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.

Principal components scores and loadings plots for visualisation of the electrospray ionisation liquid chromatography mass spectra of a mixture of chlorophyll degradation products at different cone voltages

“…Chlorophyll allomers have been characterised in previous papers. 11,15,23,24 The next phase is to scale the data as described above, and then perform PCA, retaining the three most significant principal components. Three-dimensional (3-D) plots of scores and loadings were generated, and rotated to emphasise the most important features.…”

“…Chlorophyll a was extracted from spinach according to the methods used by Svec, 13 Iriyama et al 14 and Brereton et al 11 Pure chlorophyll a was dissolved in methanol, and subjected to illumination at 4000 lux in a water bath at 40°C in the presence of oxygen. 10 After 60 min, a sample was taken and methanol was removed in vacuo to produce a sample that was stored as a residue for subsequent analysis.…”

Chemometrics and modernization of traditional Chinese medicine

“…The results were satisfactory, which will be a good basis for exploring the chemical background of TCMs and for further investigating working mechanisms of TCMs at the molecular level [45][46][47][48][49] . In fact, the multivariate resolution methods have been developed rapidly in recent years [50][51][52][53][54][55][56][57][58][59][60][61] and used to solve some difficult problems [62][63][64][65][66][67] . However, there is a long distance for us to completely solve the problems concerning on-line qualitative and quantitative analysis and structural identification for complex systems [68] .…”

Application of combined approach to analyze the constituents of essential oil from Dong quai

“…Many associated methods have also been developed to provide more information for chemical analysis both in chromatographic separation and in spectral identification, which makes it possible to interpret these complex systems [5,6,7,8,9,10,11,12,13,14,15,16,17,18].…”