Joseph J. Katz

Spectral absorption properties of ordinary and fully deuteriated chlorophylls a and b

The spectral absorption properties of fully deuteriated chlorophylls a and b, ordinary chlorophylls a and b, and methyl chlorophyllides a and b, isolated in a state of high purity, have been compared. Substitution of phytyl by methyl did not affect the wavelengths of the absorption maxima or the ratio of the absorption at the principal maxima. Deuteriation reduced the absorption in proportion to the molecular weight of the deuteriated pigments. Deuteriation also shifted the absorption maxima to slightly shorter wavelengths, an effect attributable primarily to the 6 deuterium atoms attached to carbon atoms in the conjugated system.

Proton Magnetic Resonance of Proteins Fully Deuterated except for 1H-Leucine Side Chains

The fully deuterated proteins C-phycocyanin, C-phycoerythrin, and cytochrome c have been obtained by biosynthesis with the leucine side chains, and only the leucine side chains, of normal (1H) isotopic composition. In these (isotopic hybrid) proteins, proton magnetic resonance analysis shows that the 1H-leucine side chains are in a variety of environments. During protein biosynthesis, the alpha hydrogen of leucine is exchanged with a hydrogen (2H) from the aqueous medium.

Evidence for 5- and 6-coordinated magnesium in bacteriochlorophyll a from visible absorption spectroscopy☆

The visible absorption spectrum of bacteriochlorophyll a (Bchl) in the 560-620 nm (yellow) region can be used to establish the coordination state of the central Mg atom. Five-coordinated Mg species absorb near 580 nm, whereas 6-coordinated Mg species are red-shifted to about 610 nm. Five-coordination is confirmed to be the principal coordination state of Mg in self-aggregated bacteriochlorophyll. The equilibrium constant for the reaction Bchl - Py + Py equilibrium Bchl - Py2 has been determined from computer-assisted analyses of spectral data, where Py represents pyridine. The spectral criteria for the coordination state of Mg in bacteriochlorophyll advanced here are shown to be applicable to both in vitro and in vivo systems. Similar spectral behavior is exhibited by chlorophylls a and b, and a band at 633 nm is shown to be associated with the presence of 6-coordinated Mg in chlorophyll a.

STUDIES OF CHLOROPHYLL‐CHLOROPHYLL AND CHLOROPHYLL‐LIGAND INTERACTIONS BY VISIBLE ABSORPTION AND INFRARED SPECTROSCOPY AT LOW TEMPERATURES

Abstract— A comparison of the visible absorption and infrared spectra of various chlorophyll‐chlorophyll (Chl) and Chi‐nucleophile aggregates at room temperature and at low temperatures has been made. The IR data provide structural information indispensable for the interpretation of the visible spectra. As a necessary preliminary, it is shown that Chl a solutions in nonpolar solvents can be prepared by appropriate drying techniques that contain at a conservative estimate ≤ 3 mol % of water (i.e. Chl a/H2O > 30:1). Very dry solutions of Chl a or Pyrochl a(≥ 10 mM) in toluene or methylcyclohexane‐isopentane solution show only slight changes in visible spectra on cooling to 77 K. From IR, additional Chl‐Chl aggregation occurs on cooling in methylcyclohexane‐isopentane but not to a significant extent in toluene. Dilute (10 μM) solutions of Chl a or Pyrochl a in nonpolar solvents form a new absorption peak near 700 nm at low temperatures, which we attribute to traces of water in the solvent or other residual nucleophiles not removed during the Chl purification. Addition of stoichiometric amounts of water increases the size of the ˜700 nm peak even in dilute Chl solutions. Chlorophyll a, Pyrochl a, but not pheophytin a are shown to interact with nucleophiles of the general type RXH (where R= H or alkyl, and X = O, N, or S). Such nucleophiles can coordinate to the Mg atom of one Chl molecule by lone pairs on O, N, or S, and hydrogen bond to oxygen donor functions in another Chl molecule. A ˜0.1 M solution of Chl a or Pyrochl a in toluene containing 1.5 equivalents of ethanol is converted almost entirely to a species absorbing at ˜700 nm at 77 K. Infrared spectroscopy shows conclusively that it is the keto C=O function that is involved in the cross‐linking by hydrogen bonding, a conclusion supported by the observation that Pyrochl a forms a very similar red‐shifted species at low temperatures, despite the absence of a carbomethoxy C=O function. n‐Butylamine and ethanethiol interact in much the same way as does ethanol to form species red shifted to ˜700 nm. A variety of possible structures for the low temperature forms is discussed, and the use of these red shifted species as paradigms for photoreaction center Chl is described.

Chlorophyll and Light Energy Transduction in Photosynthesis1

Publisher Summary Photosynthetic organisms use the energy of light quanta to convert the inorganic compounds carbon dioxide and water to the organic compounds required for survival by all other living organisms. Thus, the transformation of the energy of light quanta into oxidants and reductants useful for chemical synthesis assumes central importance in biology. In the sequence of events that begins with the absorption of light and culminates in the vast array of organic compounds characteristic of living organisms, chlorophyll plays a crucial role. The chlorophylls constitute a small family of closely related pigments long recognized as the primary photo-acceptors of photosynthetic organisms. Most of what is known or inferred about the function of chlorophyll in the plant is based on absorption and fluorescence spectroscopy in the visible. Consequently, an appreciation of the nuances of chlorophyll electronic transition spectra both in vitro and in vivo is essential. In all photosynthetic organisms, the main long wavelength absorption band in the red is due to chlorophyll

Chlorophyll-chlorophyll and chlorophyll-water interactions in the solid state☆

Abstract Absorption spectra in the visible and infrared have been recorded for films of dry chlorophyll a , chlorophyll b , bacteriochlorophyll, pyrochlorophyll a , and pheophytin a . The effects of water on the spectra have also been studied. From the infrared it is possible to deduce structures for the chlorophyll species involved, and to correlate the visible absorption spectra of both chlorophyll oligomers and chlorophyll-water adducts with particular chlorophyll species. Hydration may cause large red shifts in the visible absorption spectra, probably as a result of changes in the orientation of the chlorophyll molecules relative to each other.

State of chlorophyll a in vitro and in vivo from electronic transition spectra, and the nature of antenna chlorophyll☆

Abstract The computer deconvolution of the red envelopes of visible spectra of chlorophyll a solutions in dry nonpolar solvents is described. In solvents such as carbon tetrachloride and benzene chlorophyll a is present in dimer form in the concentration range of 10−6–10−2. In solvents such as hexane and n-alkanes oligomeric species of chlorophyll a are present. The relative intensities of the Gaussian components used in the spectral deconvolution can be related to the size and conformation of the chlorophyll a aggregates. While the computer deconvolution is basically a heuristic method, it is suggested that it is based on a sound physical rationale. A comparison of the computer deconvoluted visible spectra of photosynthetic organisms with those of solutions of chlorophyll a oligomers in nonpolar media provides good evidence that antenna chlorophyll in green plants has spectral properties that are very similar, if not identical, to those of chlorophyll a oligomers, (chlorophyll2)n.

HIGH-PERFORMANCE LIQUID CHROMATOGRAPHIC STUDY OF THE CHLOROPHYLL ALLOMERIZATION REACTION*

Abstract The allomerization of chlorophyll has been investigated by reversed-phase HPLC. Four major product peaks (two sets of doublets) are observed in the HPLC chromatogram. These are shown to be pairs of stereoisomers of two major components. Similar results have been obtained from the allomerization of fully deuterated chlorophyll a. Attempts to characterize the reaction products and to delineate the reaction mechanism were initially studied by the oxidation of chlorophyll in the absence of extraneous nucleophiles. The structure of the reaction products of chlorophyll allomerization were conclusively identified as 10-hydroxychlorophyll and the 10-methoxylactone by co-chromatography, NMR, and 252Cf plasma-desorption mass spectroscopy. Exploratory studies on the allomerization products of bacteriochlorophyll have also been carried out.

Aggregation of chlorophyll in nonpolar solvents from molecular weight measurements

The state of aggregation of chlorophylls a and b, pyrochlorophyll a, bacteriochlorophyll, pheophytin a and pyropheophytin a in a variety of nonpolar solvents has been deduced from molecular weight measurements made by vapor-phase osmometry. In carbon tetrachloride and benzene solution, chlorophyll a and pyrochlorophyll a exist predominately as dimers over a wide concentration range, although there is evidence for oligomer formation in concentrated solutions. In aliphatic or cycloaliphatic hydrocarbon solvents, chlorophyll a can form oligomers with aggregation numbers as large as 20. In even the most dilute cyclohexane solutions studied, chlorophyll a exists as a tetramer. Chorophyll b and bacteriochlorophyll aggregate more strongly. The basic unit for these tow chlorophyll is the trimer which has a strong tendency to form hexamer and higher oligomers in aliphatic hydrocarbon solvents. Infrared spectra for solutions of chlorophyll of known states of aggregation are presented.

Loroxanthin, a unique xanthophyll from scenedesmus obliquus and chlorella vulgaris☆

Abstract A unique xanthophyll, which had been detected before in certain green algae, has now been isolated from Scenedesmus obliquus and Chlorella vulgaris . This pigment, here called loroxanthin, has also been isolated in its deuterated form from fully deuterated Chlorella . It forms a triacetate, and with methanol plus HCl, it yields monomethyl and dimethyl ethers. It can be oxidized to an aldehyde, loroxanthal. It has been characterized as a hydroxy lutein with the additional hydroxy group on a chain methyl group, probably that on C 9 . The previously reported absence of this pigment in C. pyrenoidosa has been confirmed. It was present in two marine Cladophora species and in Ulva rigida but absent in Spirogyra sp. and in two marine siphonalean green algae. It is probably not identical with certain similar pigments reported in other vegetable sources and variously described as trollein and trollein-like.