chlorophyll-a and chlorin

The rapid growth of drug-resistant bacteria all over the world has given rise to a major research challenge, namely a search for alternative treatments to which bacteria will be unable to develop resistance. Photodynamic therapy is an approach of this kind. It involves the use of photosensitizers in combination with visible light at a certain wavelength to excite the former and generate reactive oxygen species. Various synthetic heterocyclic compounds are used as photosensitizers. Of these, derivatives of natural chlorophylls have a special place due to their properties. This review deals with the use of such compounds in antimicrobial PDT.

Topics: Amino Acid Sequence; Anti-Bacterial Agents; Cations; Chlorophyll; Photochemotherapy; Porphyrins

Chlorophylls make Earth green, are the central constituents in the engine of photosynthesis, and not surprisingly have garnered immense attention. A chlorin, the core chromophore of a chlorophyll, is a dihydroporphyrin macrocycle that contains one pyrroline ring and three pyrrole rings. The dominant method for the synthesis of chlorins has entailed the derivatization of porphyrins. The present review covers the ostensibly simple conversion of porphyrins, regardless of synthetic or biological origin, to chlorins. The period covered encompasses the entire history since the beginnings of chlorin synthetic chemistry in the early 20th century through 2015. Representative transformations include hydrogenation, cycloaddition, annulation, and diverse "breaking and mending" approaches. Altogether, the synthesis of >1000 chlorins or chlorin-like compounds (containing >50 distinct pyrroline motifs) is described. Such diversity animates the question "what structural features are essential for a chlorin to resemble chlorophyll?" To begin to address the structure-spectrum relationship, > 250 absorption spectra are provided for representative structures. The synthesis and spectral properties of the vast collection of compounds described herein are expected to illuminate the scope to which synthetic chlorins can serve as surrogates for chlorophylls and be exploited in diverse ways.

Topics: Chlorophyll; Porphyrins; Stereoisomerism

Topics: Chlorophyll; Molecular Structure; Porphyrins

Topics: Amino Acids; Carcinogens; Chlorophyll; Coenzymes; DNA; Glucagon; Luminescent Measurements; Nucleic Acids; Peptides; Porphyrins; Proteins; Spectrum Analysis

C3-(Trans-2-arylethenyl)carbonylated chlorophyll derivatives possessing a bacteriochlorin or chlorin π-system were synthesized by cross-aldol (Claisen-Schmidt) condensation of methyl pyrobacteriopheophorbide-a or 3-acetyl-3-devinyl-pyropheophorbide-a bearing the C3-acetyl group with p-(un)substituted benzaldehydes under basic conditions. The corresponding porphyrin-type chlorophyll derivatives were prepared by the oxidation (17,18-didehydrogenation) of the chlorin-type. Their Qy absorption and fluorescence emission maxima in dichloromethane correlated well with Hammett substituent constants of the p-substituents. Several electron-withdrawing p-substituents suppressed the emission due to photoinduced electron transfer quenching in a molecule. The substitution sensitivities for their maxima and fluorescence quantum yields decreased in the order of bacteriochlorin-, chlorin- and porphyrin-type derivatives.

Topics: Chalcone; Chlorophyll; Mass Spectrometry; Porphyrins; Spectrometry, Fluorescence; Spectrophotometry, Ultraviolet

Zinc 3-hydroxymethyl-13

Topics: Alkylation; Bacterial Proteins; Bacteriochlorophylls; Chlorophyll; Circular Dichroism; Isomerism; Micelles; Models, Molecular; Porphyrins; Solutions; Water; Zinc

Topics: Avidin; Chlorophyll; Fluorescence; Fluorescent Dyes; Hydrophobic and Hydrophilic Interactions; Magnetic Resonance Spectroscopy; Mass Spectrometry; Micelles; Molecular Structure; Photochemistry; Polyethylene Glycols; Porphyrins; Solvents; Structure-Activity Relationship; Surface Properties; Water

Photodynamic therapy (PDT) is entering the mainstream of the cancer treatments recently. Pyropheophorbide-a (Pa), as a degradation product of chlorophyll-a, has been shown to be a potent photosensitizer in photodynamic therapy. In this paper, we investigated the in vitro photodynamic therapy of 13

Topics: Cell Survival; Chlorophyll; Dose-Response Relationship, Drug; HeLa Cells; Humans; Hydrazones; Photochemotherapy; Photosensitizing Agents; Porphyrins

Photodynamic therapy (PDT) is one of the most promising methods of specific cancer treatment. However, commercially available photosensitizers (PSs) show significant drawbacks, such as side toxicity, low penetration ability, low blood solubility, low tumor selectivity etc. In addition, as was shown previously, a conjugation of polyamines with several toxic agents led to an increased toxicity to cancer cells. Here, we synthesized conjugates of two chlorine photosensitizers, purpurin 18 and pheophorbide a, with spermine in natural and Boc-protected form. Using specialized software, we calculated octanol-water partition coefficients for single protonation state (logP) of single PSs and PS/spermine conjugates. We found that the addition of spermine to chlorine PSs shifted the logP towards higher hydrophilicity in comparison to logP of single chlorines. In vitro studies on several cancer cells indicated that conjugation of purpurin 18 with spermine increased its retention in cancer cells. Using various concentrations of this conjugate, we found that lower concentrations (under 0.2μM) of purpurin 18/spermine conjugate launched apoptosis in HeLa cells. This combined with its high phototoxicity makes the purpurin 18/spermine conjugate a promising photosensitizer for PDT. Obtained results might serve as a basis for further studies of this potential third-generation PS on mammalian models in vivo.

Topics: Animals; Apoptosis; Cell Line, Tumor; Chlorophyll; HeLa Cells; Humans; Hydrophobic and Hydrophilic Interactions; Photochemotherapy; Photosensitizing Agents; Porphyrins; Spermine

In the present study, a biomimetic reaction center model, that is, a molecular triad consisting of a chlorin dimer and an azafulleroid, is synthesized and its photophysical properties are studied in comparison with the corresponding molecular dyad, which consists only of a chlorin monomer and an azafulleroid. As evidenced by (1) H NMR, UV/Vis, and fluorescence spectroscopy, the chlorin dimer-azafulleroid folds in nonpolar media into a C2 -symmetric geometry through hydrogen bonding, resulting in appreciable electronic interactions between the chlorins, whereas in polar media the two chlorins diverge from contact. Femtosecond transient absorption spectroscopy studies reveal longer charge-separated states for the chlorin dimer-azafulleroid; ≈1.6 ns in toluene, compared with the lifetime of ≈0.9 ns for the corresponding chlorin monomer-azafulleroid in toluene. In polar media, for example, benzonitrile, similar charge-separated states are observed, but the lifetimes are inevitably shorter: 65 and 73 ps for the dimeric and monomeric chlorin-azafulleroids, respectively. Nanosecond transient absorption and singlet oxygen phosphorescence studies corroborate that in toluene, the charge-separated state decays indirectly via the triplet excited state to the ground state, whereas in benzonitrile, direct recombination to the ground state is observed. Complementary DFT studies suggest two energy-minima conformations, that is, a folded chlorin dimer-azafulleroid, which is present in nonpolar media, and another conformation in polar media, in which the two hydrophobic chlorins wrap the azafulleroid. Inspection of the frontier molecular orbitals shows that in the folded conformation, the HOMO on each chlorin is equivalent and is shared owing to partial π-π overlap, resulting in delocalization of the conjugated π electrons, whereas the wrapped conformation lacks this stabilization. As such, the longer charge-separated lifetime for the dimer is rationalized by both the electron donor-acceptor separation distance and the stabilization of the radical cation through delocalization. The chlorin folding seems to change the photophysical properties in a manner similar to that observed in the chlorophyll dimer in natural photosynthetic reaction centers.

Topics: Biomimetic Materials; Chlorophyll; Dimerization; Fullerenes; Models, Molecular; Photosynthesis; Porphyrins; Rhodopseudomonas

A series of chlorophyll a-based chlorins conjugated with pyridyl or quinoxalyl group at different positions were synthesized, characterized and evaluated for their photodynamic effect in vitro. It was found that all the pyridyl and quinoxalyl chlorins showed promising photocytotoxicities but nontoxic without irradiation in HeLa cells, and the substituted types and positions had a significant influence on the photocytotoxicities of the chlorophyll a-based chlorins. All the chlorins with a pyridyl group at the C-D ring end exhibited relatively high photocytotoxicity as compared to those with 3(2)-pyridyl. Among them, compound 12 conjugated with a pyridyl group at its C12 position showed the best photodynamic effect in HeLa cells with an IC50 value of 0.033μM. These facts, associated with the relative high long wavelength absorptions of those chlorins may provide valuable ways to design and prepare promising photosensitizers for application in photodynamic therapy.

Topics: Chlorophyll; Chlorophyll A; HeLa Cells; Humans; Neoplasms; Photochemotherapy; Photosensitizing Agents; Porphyrins; Pyridines; Quinoxalines

We developed novel methods to convert the C3-vinyl group of a chlorophyll derivative, methyl pyropheophorbide-a, into an acetyl group, an epoxy group, and a formyl group via iodination with I2 and phenyliodine(III) bis(trifluoroacetate). Reaction of the iodinated intermediate with ethylene glycol and subsequent treatment with base led to formation of the C3-acetyl chlorin. Reaction of the iodinated intermediate with ethylenediamine afforded the C3-oxiranyl chlorin. The C3-formyl chlorin was readily derived from the epoxide without hazardous reagents such as OsO4. These reactions were facile and useful alternatives to the previous methods.

Topics: Acetylation; Chlorophyll; Epoxy Compounds; Halogenation; Iodobenzenes; Methylation; Porphyrins; Trifluoroacetic Acid

Compositions and contents of sedimentary pigments were examined using high performance liquid chromatography in order to discuss the spatial distributions of phytoplankton primary production, phytoplankton functional type and the preservation efficiency of phytoplankton pigments and their influencing factors. The results showed that: chloropigments [Chlorins, including chlorophyll-a (Chl-a) and pheopigments (Pheo-a), such as pheophytin-a (PHtin-a), pheophorbide-a (PHide-a), pPheophytin-a (pPHtin-a), sterol chlorin esters (SCEs) and carotenol chlorin esters (CCEs)] were the major type of sedimentary pigments. The nutrients inputs from Changjiang Diluted Water and upwelling in the Zhe-Min coastal mud area were the major cause for the patchy distribution with high sedimentary chloropigment contents. Carotenoid contents showed no trending changes and exhibited high values in the Changjiang Estuary and Zhe-Min Coasts. Based on the relative proportions of each diagnostic carotenoid to the total diagnostic carotenoids in the sediments, the relative contributions of diatoms, dinoflagellates, prymnesiophytes, prasinophytes, cryptophytes and cyanobacterias in the phytoplankton fuctional types were 48.8% +/- 17.4%, 10.7% +/- 11.5%, 8.1% +/- 7.2%, 18.6% +/- 8.2%, 9.4% +/- 6.4% and 4.3% +/- 3.2%, respectively. The preference for external environmental conditions (e.g., nutrient level and water salinity) was the main cause for the decreasing trends of diatoms and dinoflagellates proportions and the increasing trends of prasinophytes, cryptophytes and cyanobacterias seawards. Based on the spatial distribution of Chl-a/Pheo-a ratios, the higher preservation efficiencies of sedimentary pigments in the coastal regions (e.g., outer edge of maximum turbidity zone in the Changjiang Estuary, mouth of the Hangzhou Bay and upwelling region in the Zhe-Min Coast) were mainly due to the higher sedimentation rate and seasonal occurrences of hypoxia in bottom water, and these regions with higher sedimentary pigment preservation efficiencies were probably ideal areas for the marine eco-environmental evolutions. The bad sedimentary environment caused by the water exchange inside and outside of Hangzhou Bay was the dominant reason for the low sedimentary pigment contents and preservation efficiencies in this region.

Topics: Carotenoids; China; Chlorophyll; Chlorophyll A; Cryptophyta; Diatoms; Dinoflagellida; Estuaries; Geologic Sediments; Haptophyta; Phytoplankton; Porphyrins; Water

Zinc complex of pyropheophorbide-b, a derivative of chlorophyll-b, was covalently dimerized through ethylene glycol diester. The synthetic homo-dyad was axially ligated with two methanol molecules from the β-face and both the diastereomerically coordinating methanol species were hydrogen bonded with the keto-carbonyl groups of the neighboring chlorin in a complex. The resulting folded conformer in a solution was confirmed by visible, (1) H NMR and IR spectra. All the synthetic zinc chlorin homo- and hetero-dyads consisting of pyropheophorbides-a, b and/or d took the above methanol-locked and π-π stacked supramolecules in 1% (v/v) methanol and benzene to give redmost (Qy) electronic absorption band(s) at longer wavelengths than those of the corresponding monomeric chlorin composites. The other zinc chlorin and bacteriochlorin homo-dyads completely formed similar folded conformers in the same solution, while zinc inverse chlorin and porphyrin homo-dyads partially took such supramolecules. The J-type aggregation to folded conformers and the redshift values of composite Qy bands were dependent on the electronic and steric factors of porphyrinoid moieties in dyads.

Topics: Chlorophyll; Molecular Conformation; Porphyrins; Zinc

A chlorophyll-a derivative bonded directly with epoxide at the peripheral position of the chlorin π-system was reacted with N-urethane and C-ester protected amino acids bearing an alcoholic or phenolic hydroxy group as well as a carboxy group at the residue to give chlorophyll-amino acid conjugates. The carboxy residues of N,C-protected aspartic and glutamic acids were esterified with the epoxide in high yields. The synthetic conjugates in dichloromethane had absorption bands throughout the visible region including intense red-side Qy and blue-side Soret bands. By their excitation at the visible bands, strong and efficient fluorescence emission was observed up to the near-infrared region. The chromo/fluorophores are promising for preparation of functional peptides and modification of proteins.

Topics: Amino Acids; Catalysis; Chlorophyll; Chlorophyll A; Epoxy Compounds; Esterification; Models, Molecular; Porphyrins; Proteins; Spectrophotometry, Infrared

We investigated the effects of photodynamic therapy (PDT) on anti-tuberculosis (TB) activity by measuring inactivation rates, expressed as D-value, of MDR- and XDR-Mycobacterium tuberculosis (M. tb) clinical strains in vitro. Approximately 10(6) colony forming unit per milliliter (CFU/ml) of the bacilli were irradiated with various doses of laser light after exposure to photosensitizers. Survival of M. tb was measured by enumerating CFU in 7H10 medium to measure D-values. No inactivation of M. tb was observed when exposed to photosensitizers (radachlorin or DH-I-180-3) only or laser light only (P>0.1). Treatment with a combination of photosentizer and laser inactivated M. tb although there was a significant difference between the types of photosensitizers applied (P<0.05). Linear inactivation curves for the clinical M. tb strains were obtained up to laser doses of 30 J/cm(2) but prolonged irradiation did not linearly inactivate M. tb, yielding sigmoid PDT inactivation curves. D-values of M. tb determined from the slope of linear regression lines in PDT were not significantly different and ranged from 10.50 to 12.13 J/cm(2) with 670 nm laser irradiation at 100 mW/cm(2) of the fluency rate, except for a drug-susceptible strain among the clinical strains tested. This suggests that PDT inactivated M. tb clinical strains regardless of drug resistance levels of the bacilli. Intermittent and repeated PDT allowed acceleration of the inactivation of the bacilli as a way to avoid the sigmoid inactivation curves. In conclusion, PDT could be alternative as a new option for treatment for MDR- and XDR-tuberculosis.

Topics: Apoptosis; Chlorophyll; Dose-Response Relationship, Drug; Dose-Response Relationship, Radiation; Drug Resistance, Multiple, Bacterial; Mycobacterium tuberculosis; Photochemotherapy; Photosensitizing Agents; Porphyrins; Treatment Outcome

Using stereoselective aldol-like condensation as a key methodology, a series of chlorophyll a-based long wavelength cationic chlorins were synthesized using methyl pyropheophorbide a (MPPa) and purpurin-18-N-methoxylimide methyl ester as starting materials. Such long wavelength cationic chlorins possess covalently linked cationic moieties (pyridinium or quinolinium) on the peripheral of their tetrapyrrole macrocycles. It was found that all long wavelength cationic chlorins showed their longest absorption maxima in the range of 712-763nm, making them potential photosensitizers in photodynamic therapy. The results of preliminary experiments probing in vitro photodynamic effects showed that the purpurinimide derivatives exhibit relatively high phototoxicity in HeLa cells as compared to MPPa derivatives.

Topics: Cell Survival; Chlorophyll; Chlorophyll A; HeLa Cells; Humans; Neoplasms; Photochemotherapy; Photosensitizing Agents; Porphyrins; Stereoisomerism

Natural-chlorophyll-related porphyrins, including (2H, Zn, Cu)-protoporphyrin IX (Por-1) and Zn-mesoporphyrin IX (Por-2), and chlorins, including chlorin e₆ (Chl-1), chlorin e₄ (Chl-2), and rhodin G₇ (Chl-3), have been used in dye-sensitized solar cells (DSSCs). For porphyrin sensitizers that have vinyl groups at the β-positions, zinc coordinated Por-1 gives the highest solar-energy-to-electricity conversion efficiency (h) of up to 2.9%. Replacing the vinyl groups of ZnPor-1 with ethyl groups increases the open-circuit voltage (V(oc)) from 0.61 V to 0.66 V, but decreases the short-circuit current (J(sc)) from 7.0 mA·cm⁻² to 6.1 mA·cm⁻² and the value of h to 2.8%. Density functional theory (DFT) and time-dependent DFT (TD-DFT) calculations suggest that the higher J(sc) values of Zn-based porphyrin sensitizers result from the favorable electron injection from the LUMO at higher energy levels. In the case of the chlorin sensitizers, the number of carboxyl protons has a large effect on the photovoltaic performance. Chl-2 with two carboxyl protons gives much higher values of J(sc), V(oc), and h than does Chl-1 with three carboxyl protons. Replacing the protons of Chl-1 with sodium ions can substantially improve the photovoltaic performance of Chl-1-based solar cells. Furthermore, the sodium salt of Chl-3 with an aldehyde group at the C7 position shows poorer photovoltaic performance than does the sodium salt of Chl-1 with methyl groups at the C7 position. This is due to the low light-harvesting capability of Chl-3.

Topics: Chlorophyll; Electrochemistry; Photochemical Processes; Porphyrins; Solar Energy

The crystal structure of Photosystem II (PSII) analyzed at a resolution of 1.9 Å revealed deformations of chlorin rings in the chlorophylls for the first time. We investigated the degrees of chlorin ring deformation and factors that contributed to them in the PSII crystal structure, using a normal-coordinate structural decomposition procedure. The out-of-plane distortion of the P(D1) chlorin ring can be described predominantly by a large "doming mode" arising from the axial ligand, D1-His198, as well as the chlorophyll side chains and PSII protein environment. In contrast, the deformation of P(D2) was caused by a "saddling mode" arising from the D2-Trp191 ring and the doming mode arising from D2-His197. Large ruffling modes, which were reported to lower the redox potential in heme proteins, were observed in P(D1) and Chl(D1), but not in P(D2) and Chl(D2). Furthermore, as P(D1) possessed the largest doming mode among the reaction center chlorophylls, the corresponding bacteriochlorophyll P(L) possessed the largest doming mode in bacterial photosynthetic reaction centers. However, the majority of the redox potential shift in the protein environment was determined by the electrostatic environment. The difference in the chlorin ring deformation appears to directly refer to the difference in "the local steric protein environment" rather than the redox potential value in PSII.

Topics: Bacterial Proteins; Chlorophyll; Crystallography, X-Ray; Cyanobacteria; Models, Molecular; Molecular Structure; Oxidation-Reduction; Photosystem II Protein Complex; Porphyrins; Static Electricity

Effects of peripheral substituents in the A- and B-rings of the chlorin macrocycle on demetalation of zinc chlorophyll derivatives were examined. The demetalation rate constants of zinc 8-formyl-chlorin (1) were comparable to those of zinc 3-formyl-chlorin (2). Both 1 and 2 exhibited significantly slower demetalation kinetics than the corresponding zinc chlorin 3, which had no formyl group. Comparison of demetalation kinetics between zinc 3-ethyl-chlorin (3) and zinc 3-vinyl-chlorin (4) indicated that the substitution at the 3-position of the chlorin macrocycle from a vinyl to an ethyl group accelerated removal of the central zinc. Activation energies of demetalation of zinc chlorins 1, 2, and 5 possessing a formyl group in the A- or B-ring of the chlorin macrocycle were larger than those of zinc chlorins 3 and 4 lacking a formyl group. These results indicate that the formyl groups in the A- or B-ring provide tolerance to demetalation of chlorin molecules due to their electron-withdrawing effects.

Topics: Chlorophyll; Kinetics; Organometallic Compounds; Porphyrins; Spectrum Analysis; Temperature; Zinc

A chlorophyll derivative with a central zinc ion, a methoxy functionality at its 3(1)-position, and functionalized with a second-generation dendron (3,4-3,4,5)12G2-CH(2)OH at its 17(2)-position was synthesized starting from natural chlorophyll a (Chl a). This compound exhibits liquid crystalline (LC) behavior and its mesomorphic properties have been characterized by differential scanning calorimetry (DSC), polarisation optical microscopy (POM), powder X-ray diffraction (XRD), and scanning probe microscopy (SPM). A combination of powder XRD, high resolution scanning tunneling microscopy (STM), and atomic force microscopy (AFM) experiments revealed the formation of nano-segregated well-ordered columnar tubular superstructures consisting of about five molecules in the column stratum. These self-assembled columns are further self-organized into a two-dimensional oblique unit cell lattice. Semiconducting behavior of this compound has been studied by pulse-radiolysis time-resolved microwave conductivity (PR-TRMC) method and charge carrier mobility values of ∼10(-2) cm(2) V(-1) s(-1) are observed. Such organized columnar superstructures constructed from semisynthetic zinc chlorins are reminiscent of the tubular organization of the bacteriochlorophyll dyes in the light-harvesting chlorosomal antennae of green sulphur bacteria.

Topics: Bacteriochlorophylls; Chlorophyll; Chlorophyll A; Metalloporphyrins; Microscopy, Polarization; Microscopy, Scanning Tunneling; Molecular Structure; Pigments, Biological; Porphyrins; X-Ray Diffraction; Zinc

Demetalation of three synthetic zinc cyclic tetrapyrroles that possess identical peripheral substituents, zinc methyl bacteriopyropheophorbide a (zinc bacteriochlorin 1), zinc methyl 3-devinyl-3-acetyl-pyropheophorbide a (zinc chlorin 2), and zinc methyl 3-devinyl-3-acetyl-protopyropheophorbide a (zinc porphyrin 3), was kinetically analyzed under acidic conditions to examine the effects of macrocyclic structures on demetalation without peripheral substitution effects. Zinc bacteriochlorin 1 exhibited much slower demetalation kinetics than zinc chlorin 2 and zinc porphyrin 3. These results indicate that the bacteriochlorin skeleton provides significant resistance to the removal of the central metal from the tetrapyrrole ligand. Comparison of demetalation kinetics of 3-acetyl zinc complexes 2 and 3 with that of 3-vinyl zinc complexes under the same reaction condition demonstrated that the relative ratio (5.0 × 10(-2)) of the demetalation rate constant of the 3-acetyl zinc chlorin 2 to that of the corresponding 3-vinyl zinc chlorin 4 resembled the case of the 3-acetyl zinc porphyrin 3 to the 3-vinyl zinc porphyrin 5 (the relative ratio was 6.8 × 10(-2)). These suggest that the electron-withdrawing 3-acetyl group slows down the demetalation from the tetrapyrrole ligands more than the 3-vinyl group and that the 3-acetyl effect is analogous in both chlorin and porphyrin π-systems.

Topics: Chlorophyll; Kinetics; Porphyrins; Tetrapyrroles; Zinc

Metal complexes of methyl 13(1)-(18)O-labelled pyropheophorbide-a1-M-(18)O (M=Zn, Cu and Ni) were prepared by metallation of the (18)O-labelled free base (1-(18)O) and (18)O-labelling of unlabelled nickel complex (1-Ni). The FT-IR spectra of 1-Zn and 1-Zn-(18)O in CH(2)Cl(2) showed that the 13-keto carbonyl stretching vibration mode moved to about a 30-cm(-1) lower wavenumber by (18)O-labelling of the 13(1)-oxo moiety. In 1-Cu-(18)O and 1-Ni-(18)O, the 13-C==(18)O stretching modes were close to the highest-energy wavenumber mode of chlorin skeletal C-C/C-N vibrations at around 1650cm(-1) and they were coupled in CH(2)Cl(2) to give two split IR bands (Fermi resonance). A similar coupling was observed in the resonance Raman scattering of 1-Ni-(18)O in the solid state. The hydrogen-bonded 13-C==(16)O vibration mode of 1-Ni similarly coupled with the skeletal C-C/C-N mode in CCl(4) containing 1% (v/v) 1,1,1,3,3,3-hexafluoro-2-propanol, while such a coupling was not observed in a neat CCl(4) solution of 1-Ni possessing the 13-C==(16)O free from any interaction. The skeletal C-C/C-N band selectively coupled with the 13-C==O, not with the 3-C==O, when the difference in their peak maxima was less than 20 cm(-1).

Topics: Carbon; Chlorophyll; Hydrogen Bonding; Ions; Molecular Structure; Porphyrins; Spectroscopy, Fourier Transform Infrared

The ability to introduce substituents at designated sites about the perimeter of the chlorin or 13(1)-oxophorbine macrocycle is essential for fundamental studies related to chlorophylls. A chlorin is a dihydroporphyrin, whereas a 13(1)-oxophorbine is a chlorin containing an annulated oxopentano ring spanning positions 13 and 15. 13(1)-Oxophorbines bearing auxochromes at the 7-position of the macrocycle are valuable targets given their resemblance to chlorophyll a or b, which contains the 13(1)-oxophorbine skeleton and bears a 7-methyl or 7-formyl group, respectively. A rational route to 7-substituted 13(1)-oxophorbines was developed that relies on a new method for regioselective bromination. Under neutral conditions, a 13-acetyl-10-mesitylchlorin (FbC-M(10)A(13)) undergoes bromination (with 1 molar equiv of NBS in THF) both in ring B (7-position) and at the 15-position (42% versus 28% isolated yield), thereby thwarting installation of the isocyclic ring (ring E, spanning the 13-15 positions). Under acidic conditions (10% TFA in CH(2)Cl(2)), ring B is deactivated, and bromination occurs preferentially at the 15-position (87% yield). The capability for preferential 15-bromination is essential to install the isocyclic ring, after which bromination can be directed to the 7-position of ring B (neutral conditions, 86% yield). The ability to suppress bromination in ring B (under acidic media) has been exploited in syntheses of sparsely substituted analogues of chlorophyll b. The analogues contain a 7-substituent (acetyl, formyl, or TIPS-ethynyl), a 10-mesityl group, and the 18,18-dimethyl group as the only substituents in the 13(1)-oxophorbine skeleton. The three analogues exhibit absorption spectral features that closely resemble those of free base analogues of chlorophyll b. Taken together, the facile access to chlorins and 13(1)-oxophorbines bearing substituents at distinct sites should enable fundamental spectroscopic studies and diverse applications.

Topics: Chlorophyll; Halogenation; Porphyrins; Spectrum Analysis; Stereoisomerism; Substrate Specificity

Amide linked pyro-pheophorbide a dimers, equipped with a suitable length of linkage, assemble in non-polar solvents by internal hydrogen bonding into C2-symmetric stacked structures as evidenced by UV-vis, fluorescence, IR, CD, 1H NMR spectroscopy and DFT molecular modelling studies.

Topics: Amides; Chlorophyll; Circular Dichroism; Dimerization; Fluorescence; Hydrogen Bonding; Magnetic Resonance Spectroscopy; Molecular Structure; Porphyrins; Solvents; X-Ray Diffraction

Demetalation kinetics of natural chlorophyll (Chl) d purified from Acaryochloris marina was first studied and compared with those of Chls a and b. The demetalation rate constant of Chl d, which possessed a formyl group at the 3-position, was five-fold smaller than that of Chl a possessing a vinyl group at the same position in aqueous acetone at the proton concentration of 1.2 x 10(-3) M at 25 degrees C. In contrast, the demetalation rate constant of Chl b possessing a formyl group at the 7-position was 26 times smaller than that of Chl a. The activation energy of demetalation reaction of Chl d was larger than that of Chl a, but smaller than that of Chl b. These indicate that the substitution effect of 3-formyl group on the acidic removal of central magnesium in Chls was smaller than that of 7-formyl group.

Topics: Chlorophyll; Chlorophyll A; Cyanobacteria; Kinetics; Magnesium; Photosynthesis; Porphyrins; Spectrophotometry

Zinc 3(1)-hydroxy-13(1)-oxo-chlorins possessing various perfluorooctyl groups in the 17-propionate were prepared by modifying naturally occurring chlorophyll-a. The synthetic compound having four perfluorooctyl groups readily self-assembled to form large J-aggregates, which were dissolved in fluorous phases with more ordered supramolecular structures than in a hydrocarbon solvent.

Topics: Bacteriochlorophylls; Chemistry, Pharmaceutical; Chlorophyll; Circular Dichroism; Drug Design; Hexanes; Macromolecular Substances; Models, Chemical; Molecular Conformation; Molecular Structure; Porphyrins; Solvents; Zinc; Zinc Compounds

The highly efficient, light-harvesting antennae systems present in chlorosomes of green phototrophic bacteria, such as Chloroflexus aurantiacus, serve as a model for the design of bioinspired nanostructured materials. A semisynthetic zinc chlorin, derived from natural chlorophyll a, is used as a building block that self-assembles into excitonically coupled chromophore stacks. Temperature-dependent UV/Vis and circular dichroism spectroscopic measurements show the reversible formation of soluble chiral aggregates of this new zinc chlorin dye. High-resolution scanning tunneling microscopy and atomic force microscopy studies reveal the formation of two types of well-ordered pi-stacked aggregates on highly ordered pyrolytic graphite.

Topics: Chlorophyll; Circular Dichroism; Graphite; Microscopy, Atomic Force; Microscopy, Scanning Tunneling; Models, Molecular; Porphyrins; Spectrophotometry, Ultraviolet; Spirulina; Temperature

An octaethylporphyrin derivative, 1, possessing an exo-five-membered ring fused at the 13- and 15-positions was oxidized by osmium tetroxide to give two isomeric chlorins, 3 and 5, possessing beta,beta'-dihydroxy groups at the A- and C-rings, respectively. Single dehydration of 2,3-dihydroxychlorin 3 gave a mixture of 2- and 3-(1-hydroxyethyl)porphyrins 7, while that of 12,13-dihydroxychlorin 5 resulted in the sole formation of 131-hydroxyporphyrin 9. The latter was modified smoothly to the phytoporphyrin analogue 2, whose molecular skeleton was similar to that of naturally occurring chlorophylls possessing a 131-oxo group fixed on an exo-five-membered ring.

Topics: Chlorophyll; Porphyrins; Pyrroles

Synthetic chlorins bearing diverse auxochromes at the 3- and 13-positions of the macrocycle are valuable targets given their resemblance to chlorophylls a and b, which bear 3-vinyl and 13-keto groups. A de novo route has been exploited to construct nine zinc chlorins bearing substituents at the 3- and 13-positions and two benchmark zinc chlorins lacking such substituents. The chlorins are sterically uncongested and bear (1) a geminal dimethyl group in the reduced pyrroline ring, (2) a H, an acetyl, a triisopropylsilylethynyl (TIPS-ethynyl), or a vinyl at the 3-position, (3) a H, an acetyl, or TIPS-ethynyl at the 13-position, and (4) a H or a mesityl at the 10-position. The synthesis of the 13-substituted chlorins relied on p-TsOH x H2O-catalyzed condensation of an 8,9-dibromo-1-formyldipyrromethane (eastern half) and 2,3,4,5-tetrahydro-1,3,3-trimethyldipyrrin (western half), followed by metal-mediated oxidative cyclization, affording the 13-bromochlorin. Similar use of a bromo- or TIPS-ethynyl-substituted western half provided access to 3-substituted chlorins. A 3-bromo, 13-bromo, or 3,13-dibromochlorin was further transformed by Pd-coupling to introduce the vinyl group (via tributylvinyltin), TIPS-ethynyl group (via TIPS-acetylene), or acetyl group (via tributyl(1-ethoxyvinyl)tin, followed by acidic hydrolysis). In the 10-mesityl-substituted zinc chlorins, the series of substituents, 3-vinyl, 13-TIPS-ethynyl, 3-TIPS-ethynyl, 13-acetyl, 3,13-bis(TIPS-ethynyl), 3-TIPS-ethynyl-13-acetyl, or 3,13-diacetyl, progressively causes (1) a redshift in the absorption maximum of the B band (405-436 nm) and the Q(y) band (606-662 nm), (2) a relative increase in the intensity of the Q(y) band (I(B)/I(Q) = 4.2-1.5), and (3) an increase in the fluorescence quantum yield phi(f) (0.059-0.29). The zinc chlorins bearing a 3-TIPS-ethynyl-13-acetyl or a 3,13-diacetyl group exhibit a number of spectral properties resembling those of chlorophyll a or its zinc analogue. Taken together, this study provides access to finely tuned chlorins for spectroscopic studies and diverse applications.

Topics: Chlorophyll; Molecular Structure; Porphyrins

The spin-correlated radical pair [P(700)(+)A(1)(-)] gives rise to a characteristic "out-of-phase" electron spin-echo signal. The electron spin-echo envelope modulation (ESEEM) of these signals has been studied in thylakoids prepared from the wild-type strain of Chlamydomonas reinhardtii and in two site-directed mutants, in which the methionine residue which acts as the axial ligand to the chlorin electron acceptor A(0) has been substituted with a histidine either on the PsaA (PsaA-M684H) or the PsaB (PsaB-M664H) reaction center subunits. The analysis of the time domain ESEEM provides information about the spin-spin interaction in the [P(700)(+)A(1)(-)] radical pair, and the values of the dipolar (D) and the exchange (J) interaction can be extracted. From the distance dependence of the dipolar coupling term, the distance between the unpaired electron spin density clouds of the primary donor P(700)(+) and the phyllosemiquinone A(1)(-) can be determined. The [P(700)(+)A(1)(-)] ESEEM spectrum obtained in wild-type thylakoids can be reconstructed using a linear combination of the spectra measured in the PsaA and PsaB A(0) mutants, demonstrating that electron transfer resulting in charge separation is occurring on both the PsaA and PsaB branches. The [P(700)(+)A(1B)(-)] distance in the point dipole approximation in the PsaA-M684H mutant is 24.27 +/- 0.02 A, and the [P(700)(+)A(1A)(-)] distance in the PsaB-M664H mutant is 25.43 +/- 0.01 A. An intermediate value of 25.01 +/- 0.02 A is obtained in the wild-type membranes which exhibit both spin-polarized pairs.

Topics: Animals; Chlamydomonas reinhardtii; Chlorophyll; Dimerization; Electron Spin Resonance Spectroscopy; Electron Transport; Free Radicals; Histidine; Light-Harvesting Protein Complexes; Methionine; Mutagenesis, Site-Directed; Photosystem I Protein Complex; Plant Proteins; Porphyrins; Spin Labels; Thylakoids; Tyrosine

The ethylene ketal of pyropheophorbides, chlorophylls possessing the 13-keto carbonyl group and lacking the 13(2)-methoxycarbonyl group, reacted with H(2)(18)O (ca. 95% 18O atom) by acidic hydrolysis to give efficiently and regioselectively 13(1)-18O-oxo-labelled compounds (ca. 92% 18O). The resulting 18O-labelled chlorin was modified by several chemical reactions to afford some derivatives with little loss of the 18O atom. Following the same procedures, 3(1),13(1)-doubly-18O-labelled pyrochlorophyll derivatives were also prepared. All the synthetic 18O-labelled compounds were identified by FAB-mass and vibrational spectra. Especially, in the vibrational spectroscopic results including IR and resonance Raman spectra, an about 30 cm(-1) wavenumber down-shift of the 3- and/or 13-C[double bond]O stretching vibrational bands was observed by exchanging 3(1)- or 13(1)-oxo-oxygen atom from 16O to 18O.

Topics: Acetals; Chlorophyll; Chlorophyll A; Dioxolanes; Hydrogen-Ion Concentration; Hydrolysis; Isotope Labeling; Magnetic Resonance Spectroscopy; Molecular Structure; Oxidation-Reduction; Oxygen; Oxygen Isotopes; Porphyrins; Spectroscopy, Fourier Transform Infrared

We present a systematic study of the effect of antenna size on energy transfer and trapping in photosystem II. Time-resolved fluorescence experiments have been used to probe a range of particles isolated from both higher plants and the cyanobacterium Synechocystis 6803. The isolated reaction center dynamics are represented by a quasi-phenomenological model that fits the extensive time-resolved data from photosystem II reaction centers and reaction center mutants. This representation of the photosystem II "trapping engine" is found to correctly predict the extent of, and time scale for, charge separation in a range of photosystem II particles of varying antenna size (8-250 chlorins). This work shows that the presence of the shallow trap and slow charge separation kinetics, observed in isolated D1/D2/cyt b559 reaction centers, are indeed retained in larger particles and that these properties are reflected in the trapping dynamics of all larger photosystem II preparations. A shallow equilibrium between the antennae and reaction center in photosystem II will certainly facilitate regulation via nonphotochemical quenching, and one possible interpretation of these findings is therefore that photosystem II is optimized for regulation rather than for efficiency.

Topics: Chlorophyll; Cyanobacteria; Cytochrome b Group; Energy Transfer; Kinetics; Light-Harvesting Protein Complexes; Models, Chemical; Photosynthetic Reaction Center Complex Proteins; Photosystem II Protein Complex; Pisum sativum; Porphyrins

On-line high performance liquid chromatography/atmospheric pressure chemical ionization mass spectrometry (HPLC/APCI-MS) has been applied to the identification of a number of tetrapyrrolic pigments as well as several magnesium-free analogs. Mass spectra, acquired both in positive (+) and negative (-) ionization mode, allow not only the determination of the structural features of the pigments, but also a very easy differentiation of the Mg-chelated pigments from the free bases. In the positive ionization mode, all pigments show mainly a [M + H]+ ion and a [M + H - C20H38]+ fragment ion corresponding to the loss of the phytyl chain via a hydrogen rearrangement. In the negative ionization mode, on the other hand, although all pigments give an abundant molecular anion [M]-*, only the Mg-chelated chlorin spectra show a prominent fragment [M - C20H39]- produced by a formal loss of the phytyl radical.

Topics: Bacteriochlorophylls; Chlorophyll; Chromatography, High Pressure Liquid; Magnesium; Mass Spectrometry; Molecular Structure; Porphyrins

In isolated, chlorosome-free reaction centers from Chlorobium limicola f thiosulphatophilum, a chlorin pigment exhibits a Qy absorption band at 672 nm (Feiler, U., Nitschke, W., & Michel, H. (1992) Biochemistry 31, 2608-2614). To characterize the chemical nature of this chlorin pigment and its interactions within the reaction-center protein, selective enhancement of its Raman scattering was achieved by resonant excitation within its Soret band. This is the first time that structural studies of this pigment were performed on the native reaction-center protein. The obtained resonance Raman spectra were consistent with a single population of a chlorophyll a(-like) pigment, possessing a vinyl group on ring I, but not with bacteriochlorophyll c or bacteriophaeophytin c. The stretching frequencies of the C9-keto carbonyl of this pigment indicates that it is H-bonded to the reaction-center protein. The strength of this H-bond is very close to those of the keto carbonyls of the primary electron acceptors in purple bacterial reaction centers and D1/D2 particles. Since in membranes of Chlorobiaceae a transient bleaching at 670 nm is due to the primary acceptor in the reaction center (Nuijs, A. M., Vasmel, H., Joppe, H. L. P., Duysens, L. N. M., & Amesz, J. (1985a) Biochim. Biophys. Acta 907, 24-34), we thus conclude that the primary acceptor in Chlorobium reaction centers is the characterized chlorophyll a(-like) pigment.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Bacteria; Bacteriochlorophylls; Binding Sites; Chlorophyll; Chlorophyll A; Hydrogen Bonding; Kinetics; Light-Harvesting Protein Complexes; Magnesium; Photosynthetic Reaction Center Complex Proteins; Porphyrins; Spectrum Analysis, Raman

The tetrapyrroles in a highly immature Late Pliocene lacustrine sediment (Willershausen, Germany) show a simple distribution of both chlorin and porphyrin components as the free bases. The major components are C32 desoxophylloerythroaetioporphyrin (DPEP), a C33 bicycloalkano porphyrin, the chlorin analogue of the latter, and desoxophylloerythrin and its chlorin counterpart. The structure of the novel bicycloalkano chlorin was determined using a combination of two-dimensional phase-sensitive COSY NMR and nOe studies. Measurements of delta 13C and other data indicate that DPEP and the bicycloalkano porphyrin were derived from the chlorophyll(s) of photosynthetic organisms utilising a common source of CO2, probably diatoms. The occurrence of DPEP and other minor alkyl porphyrins indicates that the chlorophyll defunctionalisation pathway leading to these components can occur at low temperature and was probably biologically mediated, as was the condensation leading to the fused ring components.

Topics: Aluminum Silicates; Chlorobi; Chlorophyll; Clay; Diatoms; Fossils; Geologic Sediments; Germany; Porphyrins; Pyrroles; Soil Microbiology; Spectrum Analysis; Tetrapyrroles

Chlorophyll (Chl) is often viewed as having preceded bacteriochlorophyll (BChl) as the primary photoreceptor pigment in early photosynthetic systems because synthesis of Chl requires one fewer enzymatic reduction than does synthesis of BChl. We have conducted statistical DNA sequence analyses of the two reductases involved in Chl and BChl synthesis, protochlorophyllide reductase and chlorin reductase. Both are three-subunit enzymes in which each subunit from one reductase shares significant amino acid identity with a subunit of the other, indicating that the two enzymes are derived from a common three-subunit ancestral reductase. The "chlorophyll iron protein" subunits, encoded by the bchL and bchX genes in the purple bacterium Rhodobacter capsulatus, also share amino acid sequence identity with the nitrogenase iron protein, encoded by nifH. When nitrogenase iron proteins are used as outgroups, the chlorophyll iron protein tree is rooted on the chlorine reductase lineage. This rooting suggests that the last common ancestor of all extant photosynthetic eubacteria contained BChl, not Chl, in its reaction center, and implies that Chl-containing reaction centers were a late invention unique to the cyanobacteria/chloroplast lineage.

Topics: Amino Acid Sequence; Bacterial Proteins; Biological Evolution; Chlorophyll; Crystallography; Models, Molecular; Molecular Sequence Data; Multigene Family; Nitrogenase; Oxidoreductases; Oxidoreductases Acting on CH-CH Group Donors; Photosynthesis; Phylogeny; Porphyrins; Protein Structure, Tertiary; Sequence Alignment; Sequence Homology, Amino Acid

P460, an iron-containing chromophore at the active site of Hydroxylamine Oxidoreductase of the ammonia-oxidizing bacterium Nitrosomonas europaea, is a macrocycle of unknown structure with a Soret-like 460-nm absorption band in the ferrous form. The pigment can also be isolated in a peptide, "P460-Fragment". Resonance Raman spectroscopy (lambda ex = 457.9 nm) suggests that P460 is a new type of heme with symmetry properties lower than those of protophorphyrin IX or chlorins and similar to those of chlorophylls and isobacteriochlorins. Some of the resonance Raman vibrations of P460 are shifted in HAO as compared to those of P460-Fragment.

Topics: Binding Sites; Chlorophyll; Heme; Nitrosomonas; Oxidoreductases; Pigments, Biological; Porphyrins; Protein Conformation; Protoporphyrins; Spectrum Analysis, Raman

As models for chlorophyll a (Chl a), methyl ester ClFe(III)pheophorbides (1, pheophorbide a; 2, mesopheophorbide a; and 3, mesopyropheophorbide a) were examined by Fourier transform infrared (FTIR) absorption and resonance Raman (RR) spectroscopy. The infrared (IR) chlorin band above 1600 cm-1, assigned as a Ca-Cm mode (Andersson et al. (1987) J. Am. Chem. Soc. 109, 2908-2916) is shown to be metal-sensitive and responsive to spin state and coordination number for dihydroporphyrins, as well as being diagnostic for the chlorin vs. porphyrin or bacteriochlorin macrocycle. Frequency variations for this metallochlorin IR band thus parallel those of the v10 RR mode of porphyrins in their predictive utility. Qy excitation SERRS spectra of Chl a were compared with Qy excitation RR spectra of 1 and methyl Ni(II)pyropheophorbide a. The data demonstrate that 5-coordinate ClFe(III)pheophorbides are better models for chlorophylls than are ruffled 4-coordinate Ni(II)pheophorbides. Major spectral differences between the three chlorophyll models are associated with the C-9 keto and/or C-10 carbomethoxy vibrational modes. The approx. 1700 cm-1 IR band was formerly assigned solely to v(C = O) of the C-9 keto group. However, this IR feature shifts down to approx. 1685 cm-1 and nearly doubles in intensity when the C-10 carbomethoxy is removed, as for 3. Similar frequency downshifts coupled with intensity increases in the IR are found in the literature on chlorophylls. RR spectra of pheophorbides having the C-10 carbomethoxy group (1 and 2) have bands at both approx. 1700 and approx. 1735 cm-1. However, the C-9 keto v(C = O) mode of pyrophorbins also downshifts to approx. 1685 cm-1, as in the IR spectra. The approx. 1735 cm-1 ester RR mode disappears in the case of pyrophorbins, and is never RR active for nonconjugated esters of porphyrins or chlorins. These data demonstrate an interaction between the C-10 and C-9 carbonyls of phorbins. They also indicate that phorbins tend toward conjugation of the C-10 ester. Biological examples of such conjugation effects have recently been reported, e.g., for the Chl a pi-cation radical (Heald et al. (1988) J. Phys. Chem. 92, 4820-4824). Because the phorbin E ring is the major structural feature distinguishing chlorophylls from non-photosynthetic systems, the participation of the C-10 ester in ring conjugation is suggestive of its biological importance.

Topics: Chlorophyll; Fourier Analysis; Porphyrins; Spectrophotometry; Spectrophotometry, Infrared; Spectrum Analysis; Spectrum Analysis, Raman

A reversed-phase HPLC system with 88% methanol in 1 M ammonium acetate buffer pH 5.35 as the mobile phase on an ODS column was used to analyse pheophorbides a, a', b, b' and pyropheophorbides a and b. Pyropheophorbides a and b were found in samples obtained from silkworm excrement but not from spinach leaves though the preparation methods used were the same. This difference suggests that chlorophylls undergo metabolism in the body of silkworm to give pyrochlorophyll derivatives.

Topics: Animals; Bombyx; Chlorophyll; Chromatography, High Pressure Liquid; Porphyrins; Spectrophotometry, Ultraviolet

Topics: Cells, Cultured; Chlorophyll; Fibroblasts; Humans; Light; Porphyrins; Radiation-Sensitizing Agents

In recent years porphyrins and related materials have been tested as antitumor agents. A technique was devised to obtain dose-response curves for the sensitizer fraction that resists one day of elution by tissue culture medium--the retained fraction. We found a steep "threshold" dose response relationship that helps to explain tumor destruction without damage to normal tissues. The family of dose-response curves produced by a wide range of light exposures suggests that chlorins and porphyrins do not act by identical mechanisms. Moreover, they suggest that chlorins will prove superior in practical use.

Topics: Carcinoma; Cell Survival; Chlorophyll; Chlorophyllides; Colonic Neoplasms; Dose-Response Relationship, Drug; Dose-Response Relationship, Radiation; Hematoporphyrins; Humans; Light; Porphyrins; Tumor Cells, Cultured

The fluorescence decay I(t) and time-resolved spectra I(lambda, t) of some porphyrins and chlorins in ethanol and phosphate-buffered aqueous solution were investigated with a time-correlated single-photon-counting apparatus with a mode-locked Ar+ laser (514.5 nm) as the excitation source. The fluorescence of hematoporphyrin, mesoporphyrin and pheophorbide aa is considerably influenced by the conditions of aggregation (these compounds undergo aggregation in phosphate-buffered solution but not in ethanolic solution). The fluorescence decay of chlorin e6 which remains monomeric in both solvents is single exponential in all cases. The fluorescence spectra of hematoporphyrin, mesoporphyrin and pheophorbide a in phosphate-buffered solution are shifted with respect to the spectra obtained in ethanol; moreover, a new emission band (X band) appears, whose intensity increases on increasing the amount of equilibrium aggregates and shows a fast fluorescence decay. For hematoporphyrin and mesoporphyrin the appearance of the X band emission appears to be correlated with irreversible photoprocesses leading to fluorescent photoproducts. Analysis of the reported fluorescence spectra of cancer cells after incubation with hematoporphyrin derivative suggests that the fluorescent photoproducts might be formed also in vivo.

Topics: Chlorophyll; Ethanol; Hematoporphyrins; Kinetics; Mesoporphyrins; Porphyrins; Solutions; Spectrometry, Fluorescence; Structure-Activity Relationship; Time Factors; Water

Incubation of degreened Chlamydomonas reinhardtii y-1 cells in the dark with m-phenanthroline induced de novo synthesis of a chlorophyllide b-like pigment. The rate of synthesis of this pigment in the dark was greater than that of total chlorophyll in illuminated cells. Most of the newly synthesized pigment was excreted into the culture medium. The product was extracted from the medium as the metal-free pheophorbide, which had a fluorescence excitation maximum at 428 +/- 1 nm and an emission maximum at 657 +/- 1 nm (E428F657) in ethyl acetate (E427F657 in diethyl ether). Three pheophorbide species were extracted from the medium of green cells treated in the dark, a minor component with a spectrum (E410F670) identical to demetallated chlorophyll a, and two major species with spectral values of E428F657 and E433F657. The latter, predominant form had a spectrum identical to demetallated chlorophyll b, which was purified from the algal cells. E428F657 and E433F657 reacted with hydroxylamine and Girard's T-reagent, which caused a shift in the fluorescence emission maximum to 668 nm. Pheophytin b, which contains an aldehyde group, exhibited an identical spectral shift when treated in the same way, but pheophytin a or porphyrin biosynthetic intermediates did not. Proton NMR analysis of the E428F657 chlorin produced by yellow cells treated with m-phenanthroline confirmed the presence of an aldehydic proton. Chelating and nonchelating phenanthroline analogs equally stimulated synthesis of this product.

Topics: Chlamydomonas; Chlorophyll; Chlorophyllides; Chromatography, High Pressure Liquid; Chromatography, Thin Layer; Culture Media; Darkness; Magnetic Resonance Spectroscopy; Phenanthrolines; Porphyrins; Spectrometry, Fluorescence; Time Factors

Topics: Chlorine; Chlorine Compounds; Chlorophyll; Ions; Luminescent Measurements; Porphyrins