Chemical Imaging and Biodiversity of Microalgal Cells Using FTIR Microspectroscopy

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Chemical Imaging and Biodiversity of Microalgal
Cells Using FT-IR Microspectroscopy
Zuheir El-bayyari Department of Basic Sciences,
Philadelphia University- Amman - Jordan
Carol Hirschmugl Department of Physics,
University of Wisconsin Milwaukee - USA
Mario Giordano Marine Biology, University Del La
Marche- Ancona, ITALY
and Craig Sandgren Department of Biology,
University of Wisconsin Milwaukee - USA
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Motivation

• To understand the biochemistry in individual
  living cells of different microalgal cells.

• To determine the relative concentration of
  macromolecules such as proteins, lipids,
  phospholipids, phosphates, silica, and
  carbohydrates in the cells. ..etc.

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Port 031-IR Beamline at SRC
This Beamline is SRC Owned
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FT-IR Data Collection

• IR spectra were collected as absorbance spectra
  using an

• IR Scope II IR microscope (Thermo Nicolet)
  equipped with a small sized MCT/A (HgCdTe)
  detector .

• The microscope was linked to Magna System 560
  FT-IR spectrometer (Nicolt) equipped with a
  software controlled x,y stage.

• A specially designed microscope box was purged by
  dry air reducing spectral contributions from
  atmospheric water vapor and CO2.

• FT-IR spectra were recorded using
• 20 µm x 20 µm square aperture
• 8.0 sample gain,
• 4.7468 (cm/s) mirror velocity and
• The Nominal spectral resolution was 4 cm-1

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FT-IR Data Collection

• The background spectrum was recorded at a
  position of the IR window outside the algae
  sample with 256 scans.

• Three to four spectra were collected for each
  single cell of algae with 256 scans for each
  spectra.

• Data acquisition was carried out by means of
  Widows 2000 based OMNIC software package supplied
  by Thermo Nicolet.

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The Algae Family
Green Algae
Cynaobacteria
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The Algae Family
Diatom
Dino flaggelate
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Infrared Spectrum from Algae Chemical Standards
Giordano, et al., J. of Phycology, 37, 271
(2001)
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IR Spectra of Micrasterius Algae

• Numbers at the peaks show the location of the
  maximum peak

Amide I
Lipids
Amide II
Phospholipids
Carbohydrates
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Algae Samples Used
For the Algae family under study

• Samples are prepared in two forms
• Samples without fixation
• Samples with fixation (Lugol)

The feeding medium for the algae was NO3 and
NH4 With three different concentrations
Preparation of the samples was done at Marine
Sciences Dept. University of Ancona ITALY.
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Agglomerative Hierarchical (AH) clustering
(Wards technique)
The effect of Lugol in the Spectral region ( 1000
to 1500 ) cm-1
1 2 3 4 5 6
1 2 3 4 5 6
1 2 3 4 5 6
1 2 3 4 5 6
1 2 3 4 5 6
Results are skipped from this talk you might
contact me later at zbayyari_at_rocketmail.com zbayy
ari_at_uwm.edu
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Summary of IR Microspectroscopy (Ir?s) ?

• The IR spectra provide relative concentrations of
  lipids, proteins, carbohydrates , and silica in
  the specimen.

• The IR microspectroscopy demonstrates the
  potential to examine changes in the Chemistry of
  living cells.

• FTIR spectromicroscopy, in conjunction with
  clustering methods (Chemometrics) can
  consistently discriminates between different
  types of algae.

• Future outlook
• Development of environmental chamber to examine
  changes in the Chemistry of living cells while
  modifying environmental stimuli.

• Extend the chemometric study to include more
  algal species.

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Summary of IR Microspectroscopy (Ir?s) ?

• To investigate the applicability of FTIR in
  biomedical sciences, Including the pathology of
  fatal group of neurodegenerative disorder
  diseases by discriminating between healthy and
  infected tissues like.
• Cancers.
• Liver damage.
• Transmissible spongiform encephalopathies (TSE),
  such as scrapie in sheep and goats.
• Bovine spongiform encephalopathy in cattle.
• Creutzfeldt-Jakob disease in humans.

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Acknowledgements

• Collaborators

1- Prof. Carol Hirschmugl- Department of Physics.
(University of Wisconsin Milwaukee - USA).

2- Prof. Mario Giordano- Institute of Marine
Sciences. (University Del La Marche- Ancona -
Italy).
3- Stefano Mattioni-PhD. Student Dept. of
Biology (University of Ancona - Italy).
4- Prof. Craig Sandgren- Department of Biology.
(University of Wisconsin Milwaukee - USA).

• Funding

• NSF

Under Grant Number DMR-0084402 and CHE 9984931
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End of The Talk
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An Invitation to Visit Jordan
The Khazneh, Petra, of Jordan. It is carved out
from the sandstone cliff wall, is 40m high, and
is remarkably well-preserved. Thank You very much
for your kind attention And for the opportunity
to present some of my work at The 3rd Sesame
Users Meeting