The possibility of interaction of 0.1 mg/mL acetylsalicylic acid (ASA) with purified human and rat globin proteins for 24 h in vitro was investigated. It was shown that following the interaction with ASA rat globin is modified at Lys17, Lys57, Lys91, and Lys140 amino acid residues of the α-subunit as well as at Lys18 and Lys77 of the β-subunit, whereas human globin is acetylated at Lys17, Lys41, Lys57, and Lys91 residues of the α-subunit as well as Lys18, Lys96, and Lys133 of the β-subunit of the protein. Incubation of human whole blood with 0.1 mg/mL ASA for 3 h followed by globin isolation led to the identification of acetylated Lys17 and Lys57 lysine residues of the α-subunit of human globin.

A reaction of high-temperature solid-phase catalytic isotope exchange (HSCIE) was studied for the preparation of tritium- and deuterium-labeled ligands of glutamate and dopamine receptors. Tritium-labeled (5S,10R)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclopenten-5,1-imine ([G-3H]MK-801) and R(+)-7-hydroxy-N,N-di-n-propyl-2-aminotetraline ([G-3H]-7-OH-DPAT) were obtained with a specific activity of 210 and 120 Ci/mol, respectively. The isotopomeric distribution of deuterium-labeled ligands was studied using time-of-flight mass-spectrometer MX 5310 (ESI-o-TOF) with electrospray and orthogonal ion injection. Mean deuterium incorporation per ligand molecule was 11.09 and 3.21 atoms for [G-3H]MK-801 and [G-3H]-7-OH-DPAT, respectively. The isotope label was shown to be distributed all over the ligand molecule. The radioreceptor binding of tritium-labeled ligands [G-3H]MK-801 and [G-3H]-7-OH-DPAT was analyzed using the brain structure of Vistar rats. It was demonstrated that [G-3H]MK-801 specifically binds to hippocampus membranes with K d 8.3 ± 1.4 nM, B max being 3345 ± 300 fmol/mg protein. The [G-3H]-7-OH-DPAT ligand specifically binds to rat striatum membranes with K d 10.01 ± 0.91 nM and B max 125 ± 4.5 fmol/mg protein. It was concluded that the HSCIE reaction can be used for the preparation of highly tritium-labeled (+)-MK-801 and 7-OH-DPAT with retention of their physiological activities.

Для изучения биологических свойств ациламинокислот синтезированы N-арахидоноильные (АА) производные аминокислот: Gly, Phe, Pro, Val, у-аминомасляной кислоты (GABA), диоксифенилала-нина, Туг, Тгр и Ala, а также пептидов MEHFPGP и PGP. В масс-спектрах ионизации при атмосферном давлении распылением в электрическом поле все соединения дают протежированный молекулярный ион максимальной интенсивности, предел детектирования составлял 10 фмоль вещества на образец. Из всех изученных ациламинокислот наибольшую ингибиторную активность по отношению к гидролазе амидов жирных кислот из мозга крысы показал AA-Gly (IC5o 6.5 мкМ). Слабое, но регистрируемое ингибирование проявляли AA-Phe, АА-Туг и AA-GABA (1С5о 55, 60 и 50 мкМ соответственно). Сами ацилированные аминокислоты, за исключением AA-Gly, были устойчивы к гидролизу этим ферментом. Все арахидоноиламинокислоты в той или иной степени ингибировали фос-фолипазу D из капусты, наиболее активными оказались AA-GABA и AA-Phe (IC5o 20 и 27 мкМ соответственно). Опыты по выяснению возможности биосинтеза АА-Туг в гомогенатах печени и нервных тканях крысы не показали образования этой ациламинокислоты in vitro, однако были обнаружены продукты ее метаболизма: АА-дофамин и АА-(З-О-метилдофамин); наибольшее количество первого метаболита наблюдали в гомогенате печени, а второго - в гомогенате головного мозга. Ациламинокислоты не оказывали цитотоксического действия на клетки глиомы С6. Показано, что N-ацилирование гептапептида MEHFPGP (семакс) арахидоновой кислотой приводит к повышению его гидролитической стабильности и увеличению сродства к местам специфического связывания на мембранах мозжечка крысы.

N-Arachidonoyl (AA) derivatives of amino acids (glycine, phenylalanine, proline, valine, γ-aminobutyric acid (GABA), dihydroxyphenylalanine, tyrosine, tryptophan, and alanine) and peptides (Semax, MEHFPGP, and PGP) were synthesized in order to study the biological properties of acylamino acids. The mass spectra of all the compounds at atmospheric pressure electrospray ionization display the most intense peaks of protonated molecular ions; the detection limits for these compounds are 10 fmol per sample. AA-Gly showed the highest inhibitory activity toward fatty acid amide hydrolase from rat brain (IC50 6.5 μM) among all the acylamino acids studied. AA-Phe, AA-Tyr, and AA-GABA exhibited a weak but detectable inhibitory effect (IC50 55, 60, and 50 μM, respectively). The acylated amino acids themselves, except for AA-Glu, were stable to the hydrolysis by this enzyme. All the arachidonoylamino acids inhibited cabbage phospholipase D to various degrees; AA-GABA and AA-Phe proved to be the most active (IC50 20 and 27 μM, respectively). Attempts to detect the biosynthesis of AA-Tyr in homogenates of rat liver and nerve tissue in vitro were unsuccessful; however, AA-dopamine and AA-Phe, the products of its metabolism, were found. The highest contents of these metabolites were detected in liver homogenate and in the brain homogenate, respectively. Acylamino acids exert no cytotoxic effect toward the glioma C6 cells. It was shown that N-acylation of Semax with arachidonic acid results in enhancement of its hydrolytic stability and increases its affinity for the sites of specific binding in rat cerebellum membranes.