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Spectral analysis combined with advanced linear unmixing allows for histolocalization of phenolics in leaves of coffee trees.

Conéjéro G, Noirot M, Talamond P, Verdeil JL - Front Plant Sci (2014)

Bottom Line: Lastly, young leaves of Coffea pseudozanguebariae (PSE), C. eugenioides (EUG), C. arabica (ARA) and C. canephora (CAN) were compared.This confirmed the presence of xanthone in PSE and EUG, but especially its precise tissue localization.This non-invasive optical technique does not require pretreatment and is an effective experimental tool to differentiate multiple naturally-occuring fluorochores in living tissues.

View Article: PubMed Central - PubMed

Affiliation: Plant Cell Imaging platform PHIV UMR AGAP (Cirad, SupAgro, INRA), UMR B&PMP (INRA, CNRS, UM2, SupAgro) Montpellier, France.

ABSTRACT
An imaging method using spectral analysis combined with advanced linear unmixing was used to allow histolocalization of natural autofluorescent compounds such as hydroxycinnamic acid (chlorogenic acid) and xanthone (mangiferin) in living cells and tissues (mature coffee leaves). The tested method included three complementary steps: 1/ visualization of natural autofluorescence and spectrum acquisition with a multiphoton microscope; 2/ identification of some compounds using previous information on the chemical composition of the tissue, obtained from litterature; and 3/ localization of candidate compounds by spectral imaging. The second part of the study consisted of describing the histochemical structure of leaves during their development. This revealed very fast histochemical differentiation of leaves during the first week after their emergence. Lastly, young leaves of Coffea pseudozanguebariae (PSE), C. eugenioides (EUG), C. arabica (ARA) and C. canephora (CAN) were compared. This confirmed the presence of xanthone in PSE and EUG, but especially its precise tissue localization. This also highlighted the paternal CAN origin of the leaf structure in the allotetraploid species ARA. The limits and advantages of the method without staining are discussed relative to classical epifluorescence microscopy under UV light. This non-invasive optical technique does not require pretreatment and is an effective experimental tool to differentiate multiple naturally-occuring fluorochores in living tissues.

No MeSH data available.


Reference spectrum of controls (5-CQA, chlorophyll, and mangiferin purified porducts) obtained with 720 nm excitation using multiphoton microscope emission from 365 to 700 nm.
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Figure 3: Reference spectrum of controls (5-CQA, chlorophyll, and mangiferin purified porducts) obtained with 720 nm excitation using multiphoton microscope emission from 365 to 700 nm.

Mentions: Reference spectral signatures were acquired on the microscope using controls of products known to be present in mature leaves, i.e., pure 5-CQA and mangiferin powders and chlorophyll extracts. Figure 3 represents the reference spectra of 5-CQA, mangiferin and chlorophyll. Each molecule could be characterized by its absorption and emission spectra. Spectral signatures of 5-CQA, mangiferin, and chlorophyll were then stored in the Spectra Database. The emission spectrum of 5-CQA obtained with the laser system had the same profile and λmax emission (457 nm) as that obtained with a conventional spectofluorometer with a 300 nm excitation wavelength. Spectral signature of chlorophyll presented λmax at 670 nm, as expected. 5-CQA and mangiferin presented a wide spectral range with peaks at about 455 and 590 nm, respectively.


Spectral analysis combined with advanced linear unmixing allows for histolocalization of phenolics in leaves of coffee trees.

Conéjéro G, Noirot M, Talamond P, Verdeil JL - Front Plant Sci (2014)

Reference spectrum of controls (5-CQA, chlorophyll, and mangiferin purified porducts) obtained with 720 nm excitation using multiphoton microscope emission from 365 to 700 nm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3927124&req=5

Figure 3: Reference spectrum of controls (5-CQA, chlorophyll, and mangiferin purified porducts) obtained with 720 nm excitation using multiphoton microscope emission from 365 to 700 nm.
Mentions: Reference spectral signatures were acquired on the microscope using controls of products known to be present in mature leaves, i.e., pure 5-CQA and mangiferin powders and chlorophyll extracts. Figure 3 represents the reference spectra of 5-CQA, mangiferin and chlorophyll. Each molecule could be characterized by its absorption and emission spectra. Spectral signatures of 5-CQA, mangiferin, and chlorophyll were then stored in the Spectra Database. The emission spectrum of 5-CQA obtained with the laser system had the same profile and λmax emission (457 nm) as that obtained with a conventional spectofluorometer with a 300 nm excitation wavelength. Spectral signature of chlorophyll presented λmax at 670 nm, as expected. 5-CQA and mangiferin presented a wide spectral range with peaks at about 455 and 590 nm, respectively.

Bottom Line: Lastly, young leaves of Coffea pseudozanguebariae (PSE), C. eugenioides (EUG), C. arabica (ARA) and C. canephora (CAN) were compared.This confirmed the presence of xanthone in PSE and EUG, but especially its precise tissue localization.This non-invasive optical technique does not require pretreatment and is an effective experimental tool to differentiate multiple naturally-occuring fluorochores in living tissues.

View Article: PubMed Central - PubMed

Affiliation: Plant Cell Imaging platform PHIV UMR AGAP (Cirad, SupAgro, INRA), UMR B&PMP (INRA, CNRS, UM2, SupAgro) Montpellier, France.

ABSTRACT
An imaging method using spectral analysis combined with advanced linear unmixing was used to allow histolocalization of natural autofluorescent compounds such as hydroxycinnamic acid (chlorogenic acid) and xanthone (mangiferin) in living cells and tissues (mature coffee leaves). The tested method included three complementary steps: 1/ visualization of natural autofluorescence and spectrum acquisition with a multiphoton microscope; 2/ identification of some compounds using previous information on the chemical composition of the tissue, obtained from litterature; and 3/ localization of candidate compounds by spectral imaging. The second part of the study consisted of describing the histochemical structure of leaves during their development. This revealed very fast histochemical differentiation of leaves during the first week after their emergence. Lastly, young leaves of Coffea pseudozanguebariae (PSE), C. eugenioides (EUG), C. arabica (ARA) and C. canephora (CAN) were compared. This confirmed the presence of xanthone in PSE and EUG, but especially its precise tissue localization. This also highlighted the paternal CAN origin of the leaf structure in the allotetraploid species ARA. The limits and advantages of the method without staining are discussed relative to classical epifluorescence microscopy under UV light. This non-invasive optical technique does not require pretreatment and is an effective experimental tool to differentiate multiple naturally-occuring fluorochores in living tissues.

No MeSH data available.