Plant sterol oxidation products – Analogs to cholesterol oxidation products from plant origin?

Cholesterol and plant sterols are lipids which are abundantly present in a western type diet of animal and plant origin, respectively. The daily intake averages 300 mg/day each. Over the past decades, a steadily increasing consumption of plant sterol enriched dairy products (2–3 g/day) took place to lower circulating LDL cholesterol concentrations. Like all unsaturated components, plant sterols can be attacked by reactive oxygen species resulting in plant sterol oxidation products (POPs). The most widespread methods for POP determination are high-performance liquid chromatography and gas–liquid chromatography. Yet, based on the low plasma POP concentrations in normophytosterolemic subjects (POPs: ∼0.3–4.5 ng/mL), a reliable quantification yielding an appropriate limit of detection remains a challenge. While the more abundantly present cholesterol oxidation products (COPs) have elaborately been studied, research on the metabolism and biological effects of POPs is only emerging. In relation to atherogenity, biological effects including modulation of cholesterol homeostasis, membrane functioning, and inflammation are attributed to POPs. Although mostly supra-physiological concentrations are applied in in vitro assays, anti-tumor activity, cytotoxicity and estrogen-competition have been attributed to specific POPs. However, it is not obvious, if and how POPs may exert in vivo adverse or beneficial health effects similar to those attributed to COPs. In the field of nutritional science, standardized methods for the determination of POPs are required to perform relevant biological studies and to assess their presence in complex foods or biological tissues and fluids. The aim of this review is to provide an overview and evaluation of the published methods and an update on the biological effects attributed to POPs.     

Some oxidation products of ethoxyquin including those found in autoxidising systems

2,4-Dimethyl-6-ethoxyquinoline (2), 1,2-dihydro-6-ethoxy-2,2,4-trimethylquinoline nitroxide (3), 2,6-dihydro-2,2,4-trimethyl-6-quinone imine N-oxide (4), 2,6-dihydro-2,2,4-trimethyl-6-quinone imine (5), 1,8′-di(1,2-dihydro-6-ethoxy-2,2,4-trimethylquinoline) (6) and 1,2-dihydro-6-hydroxy-2,2,4-trimethylquinoline (7) have been prepared from 1,2-dihydro-6-ethoxy-2,2,4-trimethylquinoline (1) (ethoxyquin) and their spectroscopic properties (UV, IR, mass and NMR) examined.     

Intracellular oxidation of hydroethidine: Compartmentalization and cytotoxicity of oxidation products

Hydroethidine (HE) is a blue fluorescent dye that is intracellularly converted into red-emitting products on two-electron oxidation. One of these products, namely 2-hydroxyethidium, is formed as the result of HE superoxide anion-specific oxidation, and so HE is widely used for the detection of superoxide in cells and tissues. In our experiments we exploited three cell lines of different origin: K562 (human leukemia cells), A431 (human epidermoid carcinoma cells), and SCE2304 (human mesenchymal stem cells derived from endometrium). Using fluorescent microscopy and flow cytometry analysis, we showed that HE intracellular oxidation products accumulate mostly in the cell mitochondria. This accumulation provokes gradual depolarization of mitochondrial membrane, affects oxygen consumption rate in HE-treated cells, and causes cellular apoptosis in the case of high HE concentrations and/or long cell incubations with HE, as well as a high rate of HE oxidation in cells exposed to some stimuli.     

Tyrosine oxidation products: analysis and biological relevance

Summary. Dityrosine is found in several proteins as a product of UV irradiation, -irradiation, aging, exposure to oxygen free radicals, nitrogen dioxide, peroxynitrite, and lipid hydroperoxides. Interest of dityrosine in proteins is based on its potential as a specific marker for oxidatively damaged proteins and their selective proteolysis, hence it could be used as a marker for oxidative stress. Dityrosine is also the product of normal post-translational processes affecting specific structural proteins. Since post-translational modification of a given amino acid in a protein is equivalent to the substitution of that residue by an analogue, it has been proposed that the covalent modification of amino acids may serve as a marking step for protein degradation.     

Some unusual nucleic acid bases are products of hydroxyl radical oxidation of DNA and RNA

There are over 100 modified bases and their derivatives found in RNA and DNA. For some of them, data concerning their properties, synthesis and roles in cellular metabolism are available, but for others the knowledge of their functions and biosynthetic pathways is rather limited. We have analysed the chemical structure of modified nucleosides of DNA and RNA considering mainly their putative synthetic routes. On this basis we suggest, that in addition to enzymatic biosynthetic pathways well established for some odd bases, many rare nucleosides can be recognised as products of random chemical reactions. We identify them as primary or secondary products of the reaction of nucleic acids with hydroxyl radicals, the most active oxidising agent in the cell.     

Bioactive phospholipid oxidation products

Oxidation of phospholipids results in chain-shortened fragments and oxygenated derivatives of polyunsaturated sn-2 fatty acyl residues, generating a myriad of phospholipid products. Certain oxidation products of phosphatidylcholine bind to and activate the human receptor for PAF, and these PAF-like lipids are potent, selective inflammatory mediators. Formation of PAF-like lipids is nonenzymatic and so their accumulation is unregulated. PAF-like lipids are produced in vivo in response to oxidative stresses and are responsible for attendant acute inflammatory responses. PAF-like lipids almost exclusively contain an ether-linked alkyl residue at the sn-1 position of the phosphatidylcholine backbone and molecular identification of these is facilitated by phospholipase A₁ treatment to remove the bulk of the inactive phospholipids. The identity of biologically active species generated by oxidative fragmentation and oxidation can be elucidated by understanding relevant reactions leading to the formation of PAF-like lipids, and then their structure can be established by tandem mass spectrometry and chemical synthesis.     

Phytosterol oxidation products: state of the art

This editorial gives an overview of the current status of the research on the phytosterol oxidation products. The most relevant studies on their in vivo origin, biological effects and analysis in foodstuffs and biological samples are critically reviewed, in order to give concluding remarks on and to establish the future research needs in this field.     

Metabolites and dead-end products from the microbial oxidation of quaternary ammonium alcohols

Methyl-triethanol-ammonium originates from the hydrolysis of the parent esterquat surfactant, which is used as softener in fabric care. The initial steps of the catabolism were investigated in cell-free extracts of the bacterial strain MM 1 able to grow with methyl-triethanol-ammonium as sole source of carbon, energy and nitrogen. The initial degradation of methyl-triethanol-ammonium is an enzymatically catalyzed reaction, located in the particulate fraction of strain MM 1. The oxygen dependent reaction occurred also in presence of phenazine methosulfate as an alternative electron acceptor. As soon as one ethanol group of methyl-triethanol-ammonium was oxidized to the aldehyde, cyclic hemiacetals were formed by intramolecular cyclization. The third ethanol group of methyl-triethanol-ammonium was oxidized to the aldehyde and the carboxylic acid sequentially. The structurally related compounds dimethyl-diethanol-ammonium and choline were oxidized as well, whereas (±)-2,3-dihydroxypropyl-trimethyl-ammonium was not converted at all. The structures of the metabolites were established by 1D and 2D ¹H, ¹³C and ¹⁴N NMR spectroscopy and by capillary electrophoresis mass spectrometry.     

Periodate oxidation products derived from methylated alpha-amanitin: evidence for distinct aldehydic and non-aldehydic forms.

Amatoxins are cyclic octapeptides which can be purified from various mushroom species. They have found widespread use due to their potent inhibition of eukaryotic RNA polymerase II. In the course of our efforts to prepare additional semisynthetic derivatives of the amatoxin, alpha-amanitin, we examined the products formed through the periodate oxidation of 6'-O-methyl-alpha-amanitin. Periodate oxidation under conditions of near-neutral pH yielded two chemically similar, yet chromatographically separable, products. On the basis of their proton NMR spectra and their lack of reactivity with sodium chlorite these products did not contain a free aldehydic group. However, both forms were interconvertible in aqueous neutral solution and could be converted to the same product, 6'-O-methyldemethyl-gamma-amanitin, through reduction with sodium borohydride. Periodate oxidation under mildly acidic conditions generated a single product which has properties of a free aldehyde. It exhibited a proton NMR spectrum with signals characteristic of an aliphatic aldehyde and was readily oxidized to the carboxylic acid with sodium chlorite. Conditions have been defined to synthesize the free aldehyde derivative of 6'-O-methyl-alpha-amanitin in generous yield to provide a precursor for oxidation and further derivatization.     

Secondary products of lipid oxidation

In the last decade, a multitude of secondary products have been identified from the radical and photosensitized oxidations of polyunsaturated lipids. These secondary products consist of oxygenated monomeric materials including epoxy-hydroperoxides, oxo-hydroperoxides, hydroperoxy epidioxides, dihydroperoxides, hydroperoxy bis-epidioxides, and hydroperoxy bicycloendoperoxides. More recently, higher molecular weight dimeric compounds have been identified from autoxidized methyl linoleate and linolenate. Decomposition of these oxidation products form a wide range of carbonyl compounds, hydrocarbons, furans, and other materials that contribute to the flavor deterioration of foods and that are implicated in biological oxidation. The interaction of some of these degradation products with DNA may be involved in cell-damaging reactions.     

Characterization of degradation products from alkaline wet oxidation of wheat straw

Alkaline wet oxidation pre-treatment (water, sodium carbonate, oxygen, high temperature and pressure) of wheat straw was performed as a 2(4-1) fractional factorial design with the process parameters: temperature, reaction time, sodium carbonate and oxygen. Alkaline wet oxidation was an efficient pre-treatment of wheat straw that resulted in solid fractions with high cellulose recovery (96%) and high enzymatic convertibility to glucose (67%). Carbonate and temperature were the most important factors for fractionation of wheat straw by wet oxidation. Optimal conditions were 10 min at 195 degrees C with addition of 12 bar oxygen and 6.5 g l(-1) Na2CO3. At these conditions the hemicellulose fraction from 100 g straw consisted of soluble hemicellulose (16 g), low molecular weight carboxylic acids (11 g), monomeric phenols (0.48 g) and 2-furoic acid (0.01 g). Formic acid and acetic acid constituted the majority of degradation products (8.5 g). The main phenol monomers were 4-hydroxybenzaldehyde, vanillin, syringaldehyde. acetosyringone (4-hydroxy-3,5-dimethoxy-acetophenone), vanillic acid and syringic acid, occurring in 0.04-0.12 g per 100 g straw concentrations. High lignin removal from the solid fraction (62%) did not provide a corresponding increase in the phenol monomer content but was correlated to high carboxylic acid concentrations. The degradation products in the hemicellulose fractions co-varied with the pre-treatment conditions in the principal component analysis according to their chemical structure, e.g. diacids (oxalic and succinic acids), furan aldehydes, phenol aldehydes, phenol ketones and phenol acids. Aromatic aldehyde formation was correlated to severe conditions with high temperatures and low pH. Apart from CO2 and water, carboxylic acids were the main degradation products from hemicellulose and lignin.     

Separation of phosphatidylethanolamine from its oxidation and hydrolysis products by high-performance liquid chromatography

1-Palmitoyl-2-linoleoyl-phosphatidylethanolamine degrades relatively quickly when subjected to common storage and handling procedures. The degradation products consist of compounds in which double bonds in the sn-2 position acyl chain are partially oxidized and of products arising from the hydrolysis of the acyl ester bonds. Thin-layer chromatography (TLC), which is widely utilized to isolate and to ascertain the purity of phospholipids, does not readily separate the oxidation products from the parent lipid class. High-performance liquid chromatography (HPLC), however, employing a normal phase column and an isocratic, UV-transparent solvent system, can be employed to produce a rapid analytical or preparative of phosphatidylethanolamine (PE) from these degradative impurities.     

Copper induced oxidation of serotonin: analysis of products and toxicity

Serotonin is a major neurotransmitter that controls many functions, ranging from mood and behaviour through to sleep and motor functions. The non-enzymatic oxidation of serotonin is of significant importance as some oxidation products are considered to be neurotoxic. An interaction between copper and serotonin has been suggested by symptoms observed in a number of neurodegenerative diseases such as Wilson's and Prion diseases. Using PC12 cells as a model of neuronal cells, we show that the interaction between copper and serotonin is toxic to undifferentiated cells. The toxicity is largely due to reactive oxygen species as cell death is significantly reduced in the presence of the antioxidant mannitol. Differentiation of the PC12 cells also confers resistance to the oxidative process. In vitro oxidation of serotonin by copper results in the eventual formation of a coloured pigment, thought to be a melanin-like polymeric species. Using spectroscopic methods we provide evidence for the formation of a single intermediate product. This dimeric intermediate was identified and characterized as 5,5'-dihydroxy-4,4'-bitryptamine. These results indicate that copper structurally alters serotonin and this process may play a role in copper related neurodegenerative diseases.