Riboflavin irradiation

Melo, T. B. lonescu, M. A. Haggquist, G. W. Naqvi, K. R. (1999). Hydrogen abstraction by triplet flavins. I time-resolved multi-chaimel absorption spectra of flash-irradiated riboflavin solutions in water Spectrochimica Acta, Part A Molecular and Biomolecular Spectroscopy, Vol.55, No.ll, (September 1999), pp. 2299-2307, ISSN 1386-1425. 

Chlorinated dibenzo ip-dioxins are contaminants of phenol-based pesticides and may enter the environment where they are subject to the action of sunlight. Rate measurements showed that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is more rapidly photolyzed in methanol than octachlorodi-benzo-p-dioxin. Initially TCDD yields 2,3,7-trichlorodiben-zo-p-dioxin, and subsequent reductive dechlorination is accompanied by ring fission. Pure dibenzo-p-dioxin gave polymeric material and some 2,2 -dihydroxybiphenyl on irradiation. Riboflavin-sensitized photolysis of the potential precursors of dioxins, 2,4-dichlorophenol and 2,4,5-trichloro-phenol, in water gave no detectable dioxins. The products identified were chlorinated phenoxyphenols and dihydroxy-biphenyls. In contrast, aqueous alkaline solutions of purified pentachlorophenol gave traces of octachlorodibenzo-p-dioxin on irradiation. 

The scavenging effect of berbamine on active oxygen radicals was studied via a spintrapping technique and a chemiluminescence (CL) method in phorbol myristate acetate (PMA) stimulated polymorphonuclear leukocytes (PMN) and in four-cell superoxide (02+) or hydroxyl radical (OH ) generating systems. The alkaloid (0.1-0.3 mM) effectively reduced active oxygen radicals in PMA-stimulated PMN, but had no obvious effect on oxygen consumption during the respiratory burst of PMN (as measured with spin probe oxymetry). In addition, berbamine (0.3 mM) inhibited the CL response of PMA-stimulated PMN, and quenched 02 in the xanthine/xanthine oxidase and irradiation riboflavin systems, as well as OH in the Fenton... 

Photochemical decomposition of riboflavin in neutral or acid solution gives lumichrome (3), 7,8-dimethyl all oxazine, which was synthesized and characterized by Karrer and his co-workers in 1934 (11). In alkaline solution, the irradiation product is lumiflavin (4), 7,8,10-trimethyhsoalloxazine its uv—vis absorption spectmm resembles that of riboflavin. It was prepared and characterized in 1933 (5). Another photodecomposition product of riboflavin is 7,8-dimethy1-10-foTmylmethy1isoa11oxazine (12). 

Riboflavin can be assayed by chemical, en2ymatic, and microbiological methods. The most commonly used chemical method is fluorometry, which involves the measurement of intense yeUow-green fluorescence with a maximum at 565 nm in neutral aqueous solutions. The fluorometric deterrninations of flavins can be carried out by measuring the intensity of either the natural fluorescence of flavins or the fluorescence of lumiflavin formed by the irradiation of flavin in alkaline solution (68). The later development of a laser—fluorescence technique has extended the limits of detection for riboflavin by two orders of magnitude (69,70). 

Figure 1 shows the graphs of the PCL that were recorded with riboflavin as the photosensitizer and luminol as the detector for free radicals [21], The course of the PCL reaction has two maxima at approximately 30 s and 3 min after the start of irradiation. It has been demonstrated by analysis of kinetics after addition of the reactants at varying times that the first maximum is riboflavin-dependent. Luminol is needed only for visualization of the superoxide radicals. 

Figure 1 PCL graphs with riboflavin (R) as the photosensitizer and luminol (L) as the detector for free radicals. I = start of irradiation. (From Ref. 21.)...

Beadle and Tatum had found that irradiation of Neurospora spores produced mutants which were incapable of carrying out certain well-defined chemical reactions, and it was at first supposed that as a result of the destruction of a specific gene, the potentiality for producing a particular enzyme was completely lost. The "wild type" of Neurospora could propagate satisfactorily when biotin was the only vitamin-like substance supplied in the culture medium. Of the many mutant strains produced, however, one needed, in addition to biotin, the vitamin riboflavin. Without a supply of riboflavin in the culture medium this so-called "riboflavinless mutant" would not grow. Since riboflavin is a part of an enzyme system always found in Neurospora, it is an obligatory cell constituent and either has to be produced by the cells themselves (as in the wild type) or supplied exogenously in... 

Carell has recently presented the study of a flavin amino acid chimera to model riboflavin in DNA photolyases [68]. This amino acid LI (Fig. 20) was synthesized in an enantiopure fashion by building the alloxazine ring onto the epsilon amine of lysine. This coenzyme chimera was applied to the problem of repairing DNA damage caused by UV irradiation. LI was incorporated into an 21-residue peptide, P-1, possessing the sequence of the DNA-binding domain of the helix-loop-helix transcription factor MyoD. 

An aqueous solution of amitrole can decompose in the following free radical systems Fenton s reagent, UV irradiation, and riboflavin-sensitized photodecomposition (Plimmer et al, 1967). Amitrole-5- C reacted with Fenton s reagent to give radiolabeled carbon dioxide, unlabeled urea, and unlabeled cyanamide. Significant degradation of amitrole was observed when an aqueous solution was irradiated by a sunlamp (L = 280-310 nm). In addition to ring compounds, it was postulated that other products may have formed from the polymerization of amitrole free radicals (Plimmer et al., 1967). 

Photolytic. When a dilute aqueous solution (1-10 mg/L) of bromacil was exposed to sunlight for 4 months, the TV-dealkylated photoproduct, 5-bromo-6-methyluracil, formed in small quantities. This compound is less stable than bromacil and upon further irradiation, the de-brominated product, 6-methyluracil was formed (Moilanen and Crosby, 1974). Acher and Dunkelblum (1979) studied the dye-sensitized photolysis of aerated aqueous solutions of bromacil using sunlight as the irradiation source. After 1 h, a mixture of diastereoisomers of 3-5ec-butyl-5-acetyl-5-hydroxyhydantoin formed in an 83% yield. In a subsequent study, another minor intermediate was identified as a 5,5 -photoproduct of 3-5ec-butyl-6-methyluracil. In this study, the rate of photooxidation increased with pH. The most effective sensitizers were riboflavin (10 ppm) and methylene blue (2-5 ppm) (Acher and Saltzman, 1980). Direct photodegradation of bromacil is not significant (Acher and Dunkelblum, 1979 Ishihara, 1963). 

Photolytic. When propachlor in an aqueous ethanolic solution was irradiated with UV light (>, = 290 nm) for 5 h, 80% decomposed to the following cyclic photoproducts W-isopropyloxindole, W-isopropyl-3 hydroxyoxindole, and a spiro compound. Irradiation of propachlor in an aqueous solution containing riboflavin as a sensitizer resulted in completed degradation of the parent compound. 3-Hydroxypropachlor was the only compound identified in trace amounts which formed via ring hydroxylation (Rejtb et al, 1984). Hydrolyzes under alkaline conditions forming W-isopropylaniline (Sittig, 1985) which is also a product of microbial metabolism (Novick et al., 1986). 

The same authors (G8, G7) also found very substantial decreases in riboflavin (approx. 80%), and niacin (P9) fared little better. When mixtures were irradiated unusual events occurred. Riboflavin and ascorbic acid were each protected by niacin. Addition of cystine or cysteine apparently sensitized the niacin (P10). Since initial rates were not given, and the doses were considerably above the oxygen breakpoint (Sec. IIIA2), no mechanistic interpretation is possible. There also appears to be some doubt about the reliability of the colormetric assay used by these workers. 

Riboflavin (CCLV) is photosensitive on irradiation in alkaline solution, it is converted into lumiflavin (CCLVI)302 and in neutral or acid solution, lumichrome (CCLVII) is produced.147 The photolysis of 9-(2 -hydroxyethyl)isoalloxazine (CCLVIII) is also a general aeid- and base-catalyzed reaction. 

Oster [174] proposed the second hypothesis to explain his results on the photopolymerization of acrylonitrile in aqueous solution, buffered at pH 7.0, and sensitized by xanthene dyes and riboflavin using ascorbic acid as the reducing agent. Whereas the monomer is efficiently polymerized when the solution is illuminated in the presence of oxygen, irradiation in its absence leads to photoreduction of the dye to its leuco form but no polymer is formed. Therefore, the author suggests that the leuco dye reacts with atmospheric... 

Evidence supporting this mechanism is presented for the case of acrylamide polymerization sensitized by riboflavin, but not for the case of fluorescein and its halogenated derivatives. Irradiation with a millisecond flash in the presence of air leads to polymer formation after an induction period of one hour. In contrast, when the irradiation is carried out with degassed solutions, polymerization starts only after the sample is exposed to atmospheric oxygen. 

Figure 3-5 Photograph of a two-dimensional thin layer (silica gel) chromatogram of a mixture of flavins formed by irradiation of 10 pg of the vitamin riboflavin. The photograph was made by the fluorescence of the compounds under ultraviolet light. Some riboflavin (RF) remains. The arrows indicate the location of the sample spot before chromatography. Chromatography solvents a mixture of acetic acid, 2-hutanone, methanol, and benzene in one direction and M-butanol, acetic acid, and water in the other. See Treadwell et al.H)2...

Irradiation of olefins and dienes in the presence of oxygen and various sensitizers produces hydroperoxides and cyclic peroxides. Effective sensitizers include not only high-energy carbonyl triplets, but also low-energy organic dyes such as methylene blue, rose bengal, chlorophyl, and riboflavin. The same species that quench the phosphorescence of these complex molecules also quench their photosensitizing ability. 

Zinc(II) cyclene was linked to phenothiazene group which served as electron-donor (Scheme 22). To the complexed zinc(II), a riboflavin tetraacetate molecule coordinated. Upon irradiation, the flavin became a strong oxidant and the transfer of electrons could be easily observed by emission quenching. 

Photosensitization for the removal of certain pollutants in photolytic processes can contribute significantly to the degradation rate. Thus, Simmons and Zepp [88] observed increases of up to 26 times of the photodegradation rates of nitroaromatic compounds due to the action of natural or commercial humic substances with solar irradiation. In another work [89], the herbicide 4-chloro-2-methylphenoxyacetic acid (MCPA) was irradiated in water with 300 nm light in the presence of different photosensitizers. This compound, which does not photolyze directly at this wavelength, could be degraded more than 95% in 5 hr when riboflavin was used as photosensitizer. 

The catalysis of riboflavin by the macromolecules is partially attributed to reversible binding of excited riboflavin molecules to macromolecules, which produce longer-lived excited species. Evidence is presented to indicate involvement of a triplet state in both the photobinding and photodecomposition of riboflavin. The enhanced binding of riboflavin to macromolecules during irradiation was detected by both precipitation and resin desorption techniques. 

Several electron donors form molecular complexes with menadione, which then suppress its photodecomposition in aqueous solutions (107). DNA-daunomycin complexes exposed to UV irradiation are reported by Gray and Philips (127) to be more stable than daunomycin alone. The photostability of nifedipine increases on the addition of PVP (64,128). However, the rate of photobleaching of riboflavin by VIS radiation is considerably enhanced by PVP (28). 

When amino acids in parenteral solutions are exposed to relatively intense illumination for 24 hours, simulating phototherapy in neonatal nurseries, most individual amino acids decrease only slightly. Decreases in the concentrations of methionine, proline, tryptophan, and tyrosine are significantly greater in the presence of riboflavine. The observed decreases in amino acid concentrations are unlikely to be nutritionally important. However, in view of the possibility that photooxidation products may exert toxic effects, it is best to shield amino acid solutions containing vitamins from strong sources of UV-VIS irradiation (86). 

Under aerobic conditions, irradiation of phenothiazine in the presence of riboflavin leads to its photo-oxidation to the corresponding sulphoxide, presumably via the agency of singlet oxygen. The reaction is retarded by /3-cyclodextrin although not by a- and y-cyclodextrins, and this has been... 

Prepare the gel solution. For each 10 mL, mix 1.8 mL of stock acrylamide solution with 0.25 mL of ampholyte solution. Make up to vol with water and degas for 2 min under vacuum. Add 0.1 mL of riboflavin, pour into the gel mold, and irradiate with the UV lamp about 50-70 cm from the gel. Polymerization takes about 1.5-2.5 h. 

The B group vitamins, thiamine and riboflavin, are destroyed on irradiation in dilute aqueous solutions. Riboflavin is reduced in air-free solutions to a semiqui-none form. Nicotinic acid is decarboxylated on irradiation in air-saturated aqueous solutions. 

Pyridoxine Hydrochloride, USP. Pyridoxine hydrochloride.. S-hydroxy-6-methyl-3.4-pyridinedimethanol hydrochloride. vitamin B, hydrochloride, rat antidermatitis factor. is a white, odorless, crystalline substance that is soluble l .S in water and 1 100 in alcohol and in.soluble in ether. It is relatively. stable to light and air in the solid form and in acid solutions at a pH no greater than S. at which pH it can be autoclaved at IS pounds at I20°C for 20 to 30 minutes. Pyridoxine is unstable when irradiated in aqueous solution.s at pH 6.8 or above. It is oxidized readily by hydrogen peroxide and other oxidizing agents. Pyridoxine is as stable in mixed vitamin preparations as riboflavin and nicotinic acid. A 1% aqueous solution has a pH of 3. The pK i values fur pyridoxine. pyridoxal. and pyridoxamine are S.OO. 4.22. and 3.40. respectively, and their pK 2 values are 8.96. 8.68. and 8.05. respectively. 

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