pigment extract, it is recommended to use the above stated pH‐range.

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