Effects of Fluorescence Self Absorption of Algae in Sea Water

Candy Barbaran

Annie Becerra

Mentors: Prof. Fred Moshary

Dr. Alex Gilerson

NYCRI C

N p

Background

• Satellite imagery of ocean color provides important information about water quality and composition.

• Abundance of phytoplankton (algae) in the ocean can be traced by Chlorophyll A concentration which exists in all types of algae and has distinct spectral features.

• Part of the light energy absorbed by algae in blue-green emits in the red as chlorophyll fluorescence.

• Chlorophyll fluorescence is a vital parameter which can be considered as an indicator of chlorophyll concentration and photosynthetic activity in the ocean.

Analyze the spectral shape of chlorophyll fluorescence and absorption in order to detect patterns in the fluorescence spectral shift - detecting patterns in the spectral shift will allow us to eliminate all other factors and measure pure fluorescence.

Goal Improve algorithms for instruments aboard

satellites in order to make fluorescence measurements of algae more accurate.

Purpose of the experiment

Water Composition for Open Ocean and Coastal Waters

Algae Algae

Colored Dissolved Organic Matter (CDOM)

Minerals

Open ocean Coastal waters

Challenges of Using Chlorophyll Fluorescence Methods For Algae

Detection

• Fluorescence overlaps with the other spectral features. Special algorithms are needed to separate fluorescence from self absorption and elastic reflectance .

Sea surface

AlgaeDepth

600 650 700 750 8000.01

0.02

0.03

0.04

0.05

Fluorescence

745nm678nm

Re

fle

cta

nc

e

Wavelength (nm)

Reflectance peakdue to min absorption

Fluorescence heightover baseline

667nm

Reflectance

Reflectance +fluorescence

Fluorescence Magnitude and Peak over Baseline

Reflectance of the sun

Reflectance of the sun with fluorescence

Reflectance=

Eu/Ed~bb /a

WET Labs AC-S Instrument

The AC-S instrument measures absorption and attenuation at 82 wavelengths from 400 to 750 nm

Algae absorption measured by WET Labs ACS instrument

350 400 450 500 550 600 650 700 750 800 850

0

5

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Ab

sorp

tio

n (

m-1)

Wavelength (nm)

TetraselmisS TSDiluted Water

Fl zone

There is strong algae absorption in the fluorescence zone which causes change of fluorescence spectra

350 400 450 500 550 600 650 700 750 800 850

0

5

10

15

20

25

Ab

sorp

tio

n (

m-1)

Wavelength (nm)

Isochrysis IsochrysisDiluted Water

Fl zone

Changes of the fluorescence spectral shape due to algae absorption

Fl(λ)~(1/r2)*Fl0(λ)*exp(-a(λ)*h)

SpectrometerFP

Laser light from argon laserλ = 488 nm

h

r

Experimental Setup for the Measurements of Fluorescence Spectral Shape

Objects tested: algae Isochrysis sp., Tetraselmis striata, concentrations up to 4x106 cells/mL with necessary dilutions

• Algae are illuminated by the laser light at different depths. Emitted fluorescence is partially absorbed during propagation through algae.

• Spectral shape of fluorescence is measured by fiber optic sensor connected to the Ocean Optics spectrometer

SpectrometerFP

Laser light from argon laserλ = 488 nm

Shift of fluorescence of Algae Isochrysis as a function of depth under 488nm excitation

Fluorescence spectra for different excitation depths

Shift of fluorescence maximum for various excitation depths

Algae absorption peak

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0

500

1000

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2000

Inte

nsi

ty (

au)

Wavelength (nm)

Depth(mm) 5 18 23 41 54 63 73 78

0 10 20 30 40 50 60 70 80684

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Wav

elen

gth

flu

ore

scen

ce M

axim

um

(n

m)

Depth (mm)

Shift of Fluorescence of Tetraselmis striata as a function of Depth under 488 nm excitation

Fluorescence spectra for different excitation depths.

Shift of fluorescence maximum for various excitation depths

600 650 700 750 800

0

1000

Inte

ns

ity

(a

u)

Wavelength (nm)

Depth(mm) 3 18 32 51 62 72 83

0 20 40 60 80

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688

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Wav

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Flu

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(n

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Depth (mm)

Conclusions

•Both types of algae in the original concentration have strong absorption and cause spectral shift of fluorescence

•This spectral shift should be taken into account in the analysis of fluorescence and reflectance data for the waters with high chlorophyll (algae) concentrations

Further Work

• After accounting for absorption and attenuation we can graph pure fluorescence.

• Compare pure fluorescence results from the experiment to the calculated ones using Matlab software

Acknowledgements

Dr. Alex Gilerson

Professor Fred Moshary

NASA-NYCRI