Immune system bioactive peptides

The following protocols can be used for the isolation and structural characterization of any natural bioactive peptides from the immune system of invertebrates. The different procedures that will be detailed below refer to the identification and primary structure determination of the Drosophila immune-induced peptides (19,20,23,27,30) and of bioactive peptides from the immune system of other Diptera (17,21,24,31). These approaches were also successfully used for the discovery of bioactive peptides from crustaceans, arachnids, and mollusks. These methods should be considered as a guideline and not as the exact procedure to follow (see Note 3). The suggested procedures will be reported following the normal order of execution, (1) induction of the immune response by an experimental infection, (2) collection of the immunocompetent cells (hemocytes), tissues (epithelia, trachea, salivary glands, etc.)... 

Because of their well-recognized physicochemical properties (relatively high hydrophobicity, cationic character, and short length) reversed-phase, size-exclusion, and cation exchange chromatographies are particularly appropriate to purify bioactive peptides from the immune system of invertebrates. The sensitivity of HPLC, MS, Edman degradation, and liquid growth inhibition assays allow one to use from narrow (e.g., 2.1-mm internal diameter) down to micro-or nano-columns. 

A relevant set of not experimentally infected individuals (control) is required for differential analysis by MALDI-TOF-MS or RP-HPLC in order to discriminate between immune-induced molecules and constitutively present substances (19). This is a prerequisite when no in vitro assays are used to select the bioactive peptides from the immune system of the model invertebrate investigated. 

The relative low cost of synthesis and the more restricted side effects of small peptides, as compared to other classes of bioactive molecules, including proteins, support their medicinal use. However, peptides derived from natural amino acids are proteases substrates and might be eliminated by the immune system as non-self molecules, limiting their use in medicinal chemistry. 

Novel foods include those providing protein (soy and dairy), and those that are intended to improve cardiovascular health (stanol esters, P-glucan, bioactive peptides, and n-3 fatty acids), bone and joint health (calcium, glucosamine, and chondroitin), eye health (lutein), the immune system (milkmicronutrients, lactoferrin, lactoperoxidase, and colostrum), gut health (probiotics and prebiotics), body fitness (conjugated linoleic acid, amino acids, and glyco-macropeptide), energy level, and for beauty. 

Bioactive peptides have been defined as specific protein fragments that have a positive impact on body functions or conditions and may influence health. Upon oral administration, bioactive peptides, may affect the major body systems—namely, the cardiovascular, digestive, immune, and nervous systems. The beneficial health effects may be classified as antimicrobial, antioxidative, antithrombotic, antihypertensive, antimicrobial, and immunomodulatory [87,88]. 

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