Direct injection of pigment–protein complexes and membrane fragments suspended in water from phototrophs to C18 HPLC
- 20 March 2020
- journal article
- research article
- Published by Springer Science and Business Media LLC in Photosynthesis Research
- Vol. 144 (1), 101-107
- https://doi.org/10.1007/s11120-020-00735-w
Abstract
We discovered that pigments including carotenoids and (bacterio)chlorophylls in pigment–protein complexes, membrane fragments, and chlorosomes suspended in water could be injected directly into C18 HPLC and analyzed without any other treatments. We applied this method to LH1-RC and chromatophores of purple bacteria, chlorosomes of green sulfur bacteria, thylakoid membranes of cyanobacteria, and PSII and thylakoid membranes of spinach. HPLC elution profiles and pigment composition were the same as those of the conventional extraction method. The principle of this method might be that samples are first trapped on top of column, followed by the immediate extraction of the pigments with the HPLC eluent and their separation using the C18 column, as usual. In the conventional extraction method, pigments are first extracted with organic solvents, followed by evaporation of the solvents. The dried pigments are then dissolved in organic solvents and injected into C18 HPLC after filtration. The advantages of this method include the preventions of pigment isomerization and oxidation and the possibility of injecting all samples. Its drawbacks include the accumulation of denatured proteins at the top of column, causing increased HPLC pressure. The use of a guard column might solve this problem. Many factors, such as samples, column, and HPLC systems, may affect this method. Nevertheless, we think that some samples can be analyzed using this method.Keywords
This publication has 15 references indexed in Scilit:
- Effects of Calcium Ions on the Thermostability and Spectroscopic Properties of the LH1-RC Complex from a New Thermophilic Purple Bacterium Allochromatium tepidumThe Journal of Physical Chemistry B, 2017
- A Novel Astaxanthin-Binding Photooxidative Stress-Inducible Aqueous Carotenoprotein from a Eukaryotic Microalga Isolated from Asphalt in MidsummerPlant and Cell Physiology, 2013
- Synechoxanthin, an Aromatic C40 Xanthophyll that Is a Major Carotenoid in the Cyanobacterium Synechococcus sp. PCC 7002Journal of Natural Products, 2008
- Accumulation of chlorophyllous pigments esterified with the geranylgeranyl group and photosynthetic competence in the CT2256-deleted mutant of the green sulfur bacterium Chlorobium tepidumPhotochemical & Photobiological Sciences, 2008
- DNA Microarray Analysis of Cyanobacterial Gene Expression during Acclimation to High LightTHE PLANT CELL ONLINE, 2001
- Characterization of carotenes in a combination of a C18 HPLC column with isocratic elution and absorption spectra with a photodiode-array detectorPhotosynthesis Research, 2000
- Pigment Composition in the Reaction Center Complex from the Thermophilic Green Sulfur Bacterium, Chlorobium tepidum: Carotenoid Glucoside Esters, Menaquinone and ChlorophyllsPlant and Cell Physiology, 1999
- [35] Characterization of carotenoids in photosynthetic bacteriaPublished by Elsevier BV ,1992
- Chromatium tepidum sp. nov., a Thermophilic Photosynthetic Bacterium of the Family ChromatiaceaeInternational Journal of Systematic and Evolutionary Microbiology, 1986
- Inactivation of Photosynthetic Oxygen Evolution and Concomitant Release of Three Polypeptides in the Photosystem II Particles of Spinach ChloroplastsPlant and Cell Physiology, 1982