Peptides

When taken as a supplement, studies show that certain peptides could help improve body composition, reduce exercise-induced muscle damage, and improve joint health and mobility in athletes and nonathletes.For example, a number of studies have found that collagen peptide supplements are effective for increasing muscle strength, reducing joint pain, and improving joint mobility in athletes and sedentary people.

Most collagen supplements are hydrolyzed — or broken down — into peptides for easier absorption. Collagen peptides may also help improve endurance performance in athletes by stimulating the uptake of glucose (sugar) and the storage of glycogen — the storage form of glucose — in muscle cells.What’s more, collagen peptide supplementation may help increase muscle mass in non-athletic people with low muscle mass like older men and women. Certain peptides, like the collagen-derived peptide dipeptide hydroxyproline-glycine, activate pathways in the body that stimulate the growth of muscle fibers.

Other peptides used to increase muscle mass include growth hormone (GH) releasing peptides. These peptides stimulate the production and release of human growth hormone (HGH), a hormone that can induce muscle growth, which is why these supplements are popular with bodybuilders. 

Lower levels of purity
Generally, the lowest purity preparation is between >50% and >70%, with an average of about >60% purity. Products at this level are referred to as “crude” or “desalted” peptides. Though unflattering by name, this purity level can serve a useful purpose while saving you money, such as in initial peptide screens. Desalted peptides are appropriate for high-throughput screenings of large numbers of peptides for the generation of leads in pharmaceutical development, for example, as well as in other applications.

A >70% pure peptide preparation, which may contain a mixture of peptides that closely resemble one another, is likely to generate an immune response that will result in the antibody that you need. However, it is also possible for impurities at this level to cause unwanted effects in the animal or even some degree of toxicity. This can be caused by organic impurities remaining in the peptide preparation after the process of peptide synthesis. If this is a problem, it might be possible to extract the offending impurities from the >70% peptide preparation instead of buying a higher purity (and possibly more expensive) preparation of the peptide.

Mid-range levels of purity
Many analytical applications use mid-range peptide purities. Generally, a >70% to >80% purity preparation is appropriate for the production, purification and testing of antibodies. These products are also useful for ELISAs, enzyme substrate studies, epitope mapping, affinity purification, bioassays or other immunological applications, and peptide screening as described above.

Peptides purified to the >85% level are appropriate for biochemistry and semi-quantitative applications, such as enzymology, tests for biological activity and epitope mapping. Other examples of applications at this level include semi-quantitative studies of enzyme-substrate interactions, phosphorylation, peptide blocking in Western blotting and cell attachment.

Higher levels of purity
The higher levels of >90% to >98% purity are designed for particular applications. This level is the most expensive, and it usually isn’t necessary for the applications described above. Peptides purified to the >90% to >95% level are best for quantitative bioassays, quantitative in vitro receptor-ligand interaction studies, biological activity with ligand binding studies, quantitative blocking and competitive inhibition assays, quantitative phosphorylation and proteolysis studies, electrophoresis markers and chromatography standards.

Usually, the highest level of >98% purity is reserved for in vivo studies, clinical trials, drug studies that use peptides as pharmaceuticals and structure-activity relationship studies. This level is also best for examining protein structure by nuclear magnetic resonance studies or protein crystallography (though these latter three applications also can be accomplished using the >95% purity preparation).

A note on packaging
Some peptide preparations may require specialized packaging to make sure they remain intact and functional. Almost invariably, the product will be delivered to you as a lyophilized powder sealed in a polypropylene or glass vial with a screw cap, sometimes packed with an inert gas to prevent oxygen or moisture from entering. Although it is fine to ship purified peptides at atmospheric temperature for relatively short durations, it is best to store the lyophilized powder at -20°C after it arrives.

Approach 1
In general, attempt to dissolve peptides in sterile distilled water or sterile dilute acetic acid (0.1%) solution to give a stock solution at a higher concentration than that required for subsequent use. This solution may later be diluted with an appropriate buffer. If the peptide persists as visible particles, sonication may prove of help as it improves the rate of dissolution. If, after sonication, the’ solution’ has gelled, has a persistent haziness, or has a scum floating on the surface, the peptide has probably not dissolved but is simply finely suspended.

Approach 2
If the peptide remains Insoluble, look at Its amino acid composition prior to proceeding further. What proportion of amino acids are hydrophobic (A, C, F, I, L, M, P, V, W, Y) and how many residues are positively charged (K, R, H, and amino terminus) or negatively charged (D, E, and carboxy terminus)? What is the overall net charge at neutral pH? If there is a net charge at neutral pH, addition of dilute acetic acid (for basic, positively charged peptides) or dilute aqueous ammonia or ammonium bicarbonate (for acidic, negatively charged peptides) with further sonication should greatly aid solubility. The final concentration of acetic acid or ammonia/ammonium bicarbonate allowable will be determined by the use to which the peptide is to be finally put. If the peptide still refuses to dissolve, these volatile buffer systems may be removed by lyophilisation and alternative solvents tried on the same peptide sample.

Approach 3
If the peptide sequence has little or no net charge at any pH, or if the number of hydrophobic residues approaches 50% or more, +he chances are +ha+ +he above procedures may prove inadequate. Addition of acetonitrile, ethanol, dimethylformamide (DMF) dimethyl sulphoxide (DMSO), or the use of chaotropic salts such as guanidine hydrochloride or urea should aid the dissolution of most peptides. The choice will depend largely on the compatibility of such reagents with the subsequent peptide application. If it is known that the peptide is slightly soluble in aqueous solution, it is better to first dissolve it completely in a small amount of neat acetic acid or DMF and then slowly dilute with water or buffer rather than to add such solvent progressively to a suspension of the peptide in aqueous systems