Determination of Glyceraldehyde Formed in Glucose Degradation and Glycation

Glyceraldehyde (GLA) was determined in glucose degradation and glycation. GLA was detected as a decahydroacridine-1,8-dione derivative on reversed phase HPLC using cyclohexane-1,3-dione derivatizing reagent. The glucose-derived GLA level was higher than the glycation-derived GLA level, because GLA was converted to intermediates and advanced glycation end products (AGE) in glycation. GLA was also generated from 3-deoxyglucosone and glucosone as intermediates of glucose degradation and glycation. This study suggests that glyceraldehyde is generated by hyperglycemia in diabetes, and that it is also formed in medicines such as peritoneal dialysis solution.     

Identification of a novel advanced glycation end product derived from lactaldehyde

Advanced glycation end products (AGEs) are implicated in the development of diabetic complications via the receptor for AGEs (RAGE). We have reported that the 3-hydroxypyridinium (3HP)-containing AGEs derived from α-hydroxyaldehydes physically interact with RAGE and show cytotoxicity. Lactaldehyde (LA) is formed from a reaction between threonine and myeloperoxidase, but no LA-derived AGEs have been characterized. Here, we identify the structure and physiological effects of an AGE derived from LA. We isolated a novel 3HP derivative, 2-acetamido-6-(3-hydroxy-5-methyl-pyridin-1-ium-1-yl)hexanoate, named as N-acetyl-LAPL (lactaldehyde-derived pyridinium-type lysine adduct), from a mixture of LA with N^α-acetyl-L-lysine. LAPL was also detected in the LA-modified protein. LAPL elicited toxicity in PC12 neuronal cells, but the effect was suppressed by the soluble form of RAGE as a decoy receptor. Moreover, surface plasmon resonance-based analysis revealed that LAPL specifically binds to recombinant RAGE. These results indicate that LA generates an AGE containing the 3HP moiety and contributes to RAGE-dependent cytotoxicity.

Abbreviations: AGEs: advanced glycation end products; RAGE: receptor for advanced glycation end products; 3HP: 3-hydroxypyridinium; LA: lactaldehyde; LAPL: lactaldehyde-derived pyridinium-type lysine adduct; BSA: bovine serum albumin; GLAP: glyceraldehyde-derived pyridinium; MPO: myeloperoxidase; HFBA: heptafluorobutyric acid; TFA: trifluoroacetic acid; HPLC: high performance liquid chromatography; LC-ESI-QTOF-MS: liquid chromatography-electrospray ionization-quadrupole time-of-flight-mass spectrometry; NMR: nuclear magnetic resonance; LA-BSA: lactaldehyde-modified bovine serum albumin; PBS: phosphate buffered saline, GST, glutathione S-transferase; SPR: surface plasmon resonance; OP-lysine: 2-ammonio-6-(3-oxidopyridinium-1-yl)hexanoate; GLO1: glyoxalase 1; MG, methylglyoxal     

Isolation and Identification of the 3-Hydroxy-5-hydroxymethyl-pyridinium Compound as a Novel Advanced Glycation End Product on Glyceraldehyde-related Maillard Reaction

Glyceraldehyde (200 mM) and N^α-acetyllysine (100 mM) were incubated in 0.2 M sodium phosphate buffer (pH 7.4) at 37°C for a week. A major compound, glyceraldehyde-related Maillard reaction product, was purified from the reaction mixture using reverse phase (ODS)-HPLC. It was identified as 1-(5-acetylamino-5-carboxypentyl)-3-hydroxy-5-hydroxymethyl-pyridinium, named as GLAP (Glyceraldehyde derived Pyridinium compound), using NMR and MS analyses. It was suggested that GLAP as a novel advanced glycation end product (AGE) is one of the key compounds in the glyceraldehyde-related Maillard reaction.     

Prevention and reversible solubilization of advanced glycation and products (AGE) by organic germanium compounds as derivatives of amino acids

Summary The amino-carbonyl reaction (The Maillard reaction) of bovine lens crystallin, serum albumin or skin collagen with glucose was investigated to find effective means to prevent the formation of Advanced Glycation End Products (AGE) and induce the reversible solubilization of polymerized glycated proteins. The organic germanium compounds (Ge-132, 373, 385), derivatives of amino acids containing germanium as the linker of framework, were combined by the box titration method to determine the dose that would be most effective, compared with Aminoguanidine-HCl (AMG),-tocopherol (VE), and pirenoxine (Catalin-K, CK). Although AMG suppressed the formation of AGE, effective concentrations were higher than 20 mM. Ge-385, when administered by itself at a low dose, induced the reversible solubilization of AGE made from crystallin, and albumin. The addition of any two reagents such as AMG, VE, CK and Ge-132 or 385 together to proteins lessened the effective range, and the peaks of smaller molecules in the profiles of HPLC and PAGE were quite remarkable. Examination was made of the effects of Ge-132 on the eyes of SAM mice, which show senescence accelerated cataracts at a relatively young age. The prevention of cataract-genesis and induction of reversible transparency of turbid lenses became evident following the administration of Ge-132 to the eyes 4 times a day. The mode of action of organic germanium compounds was demonstrated quite capable of disconnecting the sugar-parts from AGE by decarbonylation, resulting in the formation of glucosone and amino residues, and further leading subsequently to fewer AGE.Abbreviations used in this paper: BLC bovine lens crystallin; BSA bovine serum albumin; AsCol acid soluble bovine skin collagen type III; AGE advanced glycation end products; Ge-132 2-Carboxyethylgermanium sesquioxide, Ge-373 2-Carboxy-2-amino-6-phenyl germanium sesquioxide; Ge-385 2-Carboxy-ethyl-2-aminogermanium sesquioxide; AMG or AG aminoguanidine-HCl; V. E. vitamin E or-tocopherol; CK 1-Hydroxy-5-oxo-5H-pyrido [3, 2-a] phenoxazine-3-carboxylic acid or catalin-K or pirenoxine; PACE polyacrylamide gel electrophoresis; SAM senescence accelerated mouse; HPLC high pressure liquid chromatography; SDS sodium laurylsulfate; FT fructose-p-toluidine.     

Effect of pyridoxamine on chemical modification of proteins by carbonyls in diabetic rats: characterization of a major product from the reaction of pyridoxamine and methylglyoxal

Advanced glycation end products (AGEs) from the Maillard reaction contribute to protein aging and the pathogenesis of age- and diabetes-associated complications. The alpha-dicarbonyl compound methylglyoxal (MG) is an important intermediate in AGE synthesis. Recent studies suggest that pyridoxamine inhibits formation of advanced glycation and lipoxidation products. We wanted to determine if pyridoxamine could inhibit MG-mediated Maillard reactions and thereby prevent AGE formation. When lens proteins were incubated with MG at 37 degrees C, pH 7.4, we found that pyridoxamine inhibits formation of methylglyoxal-derived AGEs concentration dependently. Pyridoxamine reduces MG levels in red blood cells and plasma and blocks formation of methylglyoxal-lysine dimer in plasma proteins from diabetic rats and it prevents pentosidine (an AGE derived from sugars) from forming in plasma proteins. Pyridoxamine also decreases formation of protein carbonyls and thiobarbituric-acid-reactive substances in plasma proteins from diabetic rats. Pyridoxamine treatment did not restore erythrocyte glutathione (which was reduced by almost half) in diabetic animals, but it enhanced erythrocyte glyoxalase I activity. We isolated a major product of the reaction between MG and pyridoxamine and identified it as methylglyoxal-pyridoxamine dimer. Our studies show that pyridoxamine reduces oxidative stress and AGE formation. We suspect that a direct interaction of pyridoxamine with MG partly accounts for AGE inhibition.     

2,3-dihydrojaborosalactone A,a withanolide from Acnistus breviflorus

From Acnistus breviflorus the new 27-hydroxy-5β,6β-epoxy-1-oxo-(22R)-witha-24-enolide (2,3-dihydrojaborosalactone A) as well as seven known withanolides, withaferin A, 2,3-dihydrowithaferin A, 6α-chloro-5β-hydroxywithaferin A, 5,6-deoxywithaferin A, jaborosalactone A, jaborosalactone D and jaborosalactone E were isolated and characterized by means of spectroscopic (¹H NMR, ¹³ C NMR and mass spectral) methods. Depending on the extraction solvent (methanol or ethanol), a known artifact (3β-methoxy-2,3-dihydrowithaferin A) and the new 5α-methoxy-4,5-dihydrojaborosalactone B and 5α-ethoxy-4,5-dihydrojaborosalactone B were also isolated and characterized.     

Two new secondary metabolites,saccharochlorines A and B,from a marine bacterium Saccharomonospora sp. KCTC-19160

Two new chlorinated secondary metabolites, saccharochlorines A and B (1 and 2), were isolated from the saline cultivation of a marine-derived bacterium Saccharomonospora sp. (KCTC-19160). The chemical structures of the saccharochlorines were elucidated by 2D NMR and MS spectroscopic data. Saccharochlorines A and B (1 and 2) exhibit weak inhibition of β-secretase (BACE1) in biochemical inhibitory assay, but they induced the release of Aβ (1–40) and Aβ (1–42) in H4-APP neuroglial cells. This discrepancy might be derived from the differences between the cellular and sub-cellular environments or the epigenetic stimulation of BACE1 expression.     

A Jonah-like chymotrypsin from the therapeutic maggot Lucilia sericata plays a role in wound debridement and coagulation

Lucilia sericata larvae are used in maggot debridement therapy, a traditional wound healing approach that has recently been approved for the treatment of chronic wounds. Maggot excretion products (MEP) contain many different proteases that promote disinfection, debridement and the acceleration of wound healing, e.g. by activating the host contact phase/intrinsic pathway of coagulation. In order to characterise relevant procoagulant proteases, we analysed MEP and identified a chymotrypsin-like serine protease with similarities to Jonah proteases from Drosophila melanogaster and a chymotrypsin from Lucilia cuprina. A recombinant form of the L. sericata Jonah chymotrypsin was produced in Escherichia coli. The activated enzyme (Jonah_m) had a pH optimum of 8.0 and a temperature optimum of 37 °C, based on the cleavage of the chromogenic peptide s-7388 and casein. Jonah_m reduced the clotting time of human plasma even in the absence of the endogenous protease kallikrein, factor XI or factor XII and digested the extracellular matrix proteins fibronectin, laminin and collagen IV, suggesting a potential mechanism of wound debridement. Based on these characteristics, the novel L. sericata chymotrypsin-like serine protease appears to be an ideal candidate for the development of topical drugs for wound healing applications.