INVESTIGATION OF THE ALLOMERIZATION REACTION OF CHLOROPHYLL a: USE OF DIODE ARRAY HPLC, MASS SPECTROMETRY AND CHEMOMETRIC FACTOR ANALYSIS FOR THE DETECTION OF EARLY PRODUCTS

Abstract –The products of chlorophyll allomerization in methanol were isolated and analyzed by open column sucrose chromatography, liquid chromatography mass spectrometry (LCMS) and DAD‐HPLC (diode‐array high‐performance liquid chromatography). Four main bands were found with molecular ions of (a) 908, (b) 938, (c) 938 and (d) 938, consistent with the structures (a) 132‐hydroxy‐chlorophyll a (II), (b) and (c) Mg(II)‐31,32‐didehydro‐151‐hydroxy‐151‐methoxy‐rhodochlorin‐15 acetic acid δ‐lactone 152‐methyl 173‐phytyl ester and its epimer (III) and (d) Mg(II)‐31,32‐didehydro‐rhodochlorin‐15‐glyoxylic acid 131,152‐dimethyl 173‐phytyl ester (IV), evidence enhanced by UV/visible spectroscopy, chromatographic coelutions and chemometrics. Chlorophyll a was degraded both in the dark and light, under O2 and N2. DAD‐HPLC of the resultant degradation mixtures were analyzed using the chemometric heuristic‐evolving latent projection method for resolution. Ultraviolet/visible spectra of II and III are reproducibly extracted from the mixtures after a short degradation time, whereas III and IV are the dominant compounds after longer degradation times. Changes in relative elution order of IV using open column chromatography and reverse‐phase HPLC are established. A possible allomerization pathway is proposed.

[1]  Paul Geladi,et al.  Principal Component Analysis , 1987, Comprehensive Chemometrics.

[2]  Patricia L. Smith.  Chemometrics: Applications of Mathematics and Statistics to Laboratory Systems , 1993 .

[3]  R. Brereton,et al.  THE USE OF LIQUID CHROMATOGRAPHY‐MASS SPECTROMETRY TO MONITOR THE ALLOMERIZATION REACTIONS OF CHLOROPHYLL a and PHEOPHYTIN a:m IDENTIFICATION OF THE ALLOMERS OF PHEOPHYTIN a , 1993 .

[4]  R. Brereton,et al.  Resolution of strongly overlapping two‐way multicomponent data by means of heuristic evolving latent projections , 1993 .

[5]  J. A. López del Val,et al.  Principal Components Analysis , 2018, Applied Univariate, Bivariate, and Multivariate Statistics Using Python.

[6]  Yizeng Liang,et al.  Heuristic evolving latent projections: resolving two-way multicomponent data. 1. Selectivity, latent-projective graph, datascope, local rank, and unique resolution , 1992 .

[7]  M. Pavel,et al.  Mathematics in pattern recognition , 1992 .

[8]  N. Bratchelf,et al.  Chapter 4 The Mathematics of Pattern Recognition , 1992 .

[9]  Paul J. Lewi,et al.  Chapter 2 Multivariate Data Display , 1992 .

[10]  S. Schwartz,et al.  High-performance liquid chromatography—continuous-flow fast atom bombardment mass spectrometry of chlorophyll derivatives , 1991 .

[11]  Paul J. Gemperline,et al.  Mixture analysis using factor analysis I: Calibration and quantitation , 1989 .

[12]  Stanley B. Brown,et al.  THE DEGRADATION OF CHLOROPHYLL - A BIOLOGICAL ENIGMA. , 1987, The New phytologist.

[13]  J. Katz,et al.  High-performance liquid chromatographic study of the chlorophyll allomerization reaction , 1984 .

[14]  C. Llewellyn,et al.  The rapid determination of algal chlorophyll and carotenoid pigments and their breakdown products in natural waters by reverse-phase high-performance liquid chromatography , 1983 .

[15]  R. Brereton,et al.  Positive and negative ion fast atom bombardment mass spectrometric studies on chlorophylls: structure of 4-vinyl-4-desethyl chlorophyll B , 1983 .

[16]  R. Brereton,et al.  4-vinyl-4-desethyl chlorophyll a: characterisation of a new naturally occurring chlorophyll using fast atom bombardment, field desorption and “in beam”electron impact mass spectroscopy , 1982 .

[17]  R. Brereton,et al.  “In beam” electron impact mass spectrometry: the structure of a bacteriochlorophyll allomer , 1980 .

[18]  W. A. Svec.  8 – The Isolation, Preparation, Characterization, and Estimation of the Chlorophylls and the Bacteriochlorophylls , 1978 .

[19]  K. Iriyama,et al.  A simple method for extraction and partial purification of chlorophyll from plant material, using dioxane. , 1974, Journal of biochemistry.

[20]  M. E. Ruiz,et al.  Chlorophylls. II. Allomerization of Chlorophylls a and b. , 1973 .