PHOTOIONIZATION OF MELANIN PRECURSORS: AN ELECTRON SPIN RESONANCE INVESTIGATION USING THE SPIN TRAP 5,5-DIMETHYL-1-PYRROLINE-1-OXIDE (DMPO)

Abstract The photoionization of 3,4-dihydroxyphenylalanine (dopa) and catechol has been studied by electron spin resonance spectroscopy using the free radical scavenger 5,5-dimethyI-1-pyrroline-1 -oxide as a spin trap for hydrated electrons and hydrogen atoms. The photochemistry of these materials is shown to resemble tyrosine in that both photoionization and photohomolysis (to give H) occur, with photoionization predominating (by a factor of 2.6 for dopa). Ionization of one of the phenolic hydroxyl groups increases the yield of radicals by a factor of 2. Action spectra and quantum yields for radical production are reported.     

EFFECT OF INTENSITY AND WAVELENGTH OF FLUORESCENT LIGHT ON CHROMOSOME DAMAGE IN CULTURED MOUSE CELLS

Abstract—A single 3- to 20-hr exposure of line NCTC 9266 mouse cells to cool-white fluorescent light (4.6 W/m²) produces chromatid breaks and exchanges. The effective wavelength is in the visible range and coincides with the mercury emission peak at 405 nm. Increasing light intensity from 4.6 W to 15.3 W/m² for 20 h causes a concomitant increase both in production of chromosome damage and formation of hydrogen peroxide (H₂O₂) in the serum-free medium. Cells washed free of medium and illuminated in saline for 3 h show chromosome damage to the same extent as cells illuminated in culture medium. Addition of catalase during the exposure period of 3 h eliminates the light-induced damage. We conclude that the light-induced chromatid breaks and exchanges result from H₂O₂ production within the cell and that exogenous catalase can enter the cell and prevent the damage.     

BIOSYNTHETIC INCORPORATION OF M-FLUOROTYROSINE INTO BACTERIORHODOPSIN

Halobacterium halobium, grown in a defined medium where tyrosine had been largely replaced with m-fluorotyrosine, biosynthetically produced purple membrane. Analysis of this membrane by high pressure liquid chromatography of phenylthiocarbamyl derivatized amino acids of membrane acid hydrolysates revealed that up to 50% of the tyrosine was present as the m-fluorotyrosine form. Yields of the purple membrane decreased as the level of incorporation increased. The experimental purple membrane showed a single 19F NMR resonance at -61.983 ppm (relative to trifluoroacetic acid). The bacteriorhodopsin (bR) in the purple membrane was normal as assayed by gel electrophoresis, isoelectric focusing, circular dichroic spectra, and UV-visible spectra. However, the fluorinated tyrosine bacteriorhodopsins at near neutral pH exhibited slightly slower rates of proton uptake and a slower M-state decay with biphasic kinetics reminiscent of alkaline solutions of bR (pH > 9). These results imply that the tyrosines in bacteriorhodopsin may play a role in the photoactivated proton translocation process of this pigment.     

A SPIN TRAPPING STUDY OF THE PHOTOCHEMISTRY OF 5,5-DIMETHYL-1-PYRROLINE N-OXIDE (DMPO)

The photochemistry of 5,5-dimethyl-l-pyrroline N -oxide (DMPO) has been studied in benzene, cyclohexane and aqueous buffer solutions (pH 7.4) by means of electron paramagnetic resonance (EPR) and the spin trapping technique. Ultraviolet irradiation of DMPO in aqueous buffer with unfiltered UV radiation from a Xe arc lamp results in photoionization of the spin trap and the generation of the DMPO cation radical, DMPO⁺. The aqueous electron, e_aq⁻, was trapped by DMPO and detected as the DMPO/H adduct. The DMPO⁺- reacted with the water to yield the DMPO/OH adduct. Ultraviolet irradiation of DMPO in nitrogen-saturated benzene gave an unidentified carbon-centered DMPO adduct that was replaced by hydroperoxyl and alkoxyl adducts of DMPO when oxygen was present. Experiments employing ¹⁷O₂ gas indicated that the oxygen in the DMPO alkoxyl adduct was derived from molecular oxygen. However, UV irradiation of DMPO in cyclohexane yielded the cyclohexyl and cyclohexyloxyl adducts of DMPO in nitrogen-saturated and air-saturated solutions, respectively. These observations suggest that in aprotic solvents UV irradiation of DMPO generates a carbon-centered radical (R⁻), derived from the trap itself, which in benzene reacts with oxygen to yield an alkoxyl radical (RO⁻), possibly via a peroxyl radical (ROO⁻) intermediate. In cyclohexane R⁻ abstracts a hydrogen atom from the solvent to yield the cyclohexyl radical in the absence of oxygen and the cyclohexyloxyl radical in the presence of oxygen. These findings indicate that when DMPO is used as a spin trap in studies employing short-wavelength UV radiation (λ < 300 nm) the photochemistry of DMPO cannot be ignored.     

INCORPORATION OF 11,12-DIHYDRORETINAL INTO THE RETINAE OF VITAMIN A DEPRIVED RATS

Abstract— The incorporation of 11,12-[15–³H]-dihydroretinal, a retinal in which the crucial 11-ene is saturated, into the retinae of vitamin A deficient rats as a result of intraperitoneal injection of the corresponding alcohol was shown by the presence of the tritium label in the rod outer segments and by identification of the extracted retinals using high pressure liquid chromatography. The amplitude of the electroretinogram (ERG) b-wave, diminished as the result of vitamin A deprivation, was not affected by administration of the analogue, although similar treatment of deprived litter mates with trans retinal restored the ERG b-wave amplitude to a normal level.
The evidence that the analogue is bound to opsin forming 11,12-dihydrorhodopsin is as follows: (1) when incubated with 11- cis retinal, extracts from vitamin A deficient rats regenerate 1.4 nmol rhodopsin while extracts from rats deficient in vitamin A and supplemented with 11,12-dihydroretinal regenerate 0.6 nmol rhodopsin indicating binding of the dihydroretinal blocks rhodopsin regeneration. (2) 11,12-dihydroretinal is shown to remain unchanged in hexane-washed retinae after extraction with methylene chloride and (3) injection of retinal into animals previously injected with 11,12-dihydroretinal also fails to restore visual sensitivity as measured by the ERG b-wave. Our results indicate that the dihydro-chromophore occupies the same binding site as the natural 11- cis retinal and that occupation of the chromophore binding site of opsin is not sufficient to restore the visual sensitivity in a vitamin-A-deprived animal.     

PICOSECOND FLUORESCENCE SPECTROSCOPY ON INCORPORATION PROCESSES OF HEMATOPORPHYRIN DERIVATIVE INTO MALIGNANT TUMOR CELLS in vitro

Abstract— By using a highly sensitive streak-camera technique, we investigate incorporation processes of HpD into malignant tumor m-KSA cells in vitro. The picosecond decays of the total fluorescence spectra, the wavelength-resolved fluorescence decays and the time-resolved fluorescence spectra from HpD in the cells are measured as a function of the incubation time. The results show that the aggregate component of HpD which has a fast fluorescence lifetime of 100 ps and a red-shifted band of ∼ 660 nm selectively accumulates more and more in the cells with the increase of the incubation time.     

TS-1分子筛骨架钛原子引入过程的研究

采用廉价的四丙基溴化铵（ＴＰＡＢｒ）为模板剂合成了ＴＳ１分子筛．在晶化过程中,运用ＸＲＤ,ＩＣＰ,ＩＲ,２９ＳｉＭＡＳＮＭＲ和ＵＶＶｉｓ光谱等表征手段,系统地研究了钛原子引入分子筛骨架的机制,观察到钛原子随分子筛的形成同步进入骨架的规律．另外,尽管在晶化初期固相中没有ＴｉＯ２结晶出现,但存在分散态的ＴｉＯｘ物种．随晶化时间的延长,液相中钛物种之间聚合的几率增加,使固相中ＴｉＯ２晶体不断形成．     

ENRICHMENT OF THE SPINDLE FIBER ATTACHMENTS FROM MOUSE LIVER CELL NUCLEI

Spindle fiber attachments (SFAs) were enriched in a fraction of nuclei isolated from mouselivers. The enrichment method combines sonication, treatment with 2 mol/L NaCl and highspeed centrifugation. SFA was enriched 27-fold on the average (n = 4) when measured byradioimmunoassay. The basic method offers opportunities for further increases of yield andfor the enrichment of SFA uf other vertebrates.     

STUDIES ON INTRODUCTION OF FOREIGN GENES INTO CULTURED CELLS OF Oryza sativa INDICA USINC Agrobacterium Ti PLASMID SYSTEM

Either bacterial attachment or cellulose fibrillar elaboration was hardly observedduring the cocultivation of the cultured suspension cells of Oryza sativa Indica with thestrain C58C1 Rif~r of Agrobacterium tumefaciens that was not specially pretreated. Onthe other hand, quite a lot of Agrobacterium cells were found to adhere to the surface ofcultured rice cells and a number of cellulose fibrils were produced around the specifiedbacteria when phenolics-pretreated bacteria were cocultivated with rice suspension cellsin common culture media, especially in complex culture solutions. The complex culturesolution was the bacterium-free filtrate of hormone-containing MS medium which hadbeen utilized to incubate carrot cells and the newly wounded hypocotyl segments fromtomato and Agrobacterium cells. Detecting experiments demonstrated that both NPT Ⅱ andNOS genes, located on the T-DNA segment of chimaeric plasmid pGV3850 :: 1103neo, weretransferred and expressed in the cultured cells of O. sativa Indica in the     

THE EFFECTS OF GROWTH-INHIBITING TREATMENTS ON PHOTOREPAIR IN CULTURED FISH CELLS

Abstract Goldfish cells (RBCF-1) cultured at different cell densities were harvested and their photorepair (PR) abilities were examined in terms of survival. Photorepair ability gradually increased during the phase of logarithmic growth, reaching a maximum in cells at the confluent state. This enhancement of PR ability disappeared 12 h after replating of cells in fresh medium. A number of growth-inhibiting treatments (serum depletion, UVC, hydroxyurea [HU], change in incubation temperature) were tested for their ability to induce PR. The treatment of cells with HU and serum depletion induced PR while the other treatments did not. The increase in the ability to perform PR after treatment with HU or serum depletion returned to normal levels more rapidly than that after fluorescent light treatment.     

THE USE OF SPIN LABEL OXIMETRY IN THE STUDY OF PHOTODYNAMIC INACTIVATION OF CHINESE HAMSTER OVARY CELLS

Spin label oximetry has been used to study the effects of photosensitizer hematoporphyrin on oxygen consumption in Chinese hamster ovary cells. These measurements demonstrate that, in the presence of the sensitizer, there occurs (i) an increase in total oxygen consumption during irradiation and (ii) a decrease in oxygen consumption following irradiation. These results are attributed to the effects of photosensitized oxygen consumption and decreased cellular respiration due to cell inactivation. Thus the inhibition of oxygen consumption after irradiation was shown to correlate with decreased cell survival measured in cell culture experiments. Possible mechanisms of inactivation and extensions of the oximetric approach are discussed.     

4-THIOURIDINE INCORPORATION INTO THE RNA OF MONKEY KIDNEY CELLS (CV-1) TRIGGERS NEAR-UV LIGHT LONG-TERM INHIBITION OF DNA, RNA AND PROTEIN SYNTHESIS

Monkey kidney cells (CV-1) grown for 4 h in the presence of 0.1 m M 4-thiouridine (s⁴Urd) incorporate this photoactivable uridine analog in their RNA. A minor, 5–8%, thiolated RNA fraction can be isolated from bulk RNA by affinity chromatography. This RNA fraction contains 1.5–2.5 s⁴Urd residues per 100 nucleotides and exhibits a broad chain length distribution ranging from 700 to 7000 nucleotides. It is essentially of nuclear origin and amounts to 30% of the RNA synthesized during exposure of cells to s⁴Urd. Under the same s⁴Urd labeling conditions, no thiolated pyrimidine residues have been detected in DNA.
Irradiation with 365 nm light (45 kJ/m²) of the cells immediately after s⁴Urd exposure triggers long-term inhibition of DNA, RNA and protein synthesis accompanied by a linear decline (50% in 2 days) in the total cell mass of cultured cells. In contrast, exposure to s⁴Urd alone results in moderate but reversible inhibitory effects. The available data suggest that s⁴Urd-induced photolesions in newly synthesized RNA such as RNA-RNA cross-links as well as RNA-protein bridges are directly involved in impairment of essential cellular functions.     

TRANSFORMATION OF CULTURED HUMAN FETAL GASTRIC FIBROBLASTS WITH PHYSICO-CHEMICAL AGENTS

Treatment of early passage human fetal gastric fibrohlasts with ultraviolet (UV) light and the chemical carcinogen ethyl nitrosourea (ENU) in succession resulted in an immortally growing cell line, named GTS 8502. The cells of this line display typical transformation characteristics, such as irregularly shaped nuclei, heteroploidization of karyotype and frequent appearance of heteromorphic chromosomes, the enhanced volume ratio of nucleus to cytoplasm, multinucleoli, appearance of microvilli on the surface of the cells and agglutination reaction to lectin concanavalin A. The transformants have high growing and mitotic indices and the ability of focus-formation on monolayers and anchorage independent growth in soft agar medium. Moreover, these cells induced turnouts in nude mice or in immunosuppressed new-born rats through heterotransplantation. The results of various methods including electromicroscopy and histochemical analyses indicate that GTS 8502 cells are of fibroblast origin.Our results thus indi     

THE EFFECTS OF ULTRAVIOLET AND VISIBLE RADIATION ON MOUSE ASCITES TUMOUR CELLS

Abstract— Effects of ultraviolet and visible radiation on the viability of Landschutz ascites tumour cells have been tested by growing control and treated tumour samples in adult mice. The tumour cells were irradiated as a dilute suspension in isotonic buffered salt solution, and were equilibrated at 0°C with oxygen or with nitrogen before irradiation.
Tumour cell proliferation was measured by a variety of techniques. The preferred assay-method was the growth of solid tumours in the axillae and groins of mice after sub-cutaneous inoculation of varying dilutions of treated or control ascites tumour cells. The immune response of the mice to the injected cells was reduced by whole body irradiation with a 300r dose of x-rays two days before inoculation. Results were calculated from parallel line assays using the reciprocal of the delay in appearance of the solid tumours up to 30 days post-innoculation. This reciprocal (1/T) was linearly related to the logarithm of the number of cells inoculated.
Photoreactivation has been demonstrated for this system, in which both U.V. and visible radiations were absorbed by the same cells. Light delivered alone in oxygen or in nitrogen was without effect on cell-viability, but it increased cell-survival after u.v.-irradiation in nitrogen and decreased survival after u.v.-irradiation in oxygen. Ultraviolet radiation alone was not significantly more lethal in oxygen than in nitrogen. A further observation in this work was an interaction between irradiated and control tumour cells injected into the same animal.
It is suggested that the radiation used may affect the antigenic character of the tumour cells as well as their reproductive capadity.     

INCORPORATION OF METAL IONS INTO POLYPHENYLQUINOXALINE

FeCl_3·6H_2O, NiCl_2, CuCl_2, ZnCl_2 and CrCl_3 have been incorporated into polyphenylquinoxaline by a new method. High-quality, flexible, glass-cast films have been obtained which exhibit increased glass transition temperature and excellent thermal stability. Moduli and tensile strengths of the metal-containing polyphenylquinoxaline films increase surprisingly at elevated temperature. Electrical resistivities of these films fall in the same order range as polyphenylquinoxaline alone. X-ray photoelectron spectroscopic study of metal-containing polyphcnylquinoxaiine films shows that all metals in these films are present in the ionic state, there is charge transfer between nitrogen of polyphenylquinoxaline and Cu~(2+), Zn~(2+) of CuCl_2, ZnCl_2 containing polyphenylquinoxaline films.     

THE EFFECT OF OXYGEN ON THE INCORPORATION OF MANGANESE,IRON AND COBALT INTO HEMATOPORPHYRIN IX IN GLACIAL ACETIC ACID

Abstract

The results of polarography studies for the reaction of hematoporphyrin IX with Mn(II), Fe(II) and Co(II) ion in acetic acid in the absence and the presence of oxygen are reported. The metal incorporation reaction in the presence of oxygen is quantitative for Mn and Co and incomplete for Fe. In the absence of oxygen, the Mn reaction does not proceed at all, whereas, both the Fe and Co reactions are quantitative.     

SAPO-34分子筛晶化过程中硅进入骨架的方式和机理

采用 Ｘ Ｒ Ｄ, Ｓ Ｅ Ｍ, Ｉ Ｒ 和 Ｎ Ｍ Ｒ 等手段考察了 Ｓ Ａ Ｐ Ｏ３４ 分子 筛的晶化过 程,深入研 究了晶化过程中硅进入 Ｓ Ａ Ｐ Ｏ３４ 晶格 骨架的方式和机理． 结果表明 ,在 Ｓ Ａ Ｐ Ｏ３４ 分子筛的整个晶化过程中没有 Ａｌ Ｐ Ｏ３４ 分 子筛晶相生成． 在 初始 凝 胶的 制备 过 程中, 模板 剂 的添 加和 混 合凝 胶的 老 化处 理对 Ｓ Ａ Ｐ Ｏ３４ 晶化过程的进行起着关键性的作用． 晶化前 期（ ＜ ２５ ｈ） , 硅原子直接参与晶核的形成和晶粒的长大过程,形成 Ｓｉ（４ Ａｌ） 结构,此阶段基本上 可以排除硅取代磷机理的作用; 晶化后期（ ＞ ２５ ｈ） ,少量硅以取代方式进入分子 筛骨架形成 Ｓｉ（ ｎ Ａｌ）（ ｎ ＝ ０ ～４） 多种硅结构     

ALTERATION IN CHROMATIN CONFORMATION OF PROLIFERATING AND DIFFERENTIATING ENZYME GENES IN NORMAL MOUSE LIVER AND ASCITES HEPATOMA CELLS AND ITS RELATION TO THE GENE EXPRESSION OF ENZYMES

Nuclei from the normal mouse liver were partially digested with micrococcal nuclease, followed by DNA extraction, agarose gel clectrophoresis and dot blot hybridization with ~(32)P-labeled cDNA probes of CPS_1 and ACT complex, It was clearly shown that the CPS_1 genes were distributed on the monomer, dimer. and trimer of nucleosomes, while the genes coding for ACT complex were distributed on the condensed oligonucleosomes. An opposite manner of distribution of CPS_1 and ACT complex genes was, however, noted in the case of ascites hepatoma cells, in which the specific activity of ACT was 13 times higher than that in the normal liver, while that of CPS_1 was remarkably reduced. Similar patterns of change in mRNA level of CPS_1 and ACT complex were observed in the normal mouse liver and ascites hepatoma cells, indicating a close relationship between chromatin structure and gene expression of these enzymes.     

THE CYTOTOXIC AND MUTAGENIC EFFECTS OF UVA RADIATION ON L5178Y MOUSE LYMPHOMA CELLS

Abstract— A broad-band UVA source that emits primarily350–400 nm radiation and no measurable radiation below 340 nm was used to test toxicity and mutagenicity at the thymidine kinase locus in L5178Y, subclone 3.7.2C (TK⁺/^-) mouse lymphoma cells. Cells were exposed to a fluence of 0 to 80 × 10⁴ J/m². The relationship between UVA fluence and survival was found to have a shoulder region followed by an exponential decrease in survival at higher fluence levels. An exposure-dependent increase in mutation was observed with increasing fluences from 0 to about 60 × 10⁴ J/m². An approximately 3- to 4-fold increase in mutations (trifluorothymidine resistance) over unexposed, control cells was seen at a fluence that resulted in 90% cell killing. We conclude that UVA radiation is a mutagen in the L5178Y mouse lymphoma cells used in this study.