Ameliorative effect of Acetyl L-carnitine in Alzheimer's Disease via Downregulating of Homocysteine Levels in Hyperhomocysteinemia Induced Cognitive Deficit in mouse model

Aim: The study was aimed at exploring the role of Acetyl L-Carnitine supplementation attenuating dementia and degradation of cognitive abilities in Hyperhomocysteinemia induced AD manifestations in the mouse model. Background: Alzheimer’s disease (AD) is a neurological disorder that is marked by dementia, and degradation of cognitive abilities. There is great popularity gained by natural supplements as the treatment for AD, due to the higher toxicities of synthetic drugs. Hyperhomocysteinemia causes excitotoxicity to the cortical neurons, which brought us to the point that amino acids possibly have a role in causing cholinergic deformities, which are an important etiological parameter in AD. Acetyl L-Carnitine a methyl donor with the presence of three chemically reactive methyl groups linked to a nitrogen atom was found to possess neuroprotective activity against experimental models of AD. Objective: The objective of the present investigation was to investigate and evaluate the pharmacological effect of Acetyl L-Carnitine against hyperhomocysteinemia induced Alzheimer’s disease (AD) in the mouse model. Material and Method: The animals were divided into normal control (vehicle-treated), HHcy (dl-Homocysteine thiolactone treated) negative control, test group i.e., low dose (50mg/kg, p.o) of acetyl L-carnitine (L-ALC), high dose (100mg/kg,p.o) of acetyl L-carnitine (H-ALC), L-ALC+SOV (Sodium orthovanadate) and H-ALC+SOV. HHcy was induced by administration of dl-Homocysteine thiolactone (dl-HCT; 1 g/kg, p.o.) on day-1 to day-15 of experimental schedule to all animals except normal control. The changes in the behaviour pattern of the animals due to neuroinflammation, and cholinergic dysfunction were examined in rotarod, novel objective recognition, passive avoidance, elevated plus maze, and morris water maze analysis. Biochemical investigation includes the estimation of total homocysteine (tHcy), Creatinine Kinase (CK), Acetylcholinesterase (AChE), thiobarbituric acid reactive substances (TBARS), reduced glutathione (GSH) and IL-6 and TNF-α. Result: Supplementation of ALC in mouse considerably lowered the HHcy-induced AD manifestations in the experimental animals. It was found that ALC and SOV successfully diminished the behaviour abnormalities and lessened the Hcy-induced alteration in systemic Hcy levels, CK activity, and cholinergic dysfunction with improved bioenergetics in the Prefrontal cortex of the mice. Conclusion: ALC was found to improve the HHcy-induced cognitive disabilities which was found to be associated with the decreased systemic levels of Hcy, CK, and cholinergic abnormalities. It also combats the oxidative stress-induced neuroinflammation with diminished pro-inflammatory markers in the pre frontal cortex. The outcomes collectively indicate ALC's potential to be used as a supplementation in the pharmacotherapy of AD.

Chemical Modification of Nisin With Homocysteine Thiolactone And Its Antibacterial Activity

The full text of this preprint has been withdrawn by the authors due to author disagreement with the posting of the preprint. Therefore, the authors do not wish this work to be cited as a reference. Questions should be directed to the corresponding author.

The Use of Single Drop Microextraction and Field Amplified Sample Injection for CZE Determination of Homocysteine Thiolactone in Urine

Two cheap, simple and reproducible methods for the electrophoretic determination of homocysteine thiolactone (HTL) in human urine have been developed and validated. The first method utilizes off-line single drop microextraction (SDME), whereas the second one uses off-line SDME in combination with field amplified sample injection (FASI). The off-line SDME protocol consists of the following steps: urine dilution with 0.2 mol/L, pH 8.2 phosphate buffer (1:2, v/v), chloroform addition, drop formation and extraction of HTL. The pre-concentration of HTL inside a separation capillary was performed by FASI. For sample separation, the 0.1 mol/L pH 4.75 phosphate buffer served as the background electrolyte, and HTL was detected at 240 nm. A standard fused-silica capillary (effective length 55.5 cm, 75 μm id) and a separation voltage of 21 kV (~99 μA) were used. Electrophoretic separation was completed within 7 min, whereas the LOD and LOQ for HTL were 0.04 and 0.1 μmol/L urine, respectively. The calibration curve in urine was linear in the range of 0.1–0.5 μmol/L, with R2 = 0.9991. The relative standard deviation of the points of the calibration curve varied from 2.4% to 14.9%. The intra- and inter-day precision and recovery were 6.4–10.2% (average 6.0% and 6.7%) and 94.9–102.7% (average 99.7% and 99.5%), respectively. The analytical procedure was successfully applied to the analysis of spiked urine samples obtained from apparently healthy volunteers.

Preparation of Nanofibers Mats Derived from Task-Specific Polymeric Ionic Liquid for Sensing and Catalytic Applications

Nanofibers mats derived from the task-specific functionalized polymeric ionic liquids based on homocysteine thiolactone are obtained by electrospinning them as blends with polyvinylpyrrolidone. The presence of this functional moiety allowed the post-functionalization of these mats through the aminolysis of the thiolactone ring in the presence of an amine by a thiol–alkene “click” reaction. Under controlled experimental conditions the modification can be performed introducing different functionalization and crosslinking of the electrospun fibers, while maintaining the nanostructure obtained by the electrospinning. Initial studies suggest that the nanofibers based on these functionalized polymeric ionic liquids can be used in both sensing and catalytic applications.

Protein-peptide composition in the lungs of rats with hyperhomocysteinemia

The accumulated data indicate that a high level of homocysteine may be a central pathogenetic factor of chronic obstructive pulmonary disease. In this study, we investigated the effect of hyperhomocysteinemia on protein homeostasis in the rat lungs. The level of proteins, peptides, total proteolytic activity, as well as protein-peptide composition, were evaluated. Hyperhomocysteinemia was induced by daily intragastric administration of DL-homocysteine thiolactone (100 mg·kg-1 of body weight) to albino non-linear male rats for 28 days. Twelve hours after the last administration, the rats were sacrificed and the lungs were harvested. Our findings showed that hyperhomocysteinemia caused the disturbances in the protein homeostasis in the lungs that are manifested by a decrease in the level of proteins in the young and old animals and an increase in the level of peptides in the rats of all studied groups. We found a change in the protein composition in the lung of HM rats - a decrease in the level of proteins with a molecular weight of 50 kDa to 100 kDa simultaneously with an increase in the level of proteins with a molecular weight of less than 50 kDa. Despite the fact that the peptide profile was the same in both control animals and HM animals, the level of individual peptide fractions increased significantly in the rats with HM. Obtained data could contribute to explain, at least in part, the mechanisms involved in the pathogenesis of lung damage in hyperhomocysteinemia.

Chemical Modification of Nisin With Homocysteine Thiolactone And Its Antibacterial Activity

The objective of this study was to prepare N-homocysteine conjugate of nisin, and to evaluate the effect of N-homocysteinylation reaction on the antibacterial activity of nisin. Progression of the modification was monitored by measuring free sulfhydryl groups, and by using both acetic acid urea polyacrylamide gel electrophoresis (AAU-PAGE) and tricine sodium dodecyl sulphate polyacrylamide gel electrophoresis (tricine SDS-PAGE). Optimum conditions to attain the maximum N-homocysteinylation degree (6.30%) were determined as 6 mg/mL nisin, 150 mM homocysteine thiolactone, 150 rpm shaker speed, 3.0 pH and 6 hrs reaction time. Antibacterial activity of N-homocysteinylated nisin was determined against Staphylococcus aureus ATCC 6538, Bacillus subtilis ATCC 6633, Lactococcus lactis ssp. cremoris AÜ, Enterococcus faecium ATCC 9097, Listeria monocytogenes NCTC 5348 and Escherichia coli RSKK. Of all the bacteria tested only the growth of E. faecium was inhibited slightly by modified nisin having just about 10% of antibacterial activity of unmodified nisin. Tricine SDS-PAGE analysis applied after trypsin and thermolysin hydrolysis showed that side chains of Lys12 and Lys22 were both modified with homocysteine thiolactone reaction.

Homocysteine Thiolactone: Biology and Chemistry

Homocysteine thiolactone is a five-membered cyclic thioester of amino acid homocysteine. It is generated from homocysteine as a result of an error-editing reaction, principally, of methionyl-tRNA synthetase. An elevated level of homocysteine thiolactone is associated with cardiovascular diseases, strokes, atherosclerosis, neurological abnormalities, etc., presumably because it reacts to the side chain of protein lysine causing protein damage and autoimmune responses. It is not only an important metabolite but also a versatile building block for organic and bioorganic synthesis. This entry contains data on the homocysteine thiolactone formation, metabolism, toxicity mechanism in vivo, and the bioorganic chemistry applications as a powerful synthetic tool in polymer science, sustainable materials development, and probes.