When in direct concentration mode the following calculations will be used to calculate corrected chlorophyll and pheophytin a values for the acidification method and the corrected chlorophyll values for the non-acidification method. Acidification Method
I. Variables stored during calibration phase of fluorometer
Cstand[1] = Concentration of standard 1 Fblank = Fluorescence of Blank value Fstand[1],B = Fluorescence of standard 1 before acidification Fstand[1],A = Fluorescence of standard 1 after acidification Fm = Acidification Ratio = (Fstand[1],B – Fblank) / (Fstand[1],A – Fblank)
(Note: If using more than one standard for calibration take only pre-acidification readings for standards 2-5)
II. Variables required from the sample analysis phase
Fsamp,B = Fluorescence of sample before acidification Fsamp,A = Fluorescence of sample after acidification Vsolvent = Volume of solvent used to extract sample Vwater = Volume of water filtered
III. Interpolation equation used in end calculation of chlorophyll and pheophytin a concentrations
Interp,B = Cstand[1] * (Fsamp,B - Fblank) / (Fstand[1],B - Fblank) Interp,A = Cstand[1] * (Fsamp,A - Fblank) / (Fstand[1],B - Fblank)
IV. End calculation for corrected chlorophyll and pheophytin a
Chlorophyll a concentration = [Fm/(Fm-1)] * (Interp,B - Interp,A) * (Vsolvent/ Vwater) Pheophytin a concentration = [Fm/(Fm-1)] * [(Fm * Interp,A) - Interp,B] * (Vsolvent/ Vwater)
V. End calculation for corrected chlorophyll a (Non-Acidification Method)
Chlorophyll a concentration = (Interp,B - Interp,A) * (Vsolvent/ Vwater)
(Note: Calibration values from the Acidification Method calibration curve can be used for calculating chlorophyll a concentrations from samples processed using Non-Acidification Method) |