Selective modification of apoB-100 in the oxidation of low density lipoproteins by myeloperoxidase in vitro.
Oxidative modification of LDL may be important in the initiation and/or progression of atherosclerosis, but the precise mechanisms through which low density lipoprotein (LDL) is oxidized are unknown. Recently, evidence for the existence of HOCl-oxidized LDL in human atherosclerotic lesions has been reported, and myeloperoxidase (MPO), which is thought to act through production of HOCl, has been identified in human atherosclerotic lesions. In the present report we describe the formation of 2,4-dinitrophenylhydrazine (DNPH)-reactive modifications in the apolipoprotein (apo) by exposure of LDL to myeloperoxidase in vitro. In contrast with the complex mixture of peptides from oxidation of LDL with reagent HOCl, oxidation with MPO in vitro produced a major tryptic peptide showing absorbance at 365 nm. This peptide was isolated and characterized as VELEVPQL(*C)SFILK..., corresponding to amino acid residues 53-66...on apoB-100. Mass spectrometric analyses of two tryptic peptides from oxidation of LDL by HOCl indicated formation of the corresponding methionine sulfoxide (M=O), cysteinyl azo (*C), RS -N= N-DNP, derivatives of EEL(*C)T(M=O)FIR and LNDLNS VLV(M=O)PTFHVPFTDLQVPS(*C)K, which suggest oxidation to the corresponding sulfinic acids (RSO2H) by HOCl. The present results demonstrate that DNPH-reactive modifications other than aldehydes and ketones can be formed in the oxidation of proteins and illustrate how characterization of specific products of protein oxidation can be useful in assessing the relative contributions of different and unexpected mechanisms to the oxidation of LDL and other target substrates. The data also suggest a direct interaction of the LDL particle with the active site on myeloperoxidase and indicate that effects of the protein microenvironment can greatly influence product formation and stability. (+info)
Antimicrobial activity of the thiosulfinates isolated from oil-macerated garlic extract.
Three thiosulfinates were isolated from oil-macerated garlic extract, and their structures were identified as 2-propene-1-sulfinothioic acid S-(Z,E)-1-propenyl ester [AllS(O)SPn-(Z,E)], 2-propenesulfinothioic acid S-methyl ester [AllS(O)SMe], and methanesulfinothioic acid S-(Z,E)-1-propenyl ester [MeS(O)SPn-(Z,E)]. This is the first report of isolating these thiosulfinates from oil-macerated garlic extract. Antimicrobial activities of AllS(O)SPn-(Z,E) and AllS(O)SMe against Gram-positive and negative bacteria and yeasts were compared with 2-propene-1-sulfinothioic acid S-2-propenyl ester [AllS(O)SAll, allicin] which is well-known as the major thiosulfinate in garlic. Antimicrobial activity of AllS(O)SMe and AllS(O)SPn-(Z,E) were comparable and inferior to that of allicin, respectively. This result suggested that the antimicrobial activity of 2-propene sulfinothioic acid S-alk(en)yl esters were affected by alk(en)yl groups. The order for antimicrobial activity was: allyl > or = methyl > propenyl. (+info)
The mode of action of allicin: its ready permeability through phospholipid membranes may contribute to its biological activity.
Allicin (diallyl thiosulfinate) is the main biologically active component of the freshly crushed garlic extracts. In the present work the ability of allicin to cross through membranes (artificial and biological) was studied. Partition coefficients of allicin in water/octanol, water/hexadecane and water/phospholipids mixtures were determined. Using phospholipid vesicles loaded with hydrophilic thiols (reduced glutathione or 2-nitro-5-thiobenzoate), we observed that allicin freely permeates through phospholipid bilayers and interacts with the SH groups. The reaction rate of allicin with SH containing molecules after crossing the membrane was the same as in solution. Fast diffusion and permeation of allicin across human red blood cell membranes was also demonstrated. Allicin does not induce leakage, fusion or aggregation of membrane. The high permeability of allicin through membranes may greatly enhance the intracellular interaction with thiols. (+info)
Activities of garlic oil, garlic powder, and their diallyl constituents against Helicobacter pylori.
Chronic Helicobacter pylori disease is reduced with Allium vegetable intake. This study was designed to assess the in vivo anti-H. pylori potential of a variety of garlic substances. The garlic materials all showed substantial but widely differing anti-H. pylori effects against all strains and isolates tested. The MICs (range, 8 to 32 microg/ml) and minimum bactericidal concentrations (MBCs) (range, 16 to 32 microg/ml) of undiluted garlic oil (GO) were smaller than those of garlic powder (GP) (MIC range, 250 to 500 microg/ml; MBC range, 250 to 500 microg/ml) but greater than the MIC of allicin (4. 0 microg/ml) (Table 2) present in GP. Allicin (MIC, 6 microg/ml; MBC, 6 microg/ml) was more potent than diallyl disulfide (MIC range, 100 to 200 microg/ml; MBC range, 100 to 200 microg/ml), its corresponding sulfide, but of a strength similar to that of diallyl tetrasulfide (MIC range, 3 to 6 microg/ml; MBC range, 3 to 6 microg/ml). Antimicrobial activity of the diallyl sulfides increased with the number of sulfur atoms. Time course viability studies and microscopy showed dose-dependent anti-H. pylori effects with undiluted GO, GP, allicin, and diallyl trisulfide after a lag phase of ca. 1 to 2 h. Substantial in vitro anti-H. pylori effects of pure GO and GP and their diallyl sulfur components exist, suggesting their potential for in vivo clinical use against H. pylori infections. (+info)
Analysis of responses of garlic derivatives in the pulmonary vascular bed of the rat.
Allicin, an extract from garlic, has been shown to be a systemic and pulmonary arterial vasodilator that acts by an unknown mechanism. In the present experiments, pulmonary vascular responses to allicin (10-100 microg), allyl mercaptan (0.3-1 mg), and diallyl disulfide (0.3-1 mg) were studied in the isolated lung of the rat under constant-flow conditions. When baseline tone in the pulmonary vascular bed of the rat was raised to a high-steady level with the thromboxane A(2) mimic U-46619, dose-related decreases in pulmonary arterial pressure were observed. In terms of the mechanism of action of allicin vasodilator activity in the rat, responses to allicin were not significantly different after administration of the nitric oxide synthase inhibitor N(omega)-nitro-L-arginine methyl ester, the K(ATP)(+) channel antagonist U-37883A, or the cyclooxygenase inhibitor sodium meclofenamate, or when lung ventilation was interrupted. These data show that allicin has significant vasodilator activity in the pulmonary vascular bed of the rat, whereas allyl mercaptan and diallyl disulfide produced no significant changes in pulmonary arterial perfusion pressure. The present data suggest that pulmonary vasodilator responses to allicin are independent of the synthesis of nitric oxide, ATP-sensitive K(+) channels, activation of cyclooxygenase enzyme, or changes in bronchomotor tone in the pulmonary vascular bed of the rat. (+info)
S-Allylmercaptoglutathione: the reaction product of allicin with glutathione possesses SH-modifying and antioxidant properties.
The reaction between allicin (diallylthiosulfinate), the active component of garlic and reduced glutathione was investigated. The product of this reaction, mixed disulfide S-allylmercaptoglutathione (GSSA) was separated by high performance liquid chromatography and identified by 1H and (13)C nuclear magnetic resonance and mass spectroscopy. The reaction is fast (with an apparent bimolecular reaction rate constant of 3.0 M(-1) s(-1)). It is pH-dependent, which reveals a direct correlation to the actual concentration of mercaptide ion (GS(-)). Both GSSA and S-allylmercaptocysteine (prepared from allicin and cysteine) reacted with SH-containing enzymes, papain and alcohol dehydrogenase from Thermoanaerobium brockii yielding the corresponding S-allylmercapto proteins, and caused inactivation of the enzymes. The activity was restored with dithiothreitol or 2-mercaptoethanol. In addition, GSSA also exhibited high antioxidant properties. It showed significant inhibition of the reaction between OH radicals and the spin trap 5,5'-dimethyl-1-pyroline N-oxide in the Fenton system as well as in the UV photolysis of H2O2. In ex vivo experiments done with fetal brain slices under iron-induced oxidative stress, GSSA significantly lowered the production levels of lipid peroxides. The similar activity of GSSA and allicin as SH-modifiers and antioxidants suggests that the thioallyl moiety has a key role in the biological activity of allicin and its derivatives. (+info)
Intake of garlic and its bioactive components.
The health benefits of garlic likely arise from a wide variety of components, possibly working synergistically. The complex chemistry of garlic makes it plausible that variations in processing can yield quite different preparations. Highly unstable thiosulfinates, such as allicin, disappear during processing and are quickly transformed into a variety of organosulfur components. The efficacy and safety of these preparations in preparing dietary supplements based on garlic are also contingent on the processing methods employed. Although there are many garlic supplements commercially available, they fall into one of four categories, i.e., dehydrated garlic powder, garlic oil, garlic oil macerate and aged garlic extract (AGE). Garlic and garlic supplements are consumed in many cultures for their hypolipidemic, antiplatelet and procirculatory effects. In addition to these proclaimed beneficial effects, some garlic preparations also appear to possess hepatoprotective, immune-enhancing, anticancer and chemopreventive activities. Some preparations appear to be antioxidative, whereas others may stimulate oxidation. These additional biological effects attributed to AGE may be due to compounds, such as S-allylcysteine, S-allylmercaptocysteine, N(alpha)-fructosyl arginine and others, formed during the extraction process. Although not all of the active ingredients are known, ample research suggests that several bioavailable components likely contribute to the observed beneficial effects of garlic. (+info)
Determination of allicin, S-allylcysteine and volatile metabolites of garlic in breath, plasma or simulated gastric fluids.
Various components of garlic and aged garlic extract, including allicin, S-allylcysteine (SAC) and volatile metabolites of allicin were determined in breath, plasma and simulated gastric fluids by HPLC, gas chromatography (GC) or HPLC- and GC-mass spectrometry (MS). Data indicate that allicin decomposes in stomach acid to release allyl sulfides, disulfides and other volatiles that are postulated to be metabolized by glutathione and/or S-adenosylmethionine to form allyl methyl sulfide. SAC can be absorbed by the body and can be determined in plasma by HPLC or HPLC-MS using atmospheric pressure chemical ionization (APCI)-MS. (+info)