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The present study deals with the application ofhigh-perform ancc-liquid-chromatographyl(HPLC) method for a quantitative detection of carnosine, anserine, L-histidine and 3-methyl-L-histidine in biological material with o-phthaldialdehyde (OPA) post column derivatisation at the constant temperature of 50°C. For this purpose, some mobile-phases were prepared with scalar acetonitrile concentrations. A complete separation of all molecules, particularly for carnosine and 3-methyl-L-his- tidine, was obtained with a solution of acetonitrile and 6mM hydrochloric acid with 0.48 M sodium chloride (5%:95% v/v). Post column derivatisation reaction at temperature of 50°C permitted to obtain an increase in sensibility of all molecules. This method has been utilised for detection of histidine dipeptides in boar spermatozoa and in sheep milk. Concentrations (mean ± S.E. nmol/109 spermatozoa) of carnosine (0.96 ± 0.14) and anserine (0.83 ± 0.18) in boar spermatozoa were significantly lower than those of L-histidine (52.85 ± 4.86) and 3-methyl-L-histidine (83.07 ± 7.1). Positive correlation was found between carnosine and anserine contents (r=0.740; p<0.01) and between L-histidine and 3-methyl-L-histidine (r=0.657; p<0.01). All histidine dipeptides studied were also present in 40 samples of sheep milk. In a case of samples without unit-forming colonies (UFC) of Staphylococcus coagulase-positive, carnosine concentrations (9.17 ±0.89 nmol/ml) were higher than anserine (0.51 ±0.02 nmol/ml) and both were significantly lower in respect to L-histidine (49.51 ± 6.48 nmol/ml) and 3-metyl-L-histidine (81.21 ±6.82 nmol/ml). A negative correlation was observed between carnosine milk levels (r=-0.773; p<0.01) and UFC/ml of Staphylococcus coagulase-positive. In conclusion this very simple and fast method can be used to detect histidine dipeptides in biological compartments where their concentrations are very low.
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Opis fizyczny
p.159-163,fig.,ref.
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autor
- University of Pisa, 2 Viale delle Piagge, 56124 Pisa, Italy
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Bibliografia
- Abe H (2000) Role of histidine-related compounds as intracellular proton buffering constituents in vertebrate muscle. Biochemistry (Mosc) 65: 757-65.
- Aristoy MC, Soler C, Toldra F (2004) A simple, fast and reliable methodology for the analysis of histidine dipeptides as markers of presence of animal origin proteins in feed for ruminants. Food Chemistry 84: 485-491.
- Boldyrev A, Buligina E, Leinsoo T, Petrushanko I, Tsubone S, Abe H (2004) Protection of neuronal cells against reactive oxygen species by carnosine and related compounds. Comp Biochem Physiol B Biochem Mol Biol 137: 81-88.
- Decker EA, Ivanov V, Zhu BZ, Frei B (2001) Inhibition of low-density lipoprotein oxidation by carnosine and histidine. J Agric Food Chem 49: 511-516.
- Decker EA, Livisay SA, Zhou S (2000) A revaluation of activity of purified carnosine. Biochemistry (Mosc) 65: 766-70.
- Ducci M, Pacchini S, Niccolini A, Gazzano A, Innocenti E, Bobowiec R, Martelli F (2004a) Determinazione delle concentrazioni di dipeptidi imidazolici, 1-histidina e 3-me- til 1-histidina nel latte e sangue di capra. VI Congresso Nazionale S.I.Di.L.V., Abano Terme (PD) 1: 147-148.
- Ducci M, Pacchini S, Niccolini A, Martelli F, Buoncristiani P, Sighieri C, Gazzano (2004b) Confronto tra due metodiche HPLC per la determinazione quantitativa di carnosina, anserina. VI Congresso Nazionale S.I.Di.L.V., Abano Terme (PD) 1: 149-150.
- Gadea J, Garcia-Vazquez F, Matas C, Gardon JC, Canovas S, Gumbao D (2005) cooling and freezing of boar spermatozoa: supplementation of the freezing media with reduced glutathione preserves sperm function. J Androl 26: 396-404.
- Kohen R Yamamoto Y Cundy K Ames BN. (1988) Antioxidant activity of carnosine, homocarnosine, and anserine present in muscle and brain. Proc Natl Acad Sei (USA) 85: 3157-3179.
- Penafiel R, Ruzafa C, Monserrat F, Cremades A (2004) Gender-related differences in carnosine, anserine and lysine content of murine skeletal muscle. Amino Acids 26: 53-58.
- Quinn PJ, Boldyrev AA Formazuyk VE (1992) Carnosine: its properties, functions and potential therapeutic applications. Mol Aspects Med 13: 379-444.
- Sanocka D, Kurpisz M (2004) Reactive oxygen species and sperm cells. Reprod Biol Endocrinol 2: 12.
- Schonherr J (2002) Analysis of products of animal origin in feed by determination of carnosine and related dipeptides by high-performance liquid chromatography. J Agric Food Chem 50: 1945-1950.
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Bibliografia
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