ABSORPTION SPECTRAL SHIFTS OF CAROTENOIDS RELATED TO MEDIUM POLARIZABILITY

Abstract–Solvent induced absorption spectral shifts of the electronic transition from ground 1 A_g state to the excited 1B_u state in carotenoids have been studied. It is shown that the shift depends only on dispersion interactions in non-polar solvents. In polar media there is just a small extra contribution to the red-shift, due to other forms of interactions. The spectral shifts are well described by the theory, which expresses the shift relative to the gas phase value, as a function of solvent polarizability. The main conclusion is that the dominating mechanism behind the large red-shifted absorbance of carotenoids in the proteinacous environment, in vivo, is the mutual polarizability interactions between the carotenoids and the surrounding medium. The solution-phase values of the dipole moments of the lA_g to 1B_u transitions and the differences of isotropic polarizability between 1B_u and lA_g states of carotenoids in non-polar solvents are calculated and found to be around 13 D and 360 ų respectively. From the great overlap of absorption spectra between carotenoids in quinoline and carotenoids in vivo in purple bacterial antenna complexes, it can be expected that the carotenoids are surrounded by several aromatic amino acids in vivo. Comparisons have been done between the exicted states in carotenoids and in linear conjugated polyenes.     

LOW TEMPERATURE ABSORPTION SPECTRUM STUDIES: LIGHT-INDUCED PIGMENT CHANGES IN PORPHYRIDIUM CULTURES

Abstract— A single-beam spectrophotometer with a low-temperature sample chamber has been built. An automatic mechanical device gives a suitable baseline and allows very reproducible absorption spectra to be obtained. The red alga Porphyridium grown, during a few days, under high-intensity monochromatic light, showed a decrease in the amount of chlorophyll a forms directly or indirectly excited. Furthermore, a 585 nm absorption band is observed in red algae such as Porphyridium, Rhodymenia and Calliblepharis. This 585 nm band is labile to many damaging agents.     

THE FUNCTION OF P700 AND CYTOCHROME f IN THE PHOTOSYNTHETIC REACTION CENTER OF SYSTEM 1 IN RED ALGAE

Abstract— Kinetics and quantum yields of light-induced oxidation of P700 and the f -type cytochrome were measured in marine red algae from absorbance changes in the region 420, 435 and 705 nm. The quantum yield for cytochrome oxidation in Iridaea splendens and Schizymenia pacifica was 0.5-0.65 in far-red light, at 20 and at 0C.
Oxidation rates of P700 measured when varying amounts of cytochrome were in the oxidized state indicated that a reaction center of system 1 in Iridaea contains 1 P700 and 4 cytochrome molecules. Oxidized P700 only accumulates when all 4 cytochromes are oxidized. The rate of photochemistry of system 1, measured as the sum of the rates of P700 and cytochrome oxidation, was independent of the oxidation level of cytochrome, but decreased with accumulation of oxidized P700. This decrease was less than proportional to the fraction of P700 that was in the oxidized state, which suggested transfer of excitation energy between reaction centers.
The quantum yield for cytochrome oxidation after dark periods of 1 min or more was only about 0.2. This effect was tentatively ascribed to a dark reduction of the cytochrome coupled to phosphorylation.     

LIGHT-INDUCED SYNTHESIS OF ANTHOCYANIN IN CARROT CELLS IN SUSPENSION—IV. THE ACTION SPECTRUM

Abstract— Using carrot cell suspension in 2,4-dichlorophenoxyacetic acid (2,4-D)-depleted culture medium, fluence-response curves for the formation of anthocyanin were determined at various wavelengths from 250 to 800 nm. In the fluence-response curves at wavelengths between 260 and 330 nm, the response showed a sharp fluence-dependent increase after the fluence exceeded threshold level at the respective wavelength. Such a sharp increase in response was not observed by light at 450 nm or longer wavelengths, although the response obtained by higher fluence of such light was always higher than that in the dark control. Action spectra determined at the sharp increasing phase of the response showed the single peak at 280 nm which equals the absorption maximum of UV-B photoreceptor.
Although red (R)-light alone had a minor effect on anthocyanin accumulation, it modulated the action of UV-B light. That is, when carrot cells were irradiated with R-light either before or after UV-B irradiation, anthocyanin formation was greatly enhanced above the level enhanced by UV-B light alone. The most effective wavelength for this enhancement was 660 nm. The effect of R-light on the anthocyanin formation of the UV-B irradiated cells was reversed by immediately following it with far-red light, suggesting the involvement of phytochrome in the R-effect.     

SALT AND pH-DEPENDENT CHANGES OF THE PURPLE MEMBRANE ABSORPTION SPECTRUM

Abstract —Purple membrane suspensions change their color to blue and the absorption maximum shifts to 608 nm when the membrane is deionized on a cation exchange column or when it is washed first with < 2N NaCl followed by deionized water. The deionized chromophore is essentially identical with the chromophore produced by lowering the pH of the native membrane to < 4.0 (p K < 3.0). However, the deionized membrane does not aggregate and can be obtained in the pure state. The original purple color of the membrane is restored by addition of around 1 m M Na⁺, K⁺ or 10 μ M Mg²⁺, Ca²⁺, Sr²⁺, Mn²⁺, Pb²⁺ or La²⁺ when the protein concentration is 5μ M . The required salt concentrations decrease with decreasing pH. Direct measurement of bound Ca²⁺ by atomic absorption spectroscopy yields a ratio of Ca²⁺ to protein of <2 and a binding constant of 1.4 × 10⁶. Titration of the spectral change with salts at different pH values shows a linear relation between the pH and the logarithm of the salt concentration, with a 1:1 ratio for Na⁺ and 1:2 ratio for Ca²⁺. These relations are well predicted by Gouy-Chapman theory; however, the accompanying release of protons, changes of the CD spectrum, the complex kinetics of the spectral change during reconstitution with salt and preliminary X-ray diffraction results all suggest that conformational changes may be occurring in the protein.     

RESOLUTION OF OVERLAPPING BANDS IN THE NEAR-UV ABSORPTION SPECTRUM OF INDOLE DERIVATIVES

Abstract— The second derivative spectra of tryptophan in water and in ethylene glycol at 22°C have been integrated in order to obtain the corresponding primitive functions. The integration was carried out by making use of Tchebychev polynomials. The results show that the integrated primitive functions do not correspond to the original absorption spectra of tryptophan in various solvents, but they reflect only the contributions of the ¹L_b bands of the indolic chromophore. The identification of the electronic component, which generates the second derivative spectrum, was based on the solvent insensitivity of the derivative peaks. The comparison between the absorption spectra reported in this paper and those calculated for the ¹L_b←¹A electronic transition of indole confirmed the assumption that the derivation process eliminates the broad, although more intense, contributions coming from the ¹L_a←¹A electronic transition.     

INFLUENCE OF CHANGES IN THE PHOTON PROTECTIVE ENERGY DISSIPATION ON RED LIGHT-INDUCED DETRAPPING OF THE THERMOLUMINESCENCE Z-BAND

-Thermoluminescence emission at 110 K (Z-band) was markedly diminished when thylakoid membranes were exposed to red light during or after Z-band charging with blue light. Analysis of this phenomenon showed that deactivation of Z-band-emitting chlorophyll species occurred preferentially on the low temperature side of the glow curve, and red light of670–680 nm was most efficient in the deactivation. In order to test our hypothesis that this detrapping is related to local heating effects caused by dissipation of absorbed energy, we measured thermoluminescence glow curves and Z-band emission spectra from spinach leaf discs and thylakoid membranes during induction of nonphotochemical chlorophyll fluorescence quenching. Pretreatment of the plant material was designed to achieve different levels of (1) de-epoxidized xanthophylls in the photosynthetic apparatus and (2) the proton concentration in the thylakoid lumen. In comparison, measurements were performed in aggregated and trimeric light-harvesting pigment-protein complexes of photosystem II. We observed on all three levels of organization that a higher capacity of excitation energy dissipation was accompanied by a stronger red light-induced detrapping of Z-band thermoluminescence.     

RED LIGHT-INDUCED ACCUMULATION OF EUGLENA GRACILIS

Abstract— It has been shown that green cells of Euglena gracilis accumulate under red light. The action spectrum of this response has been determined. Its shape and the results of regreening experiments clearly indicate a role of chlorophyll. The quenching effect of chlorophenyl–dimethylurea and NH₂OH demonstrate that this cell accumulation is not directly light-induced, as in the case of phototaxis, but is due to an effect of oxygen, the evolution of which is stimulated by light.     

ROLE OF HYDROXYL-BEARING AMINO ACIDS IN DIFFERENTIALLY TUNING THE ABSORPTION SPECTRA OF THE HUMAN RED AND GREEN CONE PIGMENTS

The human red and green cone pigments differ at either 15 or 16 amino acids, depending upon which polymorphic variants are compared. Seven of these amino acid differences involve the introduction or removal of a hydroxyl group. One of these differences, a substitution of alanine for senne at position 80, was found previously to produce a 5 nm blue shift. To determine the role of the remaining six hydroxyl group differences in tuning the absorption spectra of the human red and green pigments, we have studied six site-directed mutants in which single amino acids from the green pigment have been substituted for the corresponding residues in the red pigment. Blue shifts of 7 and 14 nm were observed upon substitution of phenylalanine for tyrosine at position 277 and alanine for threonine at position 285, respectively. Single substitutions at positions 65, 230, 233, and 309 produced spectral shifts of 1 nm or less. These data are in good agreement with a model based upon sequence comparisons among primate pigments and with the properties of site-directed mutants of bovine rhodopsin. Nonadditive effects observed in comparing the absorption spectra of red-green hybrid pigments remain to be explained.     

蛋白质-铍试剂体系的吸收光谱研究及分析应用

试验发现铍试剂与无色的蛋白质能发生染色反应,用吸光光度法研究了铍试剂与牛血清白蛋白反应前后的吸收光谱及其反应机理,蛋白质结合铍试剂的结合数为１７,制作了重要蛋白质的工作曲线,线性范围约为２０～１８０μｇ．ｍｌ＾－１,用于人血清样品测定,结果满意。     

ORIGIN OF THE RED SHIFTED ABSORPTION IN PHYCOCYANIN 632 FROM Mastigocladus laminosus

A phycocyanin (PC) with γ_max= 632 nm (PC632) was isolated from extracts of the cyanobacterium Mastigocladus laminosus. The complex contained three polypeptides migrating in SDS-PAGE around 22 kDa. The N-termini of the three polypeptides are identical. They are homologous to rod-core linker polypeptides from this and another cyanobacterium and are identical to one of them. Reconstitution of PC (γ_max=614–620 nm) with the 22 kDa polypeptide produced again the red-shifted PC632. Phycobilisomes isolated at pH 6 or 7 contain, if any, only traces of 22 kDa polypeptides. Origin and functions of the polypeptides are discussed, they are most likely proteolytic fragments of allophycocyanin-PC rod-core linkers that contain the PC-binding domain.     

RESOLUTION ENHANCEMENT AND DECONVOLUTION INTO VIBRONIC BANDS OF THE FLUORESCENCE AND ABSORPTION SPECTRA IN THE RED REGION OF DILUTE SOLUTIONS OF CHLOROPHYLL a IN VARIOUS SOLVENTS

Abstract— The fluorescence spectra of chlorophyll a in less than 10–⁶ mol dm^-3 solutions of benzene, toluene, tetrahydrofurane, EPA and ethanol were determined at both room temperature and at 77 K. Resolution enhancement using Fourier transform methods revealed the presence of vibronic bands with essentially solvent-invariant separation between their origins; this was confirmed by the quantitative deconvolution of the spectra into Gaussian bands. It was concluded that the fluorescence is due to a single spectroscopically distinct species. The absorption spectra in the above solvents include a band absent from the fluorescence spectra which, from its position and intensity, may be identified as the longest wavelength X-polarized electronic transition of chlorophyll a. Differences observed between the shapes of the low-and high-temperature absorption spectra may be attributed primarily to the narrowing of bandwidths with decreasing temperature and to intensity redistribution within the bands; the apparent increase in the integrated intensity on cooling the solutions appears to be due largely to the increase in the volume concentration of the solute as a result of solvent contraction.     

RED LIGHT-INDUCED REDUCTION OF A PARTICLE-ASSOCIATED b-TYPE CYTOCHROME FROM CORN IN THE PRESENCE OF METHYLENE BLUE*

Abstract— In the presence of methylene blue, red light causes the reduction of a h-type cytochrome in particulate fractions from corn coleoptiles. Two types of difference spectra for the cytochromes in these fractions are presented: (a) red light-minus-dark in the presence of methylene blue, and (b) dithionite-reduced-minus-oxidized. Comparison of these spectra shows that photoexcited methylene blue selectively reduces a b-type cytochrome which constitutes at most only 30% of the total dithionite-reducible cytochrome present in the most active fractions. The photoreducible cytochrome has an alpha band at room temperature near 557 nm. Bleaching of methylene blue precedes cytochrome reduction under appropriate conditions, suggesting that the photoreduced dye is donating an electron to the cytochrome. This electron transfer does not involve a flavin, at least as judged by the absence of light-induced spectral changes attributable to flavins. Preliminary kinetic studies suggest that EDTA provides the pool of electrons for the reaction. The cytochrome cannot be assigned exclusively either to mitochondria or to endoplasmic reticulum, as judged by its sedimentation properties. These results and the current literature are discussed in the context of the hypothesis that this b-type cytochrome may be involved in the photoreception mechanism for blue and uv light in vivo.     

CAROTENOID PIGMENTS AND THE IN VIVO SPECTRUM OF BACTERIOCHLOROPHYLL

Abstract— The isolation of a mutant, strain PM-9, of Rhodopseudonionus spheroides with an abnormal complement of carotenoid pigments is described.
PM-9 accumulates phytoene, phytofluene, ζ-carotene and neurosporene. Semi-aerobic cultures form more ζ-carotene and neurosporene relative to total carotenoids than do photo-synthetic cultures.
PM-9 is killed on exposure to light and oxygen.
By making use of the effect of oxygen on the nature of the carotenoids in PM-9, we have shown that these pigments do not directly influence the in vivo spectrum of bacteriochlorophyll.
Diphenylamine inhibits the synthesis of coloured carotenoids in Rhodopseudonionos gelatinosa but does not change the bacteriochlorophyll spectrum.     

单波长紫外可见分光光度计测定药物的导数吸收光谱

建立单波长紫外可见分光光度计测定药物导数吸收光谱的方法。以较小的波长差连续测定药物在不同波长λ处的吸光度A,得到一系列A-λ数据,对该系列数据进行数学处理,分别计算Δλ、ΔA、ΔA/Δλ、Δ2A/Δλ2、Δ3A/Δλ3,分别以ΔA/Δλ、Δ2A/Δλ2和Δ3A/Δλ3等各阶导数为纵坐标,以λ为横坐标,绘制药物的一阶、二阶或三阶导数吸收光谱。采用本方法测定并绘制了维生素A、布洛芬两种药物的一阶、二阶及三阶导数吸收光谱,并与其常规吸收光谱进行对比,表征了药物导数吸收光谱的特性。本方法测定药物的导数吸收光谱,扩大了单波长紫外可见分光光度计的应用范围和分析功能,实现了在单波长紫外可见分光光度计上测定药物导数吸收光谱。     

THE CALCULATION OF CHEMICAL SHIFTS IN DISUBSTITUTED NAPHTHALENES

<正> The principle and method for calculating the chemical shifts of substituted benzenes have been extended to the calculation of chemical shifts in disubstituted naphthalenes. We have set up a series of empirical parameters for the calculation of chemical shifts. The calculated results of 439 8 values from 78 compounds show that the standard deviation between the calculated and the experimental values is 0.08 ppm. The combination of this calculation with that of the coupling constants can be used to provide a criterion .for the determination of molecular structure in disubstituted naphthalenes as well as to assign NMR parameters for the experiment of proton simulated spectra of disubstituted naphthalenes.     

EFFECT OF PHYTOCHROME ON THE BIOSYNTHESIS OF ACYCLIC AND CYCLIC CAROTENOIDS IN HIGHER PLANTS

Radish plants were grown in the presence of three different herbicides that interfere with the formation of the normal range of cyclic carotenoids, leading to an accumulation of acyclic biosynthetic intermediates, mainly phytoene (SAN 6706 and amitrole) and zeta-carotene (3852). Plants were then irradiated by four different light programs in order to gain more insight into the first steps of carotenoid biosynthesis and their control by light and phytochrome. In all cases, herbicide-treated and control, carotenoid biosynthesis was greatly enhanced by red light consistent with an effect of phytochrome on the early steps of the pathway. However, similar enhancement was also obtained after treatment with far-red light. Indeed with SAN 6706-treated plants synthesis of phytoene was stimulated to a much greater extent by far-red light given alone, than by red light. The involvement of phytochrome in the regulation of carotenoid biosynthesis appears not to be as simple as previously supposed.     

THE CALCULATIOM OF CHEMXCAL SHIFTS IN SUBSTITOTED PYRIDINES

<正> The principle and method for the calculation of chemical shifts of substituted benzenes have been extended to calculation of the cheiiical shifts in substituted pyridines. We have set up a series of empirical parameters for calculation of the chemical shifts. The calculated results of 154 δ values frou 54 compounds show that the standard deviation between the calculated and the experimental values is 0 . 09 ppm. The combination of the coupling constants can be used to provide a criterion for the determination of molecular structure in substituted pyridines and to assign NMR parameters for the experiment of proton simulated spectra of substituted pyridines.     

THE FLUORESCENCE OF CAROTENOIDS

Abstract The carotenoids β-carotene, lutein, lutein epoxide and violaxanthin are fluorescent compounds. Upon excitation around 280 mμ, at wavelengths close to those of their ultraviolet absorption maxima, these carotenoids fluoresce within the range 300–400 mμ, with maximal fluorescence at approximately 320–340 mμ.     

THE EFFECT OF RED AND FAR RED LIGHT ON CAROTENOID AND CHLOROPHYLL FORMATION IN PEA SEEDLINGS

Abstract— Treatment of etiolated pea seedlings with a short exposure to red light caused a stimulation of growth (size and dry wt production) and carotenoid synthesis during the following 48 hr compared with seedlings kept entirely in darkness.The effect is nullified by a following dose of far red light and thus the phenomenon is probably phytochrome-controlled.
Similar treatment with red light one hour before continuous illumination with white light tended to reduce the lag period for chlorophyll synthesis.Again a following dose of far red light reversed this response.