Peptides

Contents 3.1 Introduction ..................................................................................................................... 42 3.2 Sample Preparation and Extraction .................................................................................. 42 3.3 Fractionation .................................................................................................................... 43

3.3.1 Ultrafi ltration ....................................................................................................... 43 3.3.2 Gel Filtration Chromatography ............................................................................ 43

3.4 Analysis ............................................................................................................................ 44 3.4.1 Analysis of Dipeptides .......................................................................................... 44 3.4.2 Analysis of Glutathione .........................................................................................45 3.4.3 Analysis of Other Peptides .................................................................................... 46

3.4.3.1 Reversed-Phase HPLC ........................................................................... 46 3.4.3.2 Ion-Exchange Chromatography ..............................................................47 3.4.3.3 Capillary Electrophoresis ........................................................................47 3.4.3.4 Gel Electrophoresis ................................................................................ 48

3.5 Peptide Sequencing .......................................................................................................... 48 3.5.1 Peptides from Proteins .......................................................................................... 49

3.5.1.1 Polypeptide Digestion ............................................................................ 49 3.5.1.2 Sequencing of Tryptic Peptides .............................................................. 50

3.5.2 Free Peptides .........................................................................................................51 References ..................................................................................................................................52

3.1 Introduction Th ere are several peptides naturally present in muscle. Carnosine (β-alanyl--histidine), anserine (β-alanyl--1-methylhistidine), and balenine (β-alanyl--3-methylhistidine) are dipeptides that express some physiological properties in muscle.1,2 Th e content of these dipeptides is especially high in muscles with glycolytic metabolism, though this also depends on the animal species, age, and diet.3-5 Beef and pork contain more carnosine and less anserine, lamb has similar amounts of carnosine and anserine, while poultry is very rich in anserine. Balenine is present in minor amounts in pork muscle.6 Glutathione (GSH) is a cysteine-containing tripeptide (glutamine, glycine, and cysteine) naturally present in fresh meats. Th is tripeptide plays an essential role in the antioxidant system, as well as in the intercellular redox state, by the blockage of reactive oxygen species and free radicals. Th is thiol compound exists in two forms, the reduced (GSH) and the oxidized form (GSSG), and the ratio of the two forms is crucial for the characterization of the oxidative stress in cells. Th e interrelation among tissue GSH, nutrition, and oxidative stress is widely discussed elsewhere.7 Most meats have been found to contain approximately twice the GSH found in poultry and 2-10 times more GSH than fi sh products.8