1. Lab 3 Particulate Absorption Collin Roesler 5 July 2007

2. Particulate absorption Spectrophotometer –Quantitative Filter Technique –Corrections –Kishino method Ac9 –Calibration dependent method –Calibration independent method

3. Absorption: Filtration method Separates particles from dissolved Concentrates particles from dilute medium

4. Measure in Spectrophotometer Reference –Match optical density of filter pad –No variability Baseline –Blank filter pad in sample compartment (what is OD blankfilter :OD sample ?)

5. Compute absorption a (m -1 ) = 2.303 OD. L (m) What is L? V filtered = A eff h A h L = h = V (m 3 ) A (m 2 )

6. What about the scattering by the filter? Path length amplification Filter pad –Creates nearly isotropic light field –Increases optical path length –Increases absorption signal –How to correct for it? a (m -1 ) = 2.303 OD. V(m 3 ) A(m 2 )

7.  correction: path length amplification Approach –Thick cultures –Measure absorption in cuvette –Measure absorption on filter pad –Determine ratio,  = OD filt = optical. OD cuv geometric

8. Determining  Kiefer and SooHoo ’82, Mitchell and Kiefer ’85, ‘88 For low loadings –Very variable For high loadings –Approaches 2  = ODf ODc OD filter pad Issue: scattering in cuvette? cuvette Filter pad

9. What about the scattering by the filter? Path length amplification, empirical approach a (m -1 ) = 2.303 ODf corrected V(m 3 ) A(m 2 ) Filter pad creates multiple scattering environment The relationship between OD f and OD c is non linear of the form: OD f corrected = C 1 * OD f + C 2 * OD f 2 C 1 = 0.29 to 0.48 C 2 = 0.05 to 0.75 Mitchell 1990, Cleveland and Weidemann 1993; Moore et al. 1995; Arbones et al. 1996 among others

10. What about the scattering by the filter? Path length amplification, theoretical approach a (m -1 ) = 2.303 OD. V(m 3 ) A(m 2 ) a (m -1 ) = 2.303 ODfilt. 2 V(m 3 ) A(m 2 ) The geometric path length is t the thickness of the pad The optical path length is r, t /r = cos  where cos  is the average cosine of the light field If the filter pad creates an isotropic light field, cos  = 0.5 The optical path length is 2 * geometric path length Roesler 1998

11. Partitioning of particulate absorption First scan is total particles, a p Extract with methanol and scan again, a nap a phyt = a p – a nap Other issues –Phytoplankton “parts” –Detrital pigments –Phycobilipigments –Inorganics

12. Kishino et al. 1985

13. Summary Filter pad technique Filter sample, want high loading to overcome the variability in the blank filter pad absorption itself, but not muddy Reference? Extraction to separate particulates, nap Computation –Offset correction, Stramski and Babin 2002 –Beta correction, try all models –Absorption calculation, a p and a nap –Phytoplankton calculation, a phyt = a p - a nap

14. ac9 approach Unfiltered water samples –Pure water calibration –T/S correction –Scattering correction Particulate absorption method –a part = a total – a filt –Calibration independent method

15. Absorption from AC9 But spectra are problematic water calibration applied 1. Pure water calibration

16. Absorption from AC9 400 500 600 700 Wavelength amam c 2. Temperature and salinity correction

17. Absorption from AC9 400 500 600 700 Wavelength amam 3. Scattering correction a.Subtract a m (715 nm) “b not a function of ” spectrophotometric approach Stramski and Babin 2002

18. Absorption from AC9 400 500 600 700 Wavelength amam 3. Scattering correction b. Subtract fraction of b( ) c a m (715) * b’( ). b’(715) b’( ) = c( ) – a m ( ) a( ) = a m ( ) -

19. Absorption from AC9 Pure water calibration Temperature/Salinity correction Scattering correction Calculate scattering spectrum 400 500 600 700 Wavelength a c b

20. Particulate Absorption AC9 method Measure sample absorption, a t –Apply water cal, T/S, scattering corrections Measure sample filtrate absorpion, a f –Apply water cal, T/S, (scattering corrections?) Compute particulate absorption, a p = a t – a f Configurations –Single profiling instrument, multiple casts –Automated filtering on single instrument –Dual profiling instruments, sensor intercalibration

21. Particulate Absorption AC9 method Calibration Independent Measure sample absorption, a t –T/S, scattering corrections Measure sample filtrate absorpion, a f –T/S, (scattering corrections?) Compute particulate absorption, a p = a t – a f Configurations –Single profiling instrument, multiple casts –Automated filtering on single instrument

22. Automated shipboard flow-through method, calibration-independent Slade et al., 2006 a total a <0.2  m

23. Automated shipboard flow-through method Slade et al., 2006 a p (440) (m -1 ) 0.04 0.03 0.02 0.015 0.010 0.005 0.000

24. ac9 Summary Pure water calibrations Whole sample measurement Filtered sample measurement Data analysis –Apply water cals –Apply Temperature/Salinity corrections –Compute a p = a t – a filt, c p = c t - c filt –Apply scattering correction to a p