PHOTOREACTIONS ASSOCIATED WITH IN VITRO HEMOLYSIS IN ERYTHROPOIETIC PROTOPORPHYRIA*

Abstract— –Photohemolysis of erythrocytes. from patients with erythropoietic protoporphyria results from damage to the cell membrane following photoexcitation of the protoporphyrin molecule by 400 nm radiation. Photochemical reactions were assessed for their roles in initiating cell destruction in an in vitro model in which erythrocytes from patients with erythropoietic protoporphyria were irradiated. Electron transfer properties were demonstrated using 2,3,6-trichloroindophenol as an electron acceptor. Photohemolysis of these abnormal cells is due to cell membrane damage initiating colloid osmotic hemolysis. Additional data are offered demonstrating that this type of photohemolysis is oxygen-dependent, associated with peroxide formation, and preceded by increased osmotic fragility. An hypothesis explaining the interrelationship of the photochemistry of protoporphyrin with the photobiologic observation of hemolysis is offered.     

STUDIES ON THE PROTEIN CONFORMATION OF PHYTOCHROME

Abstract— The extinction coefficients for large rye phytochrome were found to be: Fluorescence and circular dichroism spectra of large- and small-molecular-weight rye phytochrome give no evidence for a protein conformational change on phototransformation of phytochrome. The large molecule has a fluorescence emission peak at 331 nm when excited at 290 nm, and an excitation peak for this emission at 288 nm. The circular dichroism spectra indicate that large rye phytochrome has about 17–20% a-helix content, 30%β-structure and 50% random coil, and that the small rye phytochrome has about 10–13%α-helix content. The ultraviolet difference spectra for large and small rye phytochrome are similar and differ from the difference spectrum of the small oat phytochrome in the relative size of the 296–298 nm peak. The difference spectra may reflect changes in chromophore absorbance and in the environment of amino acid residues near the chromophore, particularly of tyrosine, and perhaps of tryptophan and cysteine.     

SOME PROPERTIES OF PHOTOTRANSFORMATION OF RYE PHYTOCHROME IN VITRO

Abstract— Much of the experimental data in the phytochrome literature has been obtained using a small-molecular-weight protein fragment. Hence, several properties of phototransformation were re-examined using large-molecular-weight rye phytochrome. The kinetics of phototransformation are first-order, both for the conversion of Pr to Pfr and for the reverse reaction. The quantum yield of phototransformation was found to be 0·28 mol Einstein^-1 for the conversion of Pr to Pfr and 0·20 mol Einstein^-1 for the conversion of Pfr to Pr. Intermediates in phototransformation were measured by cycling the pigment with high-intensity mixed red and far–red light. The difference spectrum of these intermediates between 367 and 575 nm was found to be similar to that previously reported for oat and pea phytochrome. Analysis of intermediate decay indicated complex kinetics and not a single first-order species. Transient absorbancy changes in the blue region of the spectrum upon actinic illumination could be attributed to differential rates of initial bleaching of the two forms of the pigment and a consequent alteration in the proportion of the two forms in the mixture until photostationary equilibrium is re-established.     

ON THE POSSIBILITY OF INFRARED LASER-INDUCED CHEMISTRY*

Abstract— –We examine whether chemical reactions can be selectively stimulated by the absorption of infrared photons which individually do not satisfy activation energy requirements. By a modification of available rate theory we demonstrate that a non-thermal enhancement of a chemical reaction is in principle possible, and is primarily dependent upon the extent to which a molecular mode can be coherently driven. An assumed coherent response to laser radiation leading to a minimally detectable rate enhancement places realistic constraints on the allowable magnitudes of vibrational and phase relaxation times for the mode. The inclusion of anharmonic limitations on the amount of energy stored in the mode results in the placement of more stringent though not impossible limitations on the relaxation times. A non-linear reaction rate dependence on light intensity suggests that a Q-spoiled laser be used. Optimum experimental conditions should involve the use of liquids at a temperature where thermal rates are just beneath the level of detection.     

ACTION SPECTRA OF PHYTOCHROME IN VIVO*

Abstract— A method is described to determine spectral properties of phytochrome in vivo. For photochrome in 7-day-old dark-grown Cucurbita pepo L. seedlings the mole fraction of the far-red-absorbing form (P_fr) present at photoequilibrium at 664 nm was found to be 0.76 ± 0.02 in vivo. Based on reflectance measurements, the photon fluence rate just below the surface of the cotyledons was calculated. Local rates of photoconversion for known local fluence rates were measured across cotyledons after non-saturating irradiations with wavelengths between 544 and 781 nm and in situ molar photoconversion coefficients were obtained. In contrast to purified oat phytochrome, the in situ molar photoconversion coefficients for P_fr show a strong shoulder between 660 and 700 nm. The maximum of P_fr absorption is at 726 nm. An isosbestic point of phytochrome is found at 686 nm. The mole fraction of P_fr present at photoequilibrium with 686 nm light is 0.58. The ratio of photoconversion quantum yields (that for P_r→ P_fr divided by that for P_fr→ P_r) is 1.38 ± 0.06.     

聚二氧杂环己酮合成可吸收缝合线体外降解研究

通过检测缝线的力学性能、分子量、失重、大分子取向和结晶度变化研究了聚二氧杂环己酮可吸收缝合线（ＰＤＳ）在体外的初期（４星期）降解过程。研究结果表明，本文工艺条件下研制的ＰＤＳ缝线在体外降解过程中随着分子量和大分子取向程度下降，力学性能明显下降，４星期时强度基本消失。分子量明显下降而失重变化很小的结果，证明ＰＤＳ缝线的酯水解在链中和链端等活性，水解的几率只与水分子的可及程度相关。ＷＡＸＤ和ＤＳＣ测试结果表明，随着体外降解进行，缝线原晶区基本保持不变，无定形区产生新的有序区，并且含量不断增加和完善。采用结晶高聚物微原纤结构模型解释ＰＤＳ缝线无定形区在水中的降解机理与研究结果相吻合。ＰＤＳ缝线的体外降解机理与聚羟基乙酸（ＰＧＡ）缝线基本相同     

IN VITRO SYNTHESIS OF PHYTOCHROME APOPROTEIN DIRECTED BY mRNA FROM LIGHT AND DARK GROWN AVENA SEEDLINGS*

Abstract— The aim of this work was to compare the translatability of poly-A-RNA from light- and dark-grown Arena seedlings, the product of which should be phytochrome. Polysomal poly-A-RNA was isolated from 4-day-old dark-grown Avena seedlings (Avena sativa L. cv. Garry) and translated in a rabbit reticulocyte lysate system. Immunoprecipitation by anti-phytochrome serum was used for measurements of specific phytochrome translation. Characterization of the translated and immunoprectpitated protein was performed by comparison with [₃₅S]-methionine in vivo-labelled phytochrome. The specificity of the precipitation was shown by parallel use of non-specific serum and by competitive inhibition of precipitation by exogenous unlabelled phytochrome isolated by affinity chromatography.     

红外光谱法观察二茂镧系氯化物与氧及水蒸气的反应

用实时红外光谱法在特制的收池中分别观察了一些二茂镧系氯化物Ｃｐ２ＬｎＣｌ（Ｃｐ＝Ｃ５Ｈ５，Ｌｎ＝Ｈｏ，Ｇｄ，Ｌｕ）和Ｃｐ＇Ｌｕ（Ｃｐ＝Ｃ５Ｈ４ＣＨ２ＯＣＨ３）与干燥氧气及水蒸气的反应，结果证实这些二茂镧系配合物在室温干燥氧气氛中很稳定，但在水蒸气中很快水蒸气的反应，结果证实这些二茂镧系配合物在室温干燥氧气氛中很稳定，但在水蒸气中很快水解。从水解反应的最初始阶段和较后期采得的红外光谱提示可能存在...     

PHYTOCHROME IN ETIOPLASTS IN RELATION TO CHLOROPHYLL ACCUMULATION*

Abstract— De-etiolation of maize seedlings reduces their sensitivity for red light potentiation of rapid chlorophyll accumulation in white light. An earlier proposal (Raven and Spruit, 1973) attributes this to migration of the far-red absorbing form of phytochrome (P_fr) to receptors essential for chlorophyll synthesis, thereby increasing the local P_fr/total phytochrome (P_tot)ratio. We have studied etioplasts as possible loci for such P_(r receptors. The level of spectrophotometric phytochrome in purified etioplasts isolated from red preirradiated maize seedlings was higher than that of dark grown plants. The difference was marginally significant, however. We argue that migration of a fraction of cytoplasmic P_fr to the etioplasts, too small to be spectrophotometically demonstrable, could still meet the requirements of the model. Dark destruction of bulk spectrophotometric P_fr following saturating red irradiation of seedlings is not paralleled by a decrease of etioplast phytochrome. the latter remaining essentially constant over long periods. On the other hand, the potentiating effect of red light in intact seedlings is still partially reversible by far red light even after 24 h of darkness when destruction of bulk P_fr is complete. Since this demonstrates persistent presence of P_fr active in potentiation, we propose that at least part of this P_fr is associated with the etioplasts.     

低聚壳聚糖与角质层蛋白相互作用的体外研究

以脱脂鼠角质层和提取角蛋白为模型,利用电化学交流阻抗谱、SEM、DSC、FTIR等手段,研究了低聚壳聚糖及氨基葡萄糖与角质层蛋白之间的相互作用.结果发现,氨基葡萄糖溶液处理不影响脱脂角质层膜的阻抗,而低聚壳聚糖溶液处理却可明显降低脱脂角质层膜的阻抗,阻抗值由3.79×106Ω·cm2降至8.379×105Ω·cm2.此外,壳聚糖溶液和氨基葡萄糖溶液都可明显破坏角蛋白表面致密均匀的结构,并使得角蛋白二级结构中α-螺旋结构含量减少,同时推动角蛋白α-螺旋结构向β-折叠结构和无规则卷曲结构转变.这些结果表明低聚壳聚糖与角质层蛋白相互作用,一方面可有效降低鼠角质层结构的紧密程度,另一方面可影响角蛋白的微结构,使其结构松散并出现微细孔道,从而为药物的经皮吸收与传送疏通了通道.     

IMMUNOBLOT ANALYSIS OF CROSS-REACTIVITY OF MONOCLONAL ANTIBODIES AGAINST OAT PHYTOCHROME WITH PHYTOCHROME FROM SEVERAL PLANT SPECIES*

Abstract— Eighteen monoclonal antibodies (mAbs) have been produced against partially degraded phytochrome from A vena sativa cv. Trafalgar. Several of the mAbs are capable of recognising the antigen on immunoblots following sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS-PAGE) and protein electroblotting. Six of these mAbs were screened for the ability to recognise electroblotted phytochrome from eight plant species. Particularly amongst the monocot species tested, but also amongst the dicots, the mAbs showed extensive cross-reactivity. The results suggest that there are perhaps several conserved antigenic sites on the phytochrome molecule.     

OPTICAL PROPERTIES OF PHYTOCHROME IN THE BLUE- SPECTRAL REGION AS MEASURED IN VIVO AND IN VITRO

In the blue spectral region, the phototransformation difference spectrum of oat phytochrome extracted as P_fr differs from that of phytochrome extracted as P_r. The difference absorbance maximum for phytochrome extracted as P_fr is at 420 nm, while that extracted as P_r is at 412 nm. The phototransformation difference spectrum measured in the blue in oat coleoptile tips without inner leaves, corresponds very well with that of phytochrome as extracted in its P_fr form. There is, however, a slight apparent attenuation of the blue difference band relative to those in the red-far-red. In coleoptile tissue containing inner leaves, the blue difference band is relatively even more highly attenuated. A similar attenuation is observed in the blue, in the protochlorophyllide to chlorophyllide phototransformation difference spectrum. In the spectrum measured with excised coleoptile without inner leaves, there is a small attenuation, while in coleptile tissue with inner leaves the attentuation is nearly 9-fold. These data suggest that the observed attenuation is probably artifactual. Neither instrumental non-linearity nor fluorescence induced by the measuring beam could explain the observed attenuation. It is suggested that the observed attenuation is probably mainly the result of wavelength dependent scatter amplification, the amplification in the blue being attenuated by the high background absorption of other pigments in this region.     

ON THE POLARITY OF PHYTOCHROME CONTROLLED LEAF UNFOLDING IN DARK-GROWN SEEDLINGS OF TRITICUM AESTIVUM*

Abstract— The apical ends of sections cut from etiolated primary wheat leaves show a higher degree of phytochrome-mediated leaf unfolding than the basal ends, regardless of the position of the segment in the leaf. It could be clarified that this polarity is not the result of a stable gradient within the intact dark grown leaf. Partial irradiation experiments have shown that leaf unfolding can be induced much more effectively by an irradiation of the apical half of a section than of the basal half. Though only the basal end of the sections was incubated with gibberellic acid (GA₃) in darkness, the degree of the GA₃-induced leaf unfolding was highest at the apical end of these non-irradiated segments. The results suggest that in each separate leaf section a new gradient of a still unknown “unfolding-factor” is established.     

IN VITRO STUDIES ON THE PHOTOSENSITIZED OXIDATION OF LENS PROTEINS BY PORPHYRINS

Abstract Porphyrins, which may be introduced into the eye as a result of abnormal porphyrin metabolism (uroporphyrin–Uro) or when used in the diagnosis or photodynamic therapy of certain tumors, including intraocular tumors (hematoporphyrin–Hp and'hematoporphyrin derivative'–Hpd and mesotetra( P -sulfonatophenyl)porphyrin–TPPS) are efficient photosensitizers in biological systems. We have been studying the potential phototoxic side effects of these drugs in the lens of the eye. Encapsulated in the human lens is a mixture of soluble protein crystallins. With little turnover of protein in the lens, any photosensitized modifications will accumulate and may result in an opacification of the lens. To evaluate the potential of different porphyrins to induce such damage, a series of porphyrins were photolyzed (transmission above 295 nm) in the presence of calf lens protein (2 mg m⁻¹). Marked photopolymerization and histidine destruction were observed for the lens protein photolyzed in the presence of all of the drugs. We have found that the relative effectiveness of the following porphyrins to induce that damage is: Uro = TPPS Hpd = Hp. Both the singlet oxygen quencher, azide, and the free radical scavenger, penicillamine, decrease this photosensitized oxidative damage to lens protein. TPPS binds significantly to lens protein and this binding leads to conformational changes in that protein.     

THE EFFECT OF PROLONGED LIGHT EXPOSURE ON THE EFFECTIVENESS OF PHYTOCHROME IN ANTHOCYANIN SYNTHESIS IN TOMATO SEEDLINGS*

Abstract— The hypocotyl of the tomato ( Lycopersicon esculentum ) seedling synthesizes large amounts of anthocyanin if exposed to prolonged light. Single light pulses are totally ineffective. The involvement of phytochrome can be shown by light pulse treatments following a prolonged light exposure. It is predominantly the action of blue/UV light which leads to a high responsiveness of anthocyanin synthesis towards phytochrome. Moreover, the data suggest a phytochrome-independent action of blue/UV light, in particular of UV-B, on anthocyanin synthesis.     

APPARENT INCREASE IN PHOTODETECTABLE PHYTOCHROME IN THE PRESENCE OF CALCIUM CARBONATE*

Abstract— –Using squash seedling extracts ( Cucurbita pepo L.), we describe an apparent increase in photodetectable phytochrome content that is depended upon the use of either CaCO₃ or starch as a scattering agent and is also a function of preirradiation with red light of either intact tissue or crude extracts prior to sample preparation for spectrophotometric assay. This apparent increase in photodetectable phytochrome content requires about 1 h for full expression after addition of the scattering agent. If irradiations are given in vitro , the increase is partially reversed by far red light when using squash extracts, and is fully reversed with Aoena and Zea extracts. For squash extracts the magnitude of this increase. which is typically between 20 and 40%. is quantitatively correlated with both the amount of Pfr produced by a brief red irradiation and with the proportion of the total phytochrome pool that is pelletable upon centrifugation at 20,000 g . The correlation with pelletable phytochrome does not hold for Auena and Zea , at least when irradiations are gwen in uitro . The increase in photodetectable phytochrome may result from changed phytochrome extinction, changed phototransformation quantum yields, and/or specific trapping of phytochrome by the scattering agent as it settles. An important consequence of these data is that they indicate a need for caution when using a scattering agent during spectrophotometric assay of phytochrome.     

THE EFFECT OF APPLIED THICKNESS ON SUNSCREEN PROTECTION: IN VIVO AND IN VITRO STUDIES

The effect of applied thickness of sunscreen on the degree of protection against ultraviolet exposure has been investigated using an in vitro model (excised human epidermis) and phototesting in human subjects. The agreement between the protection factors obtained by each technique was excellent. It is shown that the influence of applied thickness on sunscreen protection factors is much less than would be predicted by Beer's law.     

红外发射光谱法用于煤化学研究

提出了一套可用于煤化学研究的红外发射光谱测试系统 ,设计了一个结构简单 ,使用方便的样品槽 ,讨论了操作中影响光谱质量的有关因素 ,并以观察煤在低温氧化过程中结构变化为例 ,展示了这一方法在煤化学领域中的应用前景。     

聚对二氧环己酮-乙交酯缝合线体外降解研究

用DSC和X ray衍射法对聚对二氧环己酮 乙交酯 (PDG)缝合线的初期体外降解过程进行了研究 ,并使用纤维摄影仪对纤维在降解过程中的表面形态变化进行了观察 .研究结果表明 ,在本工艺条件下得到的PDG缝合线在体外降解过程中分为三个阶段降解 ,即无定形区降解、原纤间非晶区降解和晶区水解 .在体外降解过程中 ,PDS缝合线以横向断裂为主 ,而PDG则通过表面积的大块脱落来完成降解过程