Riboflavin sensitizer

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. 

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). 

Berliner, L.J., Ogata, T. 1997. Riboflavin-sensitized singlet oxygen formation in milk. In Food and Free Radicals (M. Hiramatsu, T. Yoshikawa, M. Inoue, eds.), pp. 119-122, Plenum Press, New York. 

An efficient transformation of maline (17) into raumacline (144), which is a biotransformation product of ajmaline in cell cultures of Rauvolfia serpentina, was developed by Endress and StSckigt (168). Ajmaline (17) was reduced with NaBH4 in citrate/phosphate buffered solution (pH 6.0) to 4,21-secoajmaline (145), which, by riboflavin-sensitized photo-oxidation, afforded raumacline (144) in 86% total yield (Scheme 13). 

Silva, E., Riickert, V., Lissi, E., and Abuin, E. (1991) Effects of pH and ionic micelles on the riboflavin-sensitized photoprocess of tryptophan in aqueous solution, J. Photochem. Photobiol. B Biol., 11, 57-68. 

The excited state of ketones can thus initiate free-radical reactions, and this is probably the mechanism for many examples of enhanced photodecomposition of environmental pollutants sensitized by acetone or other simple carbonyl compounds. A good example of such reactions is the acetone-promoted photooxidation of atrazine (24) and related triazine herbicides described by Burkhard and Guth (1976). In water, atrazine absorbs almost no solar UV and was accordingly quite stable to photolysis, but in the presence of large amounts of acetone (about 0.13 M), its half-life was decreased to about 5 hr. The produets were N-dealkylation products and ring-hydroxylated triazines. Similar products were also identified in riboflavin-sensitized photooxidation of triazines (Rejto et al., 1983). Presumably, a principal mechanism of photodecomposition would be H-abstraction from the N-alkyl substituents of atrazine, perhaps in conjunction with electron transfer from the unshmed pairs of the nitrogen atoms. 

Foote [68] has proposed a slightly different mechanism for the initiation of photo-sensitized oxidation. In this mechanism, the sensitizer forms an active complex with oxygen, which then attacks the fatty acid. This process is shown below and is believed to function in the riboflavin sensitized oxidation process. 

Use of physiological functional indices in relation to riboflavin deficiency (analogous to dark adaptation for vitamin A clotting factors for vitamin K, etc.) has not proved possible, because the analogous riboflavin-sensitive physiological processes are insufficiently specific or easily measurable for use in population studies. Of the biochemical indices, urinary excretion and the flavin-dependent enzyme, erythrocyte glutathione reductase, are generally considered to be the front-runners in the race for acceptance in human studies. These have already been described in the previous section. 

The microbial assay is based on the growth of l ctobacillus casei in the natural (72) or modified form. The lactic acid formed is titrated or, preferably, the turbidity measured photometrically. In a more sensitive assay, l euconostoc mesenteroides is employed as the assay organism (73). It is 50 times more sensitive than T. casei for assaying riboflavin and its analogues (0.1 ng/mL vs 20 ng/mL for T. casei). A very useful method for measuring total riboflavin in body fluids and tissues is based on the riboflavin requirement of the proto2oan cHate Tetrahjmenapyriformis which is sensitive and specific for riboflavin. 

In biological systems, sensitizers such as porphyrins, chlorophylls, and riboflavin can sensitize 02 production and this can lead to deleterious effects including DNA damage and lipid peroxidation. 

The recognition of their structure permits the determination of vitamins by the tools of analytical chemistry, but while such methods are widely used in industrial production, the minute quantities in body fluids and tissues limit the purely chemical approach to a few members of this group present in relatively high concentration, e.g., vitamin C (K5). Microchemical methods are in use for the determination of thiamine, riboflavin, and some of the fat-soluble vitamins, based on the most sensitive colorimetric and, in particular, fluorometric techniques. Vitamin D, on the other hand, is determined by animal assay. 

Note Riboflavin solutions are fight sensitive. Store them in the dark as you prepare them. 

The SP procedure of water-soluble vitamins from multivitamin tablets is particularly challenging due to the diverse analytes of varied hydrophobicities and pfC. Water-soluble vitamins (WSVs) include ascorbic acid (vitamin C), niacin, niacinamide, pyridoxine (vitamin B ), thiamine (vitamin Bj), folic acid, riboflavin (vitamin B2) and others. While most WSVs are highly water soluble, riboflavin is quite hydrophobic and insoluble in water. Folic acid is acidic while pyridoxine and thiamine are basic. In addition, ascorbic acid is light sensitive and easily oxidized. The extraction strategy employed was a two-step approach using mixed solvents of different polarity and acidity as follows ... 

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). 

Photolytic. Acher et al. (1981) studied the dye-sensitized photolysis of terbacil in aerated aqueous solutions over a wide pH range. After a 2-h exposure to sunlight, terbacil in aqueous solution (pH range 3.0-9.2) in the presence of methylene blue (3 ppm) or riboflavin (10 ppm) decomposed to 3-ter/-5-butyl-5-acetyl-5-hydroxyhydantoin. Deacylation was observed under alkaline conditions (pH 8.0 or 9.2) affording 3-/er/-5-hydroxyhydantoin. In neutral or acidic conditions (pH 6.8 or 3.0) containing riboflavin, a mono-Wdealkylated terbacil dimer and an unidentified water-soluble product formed. Product formation, the relative amounts of products formed, and the rate of photolysis were all dependent upon pH, sensitizer, temperature, and time (Acher et ah, 1981). 

Thiamine, riboflavin, nicotinamide, pyridoxine, and folic acid can be determined together by using DAD, but pantothenic acid and biotin do not have adequate sensitivity for UV detection in complex matrices. 

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. 

Some analytes, such as riboflavin (vitamin B2)16 and polycyclic aromatic compounds (an important class of carcinogens), are naturally fluorescent and can be analyzed directly. Most compounds are not luminescent. However, coupling to a fluorescent moiety provides a route to sensitive analyses. Fluorescein is a strongly fluorescent compound that can be coupled to many molecules for analytical purposes. Fluorescent labeling of fingerprints is a powerful tool in forensic analysis.17 Sensor molecules whose luminescence responds selectively to a variety of simple cations and anions are available.18 Ca2+ can be measured from the fluorescence of a complex it forms with a derivative of fluorescein called calcein. 

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. 

Riboflavin is stable to heat but is extremely sensitive to light, a fact of some nutritional significance. Do not leave bottled milk in the sunshine (see Fig. 15-8) Many products of photolysis are formed (Fig. 3-5). Among them is lumichrome. 

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. 

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