Iron-thiolate induced oxidation of methionine to methionine sulfoxide in small model peptides. Intramolecular catalysis by histidine.

Peptides containing either glycine and methionine, or glycine, methionine and histidine at various locations were oxidized by the dithiothreitol/ferric chloride system in phosphate buffer. The yields of peptide degradation and sulfoxide formation were measured as a function of peptide sequence and pH. In general little change of the final yields of peptide degradation is observed whereas the final yields of sulfoxide formation progressively decrease on going from pH 6.0 to 8.0. The pH profiles vary with the structure of the respective peptide. Efficient sulfoxide formation occurred when histidine and methionine were present within the same peptides sequence, and particularly when methionine was located at the C-terminus of the peptide. Added superoxide dismutase, catalase, and methanol did neither promote nor inhibit both the degradation of peptide and the formation of sulfoxide excluding free superoxide, hydrogen peroxide, and hydroxyl radicals as responsible reactive oxygen species. The observations are rationalized by invoking a pH-dependent conversion of an efficiently sulfoxide yielding oxidant into another oxidant which still degrades peptides but does not form methionine sulfoxide. The first might be a metal-bound peroxide or peroxyl species which converts into a metal-bound or 'complexed' hydroxyl radical.

In vivo aging of rat skeletal muscle sarcoplasmic reticulum Ca-ATPase. Chemical analysis and quantitative simulation by exposure to low levels of peroxyl radicals.

Sarcoplasmic reticulum (SR) Ca-ATPase of young adult (5 months) and aged (28 months) Fischer 344 male rat skeletal muscle was analyzed for posttranslational modifications as a result of biological aging and their potential functional consequences. The significant differences in the amino acid composition were a 6.8% lower content of sulfhydryl groups and a ca. 4% lower content of Arg residues of the Ca-ATPase from old as compared to young rats. Based on a total of 24 Cys residues the difference in protein thiols corresponds to a loss of 1.5 mol Cys/mol Ca-ATPase as a result of in vivo aging. The loss of Cys residues was not accompanied by a loss of enzyme activity though the 'aged' Ca-ATPase was more sensitive to heat inactivation, aggregation, and tryptic digestion. A comparison of the total sulfhydryl content of all SR proteins present revealed a 13% lower amount for SR vesicles isolated from aged rats. Compared to the alterations of Cys and Arg, there was only a slight and probably physiologically insignificant increase of protein carbonyls with aging, i.e. from 0.32 to 0.46 mol carbonyl groups per mol of Ca-ATPase. When SR vesicles from young rats were exposed to AAPH-derived peroxyl radicals, there was a loss of ca. 1.38 x 10(-4) M total SR sulfhydryl groups per 4 mg SR protein/ml (corresponding to ca. 25%) and a loss of 9.6 x 10(-5) M Ca-ATPase sulfhydryl groups (corresponding to ca. 31%) per 1.6 x 10(-5) M initiating peroxyl radicals, indicating that the stoichiometry of sulfhydryl oxidation was > or = 6 oxidized thiols per initiating AAPH-derived peroxyl radical. Besides Cys, the exposure to AAPH-derived radicals caused a slight loss of Ca-ATPase Arg, Met, and Ser residues. Most importantly, the SR Ca-ATPase exposed to this low concentration of peroxyl radicals displayed physical and functional properties quantitatively comparable to those of SR Ca-ATPase isolated from aged rats, i.e. no immediate loss of activity, increased susceptibility to heat inactivation, aggregation, and tryptic digestion. Moreover, a comparison of kinetically early tryptic fragments by HPLC-electrospray MS and N-terminal sequencing revealed that similar peptide fragments were produced from 'aged' and AAPH-oxidized Ca-ATPase which were not (or kinetically significantly later) generated from the 'young' Ca-ATPase, suggesting some conformational changes of the Ca-ATPase as a result of aging and AAPH-exposure. All except one of these peptides originated from locations remote from the nucleotide-binding and calcium-binding sites. The latter results suggest that aging and AAPH-exposure may target similar Cys residues, mainly at locations remote from the nucleotide-binding and calcium-binding sites, rationalizing the fact that Cys oxidation did not immediately cause inactivation of the Ca-ATPase. Our results provide a quantitative estimate of a net concentration of reactive oxygen species, here peroxyl radicals, which induces physical and chemical alterations of the SR Ca-ATPase quantitatively comparable to those induced by in vivo aging.

Exploring oxidative modifications of tyrosine: an update on mechanisms of formation, advances in analysis and biological consequences.

Protein oxidation is increasingly recognised as an important modulator of biochemical pathways controlling both physiological and pathological processes. While much attention has focused on cysteine modifications in reversible redox signalling, there is increasing evidence that other protein residues are oxidised in vivo with impact on cellular homeostasis and redox signalling pathways. A notable example is tyrosine, which can undergo a number of oxidative post-translational modifications to form 3-hydroxy-tyrosine, tyrosine crosslinks, 3-nitrotyrosine and halogenated tyrosine, with different effects on cellular functions. Tyrosine oxidation has been studied extensively in vitro, and this has generated detailed information about the molecular mechanisms that may occur in vivo. An important aspect of studying tyrosine oxidation both in vitro and in biological systems is the ability to monitor the formation of oxidised derivatives, which depends on a variety of analytical techniques. While antibody-dependent techniques such as ELISAs are commonly used, these have limitations, and more specific assays based on spectroscopic or spectrometric techniques are required to provide information on the exact residues modified and the nature of the modification. These approaches have helped understanding of the consequences of tyrosine oxidation in biological systems, especially its effects on cell signalling and cell dysfunction, linking to roles in disease. There is mounting evidence that tyrosine oxidation processes are important in vivo and can contribute to cellular pathology.

The metal-catalyzed oxidation of methionine in peptides by Fenton systems involves two consecutive one-electron oxidation processes.

The one-electron oxidation of methionine (Met) plays an important role in the redox reactions of Met in peptides and proteins under conditions of oxidative stress, e.g., during the metal-catalyzed oxidation of beta-amyloid peptide (beta A). However, little information is available with regard to mechanisms and product formation during the metal-catalyzed oxidation of Met. Here, we demonstrate that two-electron oxidation of Met in Fenton reactions, carried out aerobically by [Fe(II)(EDTA)](2-) and H(2)O(2) (EDTA = ethylenediaminetetra acetate) is the consequence of two consecutive one-electron transfer reactions carried out by either free or complexed hydroxyl radicals, followed by the reaction of an intermediary sulfur-nitrogen bonded radical cation (sulfuranyl radical) with O(2). The model peptide Met-Met represents an ideal substrate for these investigations as its one-electron oxidation, followed by reaction with molecular oxygen, produces unique intermediates, azasulfonium diastereomers, which can be chemically isolated before hydrolysis to sulfoxide occurs.

Thiyl radicals abstract hydrogen atoms from carbohydrates: reactivity and selectivity.

Free radical damage of DNA is a well-known process affecting biological tissue under conditions of oxidative stress. Though carbohydrate-derived radicals are generally "repaired" by hydrogen transfer from thiols, the reverse possibility, namely hydrogen abstraction by thiyl radicals from carbohydrates, exists. The biological relevance of this process has been discussed controversially, especially because of the lack of rate constants. Therefore, we have measured rate constants for the hydrogen transfer reaction between thiyl radicals from cysteine and selected carbohydrates, 2-deoxy-D-ribose (dRib), 2-deoxy-D-glucose (dGls), alpha-D-glucose (Gls), and inositol (Ino). Rate constants are on the order of 10(4) M(-1)s(-1), with the highest average value for dRib, (2.7 +/- 1.0) x 10(4) M(-1)s(-1), and the lowest average value for dGls, (1.6 +/- 0.2) x 10(4) M(-1)s(-1), based on two ways of kinetic analysis, standard competition kinetics and stochastic simulation of the experimental results, respectively. In general, thiyl radicals attack preferentially the C(1)-H bond of the carbohydrates, to an extent of ca. 72% in dRib and 90% in dGls. Kinetic measurements were possible through a specifically designed competition system measuring the reaction of thiyl radicals with either the C-H bonds of the carbohydrates or the C(alpha)-H bond of cysteine under conditions where the extent of other competitive reactions of the thiyl radicals were minimized.

Diastereoselective protein methionine oxidation by reactive oxygen species and diastereoselective repair by methionine sulfoxide reductase.

Recent studies have shown that the "calcium-sensor" protein calmodulin (CaM) suffers an age-dependent oxidation of methionine (Met) to methionine sulfoxide (MetSO) in vivo. However, MetSO did not accumulate on the Met residues that show the highest solvent-exposure. Hence, the pattern of Met oxidation in vivo may give hints as to which reactive oxygen species and oxidation mechanisms participate in the oxidation of this important protein. Here, we have exposed CaM under a series of different reaction conditions (pH, [Ca(2+)], [KCl]) to various biologically relevant reactive oxygen species and oxidizing systems (peroxides, HOCl, peroxynitrite, singlet oxygen, metal-catalyzed oxidation, and peroxidase-catalyzed oxidation) to investigate whether one of these systems would lead to an oxidation pattern of CaM similar to that observed in vivo. However, generally, these oxidizing conditions led to a preferred or exclusive oxidation of the C-terminal Met residues, in contrast to the oxidation pattern of CaM observed in vivo. Hence, none of the employed oxidizing conditions was able to mimic the age-dependent oxidation of CaM in vivo, indicating that other, yet unidentified oxidation mechanisms may be important in vivo. Some oxidizing species showed a quite-remarkable diastereoselectivity for the formation of either L-Met-D-SO or L-Met-L-SO. Diastereoselectivity was dependent on the nature of the oxidizing species but was less a function of the location of the target Met residue in the protein. In contrast, diastereoselective reduction of L-Met-D-SO by protein methionine sulfoxide reductase (pMSR) was efficient regardless of the position of the L-Met-D-SO residue in the protein and the presence or absence of calcium. With only the L-Met-D-SO diastereomer being a substrate for pMSR, any preferred formation of L-Met-L-SO in vivo may cause the accumulation of MetSO unless the oxidized protein is substrate for (accelerated) protein turnover.

Nitric oxide-dependent modification of the sarcoplasmic reticulum Ca-ATPase: localization of cysteine target sites.

Skeletal muscle contraction and relaxation is modulated through the reaction of sarcoplasmic reticulum (SR) protein thiols with reactive oxygen and nitrogen species. Here, we have utilized high-performance liquid chromatography-electrospray mass spectrometry and a specific thiol-labeling procedure to identify and quantify cysteine residues of the SR Ca-ATPase that are modified by exposure to nitric oxide (NO). NO and/or NO-derived species inactivate the SR Ca-ATPase and modify a broad spectrum of cysteine residues with highest reactivities towards Cys364, Cys670, and Cys471. The selectivity of NO and NO-derived species towards the SR Ca-ATPase thiols is different from that of peroxynitrite. The efficiency of NO at thiol modification is significantly higher compared with that of peroxynitrite. Hence, NO has the potential to modulate muscle contraction through chemical reaction with the SR Ca-ATPase in vivo.

Diastereoselective reduction of protein-bound methionine sulfoxide by methionine sulfoxide reductase.

Methionine sulfoxide (MetSO) in calmodulin (CaM) was previously shown to be a substrate for bovine liver peptide methionine sulfoxide reductase (pMSR, EC 1.8.4.6), which can partially recover protein structure and function of oxidized CaM in vitro. Here, we report for the first time that pMSR selectively reduces the D-sulfoxide diastereomer of CaM-bound L-MetSO (L-Met-D-SO). After exhaustive reduction by pMSR, the ratio of L-Met-D-SO to L-Met-L-SO decreased to about 1:25 for hydrogen peroxide-oxidized CaM, and to about 1:10 for free MetSO. The accumulation of MetSO upon oxidative stress and aging in vivo may be related to incomplete, diastereoselective, repair by pMSR.

Biological aging does not lead to the accumulation of oxidized Cu,Zn-superoxide dismutase in the liver of F344 rats.

Cu,Zn-Superoxide dismutase (SOD) was isolated from the liver of 3-, 12-, and 26-month-old Fisher 344 (F344) rats. Specific activity and metal content of the enzyme, purified by ion-exchange and size-exclusion chromatography, did not significantly change with age. Electrospray ionization-mass spectrometry and amino acid analysis of Cu,Zn-SOD apoprotein, further purified by reverse-phase HPLC, showed neither significant loss of amino acids nor accumulation of oxidized isoforms with age. When bovine Cu,Zn-SOD, oxidized with H(2)O(2) in vitro, was added to rat liver homogenate, we reisolated circa 70% of the oxidized bovine Cu,Zn-SOD together with the rat isoform, showing that oxidized Cu,Zn-SOD can be recovered from tissue homogenate. Therefore, our data do not confirm an earlier hypothesis that oxidatively modified Cu,Zn-SOD protein accumulates in the liver of aged F344 rats.

Accumulation of nitrotyrosine on the SERCA2a isoform of SR Ca-ATPase of rat skeletal muscle during aging: a peroxynitrite-mediated process?

The SR Ca-ATPase in skeletal muscle SR vesicles isolated from young adult (5 months) and aged (28 months) rats was analyzed for nitrotyrosine. Only the SERCA2a isoform contained significant amounts with approximately one and four nitrotyrosine residues per young and old Ca-ATPase, respectively. The in vitro exposure of SR vesicles of young rats to peroxynitrite yielded selective nitration of the SERCA2a Ca-ATPase even in the presence of excess SERCA1a. No nitration was observed during the exposure of SR vesicles to nitric oxide in the presence of O2. These data suggest the vivo presence of peroxynitrite in skeletal muscle. The greater nitrotyrosine content of SERCA2a from aged tissue implies an age-associated increase in susceptibility to oxidation by this species.

Age-related chemical modification of the skeletal muscle sarcoplasmic reticulum Ca-ATPase of the rat.

Much emphasis has been placed on the description of age-related changes in skeletal muscle physiology. The present paper summarizes the chemical characterization of age-related post-translational modifications of the rat skeletal muscle sarcoplasmic reticulum (SR) Ca-ATPase isoforms SERCA1 and SERCA2a obtained from 5- and 28-month-old male Fischer 344 rats. Whereas the SERCA1 isoform shows an age-dependent loss of Cys and Arg, the SERCA2a isoform displays a loss of Cys but also a significant accumulation of 3-nitrotyrosine. The in vitro exposure of SR vesicles particularly rich in SERCA1 (>90%) from 5-month-old rats to low levels of peroxyl radicals yielded SR vesicles with physical properties of the SR Ca-ATPase identical to those observed for the SR Ca-ATPase obtained from 28-month-old rats. The peroxyl radical-modified SR Ca-ATPase showed a loss of Cys and Arg but also of Ser and Met, indicating that peroxyl radicals, though a good model oxidant to generate 'aged' SR vesicles, may not be the only oxidant responsible for the chemical modification of the SR Ca-ATPase in vivo. In fact, efficient thiol modification of the SERCA1 was also observed after the exposure to peroxynitrite. Peroxynitrite selectively nitrated the tyrosine residues of the SERCA2a isoform even in the presence of an excess of SERCA1. Thus, peroxynitrite may be responsible for the age-dependent modification of the SR Ca-ATPase in vivo.

Reactive oxygen species and biological aging: a mechanistic approach.

The experimental evidence for an age-dependent increased generation of reactive oxygen species and a progressive accumulation of oxidized biomolecules is growing. However, despite such facts there is no detailed mechanistic information on how the higher availability of reactive oxygen species translates into the accumulation of oxidized biomolecules. For example, open questions are which reactive oxygen species are responsible for what types of oxidation products in vivo, under what specific reaction conditions can we expect which reaction products, and why specifically are modified biomolecules eliminated whereas others accumulate? Mitochondria appear to serve as the major source for reactive oxygen species in aging tissue. Genetic experiments have demonstrated an effect of Cu,ZnSOD on life span and in the prevention of age-related oxidative damage, suggesting that extramitochondrial superoxide promotes biological aging. However, as superoxide does not easily cross membranes, potential chemical pathways that convert mitochondrial reactive oxygen species into superoxide outside the mitochondria are displayed. The chemical reactivity of individual reactive oxygen species with the amino acid side chain of methionine is surveyed to obtain mechanistic details on the oxidation pathways potentially leading to the age-dependent methionine oxidation of the protein calmodulin in vivo. It will evolve that the in vivo accumulation of oxidized calmodulin cannot be the result of the reaction of an individual reactive oxygen species with calmodulin in homogenous solution alone. Complexation of calmodulin to calmodulin-binding proteins and protein turnover are additional parameters likely contributing to the accumulation of specifically modified calmodulin.

The oxidative inactivation of sarcoplasmic reticulum Ca(2+)-ATPase by peroxynitrite.

The oxidative inactivation of rabbit skeletal muscle Ca(2+)-ATPase in sarcoplasmic reticulum (SR) vesicles by peroxynitrite (ONOO-) was investigated. The exposure of SR vesicles (10 mg/ml protein) to low peroxynitrite concentrations ( < or = 0.2 mM) resulted in a decrease of Ca(2+)-ATPase activity primarily through oxidation of sulfhydryl groups. Most of this deactivation (ca.70%) could be chemically reversed by subsequent reduction of the enzyme with either dithiothreitol (DTT) or sodium borohydride (NaBH4), indicating that free cysteine groups were oxidized to disulfides. The initial presence of 5 mM glutathione failed to protect the SR Ca(2+)-ATPase activity. However, as long as peroxynitrite concentrations were kept < or = 0.45 mM, the efficacy of DTT to reverse Ca(2+)-ATPase inactivation was enhanced for reaction mixtures which initially contained 5 mM glutathione. At least part of the disulfides were formed intermolecularly since gel electrophoresis revealed protein aggregation which could be reduced under reducing conditions. The application of higher peroxynitrite concentrations ( > or = 0.45 mM) resulted in Ca(2+)-ATPase inactivation which could not be restored by exposure of the modified protein to reducing agents. On the other hand, treatment of modified protein with NaBH4 recovered all SR protein thiols. This result indicates that possibly the oxidation of other amino acids contributes to enzyme inactivation, corroborated by amino acid analysis which revealed some additional targets for peroxynitrite or peroxynitrite-induced processes such as Met, Lys, Phe, Thr, Ser, Leu and Tyr. Tyr oxidation was confirmed by a significant lower sensitivity of oxidized SR proteins to the Lowry assay. However, neither bityrosine nor nitrotyrosine were formed in significant yields, as monitored by fluorescence spectroscopy and immunodetection, respectively. The Ca(2+)-ATPase of SR is involved in cellular Ca(2+)-homeostasis. Thus, peroxynitrite mediated oxidation of the Ca(2+)-ATPase might significantly contribute to the loss of Ca(2+)-homeostasis observed under biological conditions of oxidative stress.

Decreased plasma membrane calcium transport activity in aging brain.

We have assessed the functional properties of both calmodulin (CaM) and the plasma membrane Ca(2+)-ATPase in brains of young, middle aged, and old Fisher 344 rats. Under optimal conditions of saturating Ca2+ and ATP, the CaM-activated Ca(2+)-ATPase activity was decreased with increasing age, particularly when CaM isolated from the brains of aged rats was used to stimulate the enzyme. In the case of CaM, structural modifications within the primary sequence of the protein from aged brains were identified. We found that during normal biological aging approximately 6 methionine residues were modified to their corresonding sulfoxide per CaM, and no other amino acids were modified. Some aspects of the age-related decline in the effectiveness of CaM as an activator of Ca(2+)-ATPase could be simulated using a range of reactive oxygen species (including hydrogen peroxide and oxoperoxynitrite) and, in the latter case, the extent of oxidative modification of specific methionine residues was directly related to their surface accessibility. The pattern of oxidative modification of the methionines in the aged CaM was less straightforward, though both in vitro oxidation of CaM and aging within the brain markedly decreased the functional properties of this important Ca(2+)-regulating protein.

Rate constants for the reactions of halogenated organic radicals.

Absolute rate constants have been measured by means of pulse radiolysis for the reactions of various halogenated aliphatic compounds (ethane derivatives, including the anaesthetics halothane, enflurane, isoflurane and methoxyflurane) with hydrated electrons and .OH radicals, the reactions of halogenated carbon-centered radicals, derived thereby, with molecular oxygen, and the reactions of halogenated peroxyl radicals with various antioxidants (ascorbate, chlorpromazine, promethazine, propyl gallate, ABTS) in aqueous solutions. All oxygen addition reactions occur essentially diffusion-controlled. This finding is correlated with the stereoelectronic properties of the primary carbon-centred radicals. The oxidative power of the halogenated peroxyl radicals reflects the inductive -I effect of the halogens and accordingly increases with the degree of halogen substitution, with fluorine substituents being particularly effective. The peroxyl radicals derived from freon 113, namely CClF2CClFOO. and CCl2FCF2OO., have been identified as the best oxidants among these species.

Autocatalytic nitration of P450CAM by peroxynitrite.

Peroxynitrite (PN) gains high selectivity as a physiological oxidizing and nitrating agent through catalysis by metal ions. This was established for the heme-thiolate (P450) enzyme prostacyclin synthase which was tyrosine nitrated and inhibited at low PN levels [FEBS Lett. 382 (1996) 101]. Other P450 proteins reacted in a similar manner and a ferryl species (Compound II) has been identified as an intermediate during reactions with PN [Nitric Oxide 3 (1999) 142]. Here we investigated cytochrome P450CAM and found that it catalyzes the decomposition of PN as well as an increased nitration of phenol. The latter at the expense of phenol hydroxylation is characteristic for the proton-assisted PN action. PN also caused self-nitration of P450CAM at several tyrosine residues. Two of them, Y96 and Y305 were largely protected in the presence of the ligand metyrapone. Unlike other heme-thiolate proteins P450CAM did not form distinct spectral intermediates characteristic for Compound II. We conclude that P450CAM serves as a model for the nitration of prostacyclin synthase with respect to its autocatalytic tyrosine nitration and its prevention by blocking the active site.

Chemical stability of nucleic acid-derived drugs.

Nucleic acid-derived drugs exhibit both chemical and physical instability. This mini-review focuses on the prevalent hydrolytic and oxidative pathways of chemical degradation as they are affected by various endogenous (primary structure, chemical modifications in bases, sugars and phosphate residues) and exogenous (pH, buffer concentration, metal cation presence, oxygen presence) factors.

Oxidative degradation of pharmaceuticals: theory, mechanisms and inhibition.

Oxidation is a common pathway for drug degradation in liquid and solid formulations. The present article reviews mechanistic details of autoxidation and chain oxidation processes, as well as the oxidation of selected functional groups commonly affected in drugs.

Metal-catalyzed oxidation of human growth hormone: modulation by solvent-induced changes of protein conformation.

Metal-catalyzed oxidation (MCO) represents a prominent pathway of protein degradation. To evaluate the importance of the integrity of the metal-binding site on MCO, we subjected recombinant human growth hormone (rhGH), to MCO (ascorbate, Cu(2+), (3)O(2)) in the presence of various aliphatic alcohols (ethanol, ethylene glycol, trifluoroethanol, 1-propanol, 2-propanol, 1,2-propylene glycol, 1-butanol, 2-butanol, and tert-butanol). All alcohols inhibited MCO in a concentration-dependent and sigmoidal manner. Half-points, P(1/2), were dependent on the nature of the alcohol. Circular dichroism and fluorescence spectroscopy were used to monitor cosolvent-induced secondary and tertiary structural changes. The presence of alcohols increased the helical content of rhGH and induced a red shift in the tryptophan emission. The midpoints of the tertiary structural change correlated with the P(1/2) values. Solvent polarity at P(1/2) was determined according to the E(T)(30) scale. All alcohol/water mixtures at P(1/2) had rather similar solvent polarities between 54.5 to 56.4 kcal/mol, with the exception of ethylene glycol. On the other hand, no correlation was obtained between the protection against MCO and the hydroxyl radical-scavenging properties of the cosolvent. We conclude that the primary mechanism of MCO inhibition is a cosolvent-induced conformational perturbation of the metal-binding site as opposed to pure radical scavenging.

Photostability of 2-hydroxymethyl-4,8-dibenzo[1,2-b:5,4-b']dithiophene-4,8-dione (NSC 656240), a potential anticancer drug.

Stability studies of 2-hydroxymethyl-4,8-dibenzo[1,2-B:5,4-b']dithiophene-4,8-dione (NSC 656240, dithiophene), a poorly water-soluble (approximately 5 microg/ml) potential anticancer drug are reported. Dithiophene stability turned out to be very sensitive to laboratory fluorescent lighting. The rate of photodegradation of dithiophene was studied in aqueous solutions at room temperature (approximately 25 degrees C) at various pH values, in MeOH, CH(3)CN, DMF, DMA, and in mixed nonbuffered aqueous/organic solutions. The aqueous pH-rate profile indicated no sensitivity to changing pH values. 1H NMR and LC/MS methods were used to characterize the degradation products. Dithiophene photodegradation in the presence of air followed an apparent autoxidation pathway with dithiophene-2-aldehyde and dithiophene-2-carboxylic acid as the major degradants. The structures were confirmed against authentic samples. Dithiophene photodegradation under anaerobic conditions followed an apparent disproportionation pathway with only one identified major product, dithiophene-2-aldehyde.