312
the areas of high pigment intensity in the Euglena and Chlamydomonas cells. Both
for Euglena and Chlamydomonas, the highest abundance of carotenoids were found
in the eyespot, however, a pigment was also observed in other areas of the cell, most
probably in chloroplasts. In Chlamydomonas, due to known birefringence of its eye,
the eyespot was measured with the polarized light, showing that pigment molecules
are arranged with the long axis along the elongated direction of the eyespot and
parallelly to the body axis [31].
Raman and fluorescence microspectroscopy, in addition to matrix-free laser
desorption/ionization mass spectrometry were applied to the analysis of composition and distribution of photosynthetic apparatus components in a single cell of
Euglena gracilis and Chlamydomonas reinhardtii [32]. high-spatial resolution
( ca. 550 nm) Raman imaging combined with fluorescence imaging allowed for
simultaneous visualization of β-carotene and protoplastids distribution in a single
algal cell.
In the light of increasing interest in the application of algae as biofuel, two algal
species, a potential biofuel source, Chlorella sorokiniana and Neochloris oleoabundans, were investigated with the help of Raman microimaging [33]. the compositional analysis of healthy and nitrogen-starved algae indicated for the presence of an
unspecified carotenoid in their cells, accompanied with chlorophyll and triglyceride
in the case of nitrogen-starved algae. Raman distribution maps showed that the unspecified carotenoid is located specifically within a chloroplast [33].
Laser tweezers Raman spectroscopy (LtRS) was applied to study the carotenogenesis process in a single live cell of yeast Rhodotorula glutinis [20]. the LtRSbased analysis of the kinetics of total carotenoid accumulation inside individual
Rhodotorula glutinis cells demonstrated a significant variation of the carotenoid
level in the studied population of 100 cells, particularly for certain growth phases.
this variation indicates for high cellular heterogeneity in the studied cellular population, and provide for better understanding of carotenogenesis not only in the bulk
sample, but for individual cells [20].
Raman microspectroscopy was also demonstrated to be a suitable tool to quantify carotenoids inside subcellular structures. β-carotene quantification in lipid bodies
of the alga Trachydiscus minutus in vivo was assessed and proposed to be a method
for noninvasive cell sorting of algal strains [21]. Calculated β-carotene absolute
concentrations were correlated with lipid bodies volumes, showing high variability
between the individual specimens.
Spontaneous Raman spectroscopy was also combined with Coherent anti-Stokes
Raman Spectroscopy (CARS) to study carotenoid-related lipid accumulation under
nitrogen-repletion conditions in a green microalga Coccomyxa subellipsoidea [34].
As mentioned above, such research are of growing interest due to potential industrial usage of Raman spectroscopy as a tool for quantification of biofuels or chemicals
in algal cells. As expected, both spontaneous Raman and CARS imaging demonstrated that the lipid (triacylglycerols) level increased in N-depleted algae compared
with control algal cells containing more carotenoids and less lipids. In the case of
the applied equipment, CARS spectroscopy was demonstrated to possess higher
scanning speed and spatial resolution (< 500 nm) [34], although it is necessary to
A. Kaczor and m. Pilarczyk
the areas of high pigment intensity in the Euglena and Chlamydomonas cells. Both
for Euglena and Chlamydomonas, the highest abundance of carotenoids were found
in the eyespot, however, a pigment was also observed in other areas of the cell, most
probably in chloroplasts. In Chlamydomonas, due to known birefringence of its eye,
the eyespot was measured with the polarized light, showing that pigment molecules
are arranged with the long axis along the elongated direction of the eyespot and
parallelly to the body axis [31].
Raman and fluorescence microspectroscopy, in addition to matrix-free laser
desorption/ionization mass spectrometry were applied to the analysis of composition and distribution of photosynthetic apparatus components in a single cell of
Euglena gracilis and Chlamydomonas reinhardtii [32]. high-spatial resolution
( ca. 550 nm) Raman imaging combined with fluorescence imaging allowed for
simultaneous visualization of β-carotene and protoplastids distribution in a single
algal cell.
In the light of increasing interest in the application of algae as biofuel, two algal
species, a potential biofuel source, Chlorella sorokiniana and Neochloris oleoabundans, were investigated with the help of Raman microimaging [33]. the compositional analysis of healthy and nitrogen-starved algae indicated for the presence of an
unspecified carotenoid in their cells, accompanied with chlorophyll and triglyceride
in the case of nitrogen-starved algae. Raman distribution maps showed that the unspecified carotenoid is located specifically within a chloroplast [33].
Laser tweezers Raman spectroscopy (LtRS) was applied to study the carotenogenesis process in a single live cell of yeast Rhodotorula glutinis [20]. the LtRSbased analysis of the kinetics of total carotenoid accumulation inside individual
Rhodotorula glutinis cells demonstrated a significant variation of the carotenoid
level in the studied population of 100 cells, particularly for certain growth phases.
this variation indicates for high cellular heterogeneity in the studied cellular population, and provide for better understanding of carotenogenesis not only in the bulk
sample, but for individual cells [20].
Raman microspectroscopy was also demonstrated to be a suitable tool to quantify carotenoids inside subcellular structures. β-carotene quantification in lipid bodies
of the alga Trachydiscus minutus in vivo was assessed and proposed to be a method
for noninvasive cell sorting of algal strains [21]. Calculated β-carotene absolute
concentrations were correlated with lipid bodies volumes, showing high variability
between the individual specimens.
Spontaneous Raman spectroscopy was also combined with Coherent anti-Stokes
Raman Spectroscopy (CARS) to study carotenoid-related lipid accumulation under
nitrogen-repletion conditions in a green microalga Coccomyxa subellipsoidea [34].
As mentioned above, such research are of growing interest due to potential industrial usage of Raman spectroscopy as a tool for quantification of biofuels or chemicals
in algal cells. As expected, both spontaneous Raman and CARS imaging demonstrated that the lipid (triacylglycerols) level increased in N-depleted algae compared
with control algal cells containing more carotenoids and less lipids. In the case of
the applied equipment, CARS spectroscopy was demonstrated to possess higher
scanning speed and spatial resolution (< 500 nm) [34], although it is necessary to
A. Kaczor and m. Pilarczyk