Bovine cheese whey was investigated as a source of growth-stimulating factors that might replace or supplement fetal bovine serum in cell culture. Although some cell growth activity was demonstrated in whey or whey ultrafiltrates, enrichment on the basis of molecular size was not useful because the most abundant whey proteins, beta-lactoglobulin and alpha-lactalbumin, have molecular masses that are similar to most known growth factors. Instead, cation-exchange chromatography was selected as an enrichment process because, in contrast to the major whey proteins, growth factors generally have basic isoelectric points. Adsorption to and elution from Sepharose Fast Flow-S resin yielded an extract containing only 1 to 2% of whey protein but substantial growth-promoting activities on Balb/c 3T3 cells, L6 myoblasts, and human skin fibroblasts. The growth activity could be separated from lactoferrin, one of the prominent basic proteins present, through a stepwise elution from the resin. The resultant fraction, which contained lactoperoxidase as the most abundant protein stimulated the growth of the three cell lines at protein concentrations that were 2- to 20-fold lower than observed with fetal bovine serum. Immunoglobulin G could be removed by affinity chromatography, or lactoperoxidase could be inactivated by heat, without significant losses to the growth-promoting capacity of the fraction. These results suggest that enrichment of growth factors by cation-exchange chromatography offers a practical method for the large-scale isolation of an extract from cheese whey that stimulates cell growth.

Comparison has been made between the activity of the pineal hormone melatonin, and several analogues and metabolites in inhibiting sexual development in a protein-restricted prepubertal rat model. Eleven melatonin analogues or metabolites were tested with the aim of evaluating the model as a test of the hypothesis that melatonin acts as a prohormone and that the ring schism metabolites (kynurenamines) mediate many of the effects attributable to melatonin. Although the hypothesis could not be confirmed, modification of the melatonin structure by lengthening the acylamide side chain or by replacing the 5 methoxy function with fluorine resulted in loss of biological potency. Modification of the melatonin structure to block the two known points of metabolism resulted in no significant alteration in biological activity. Thus 6-chloromelatonin (blocking 6-hydroxylation) and 2,3-dihydromelatonin (blocking oxidative cleavage of the C2-C3 bond) and 6-chloro-2,3-dihydromelatonin remained biologically active. The metabolic products of brain indolearnine-2,3-dioxygenase, N-acetyl-N 2 -formyl-5-methoxy kynurenamine (aFoMK) and N-acetyl-5-methoxy kynurenamine (aMK), paradoxically were also biologically active.

We have previously reported the presence of a 70 kDa insulin-like growth factor (IGF)-II-specific binding protein in chicken serum using Western ligand blotting approaches. In order to ascertain the identity of this 70 kDa IGF-II binding species, the protein has been purified from chicken serum using a combination of ion-exchange and gel-permeation chromatography. Interestingly, amino acid sequencing of the purified protein revealed that it has the same N-terminal sequence as chicken vitronectin (VN). The protein has the ability to specifically bind IGF-II and not IGF-I as determined by ligand blotting, cross-linking and competitive binding assay approaches. In addition, the protein binds 125I-des(1-6)-IGF-II, suggesting that the interaction with IGF-II is different to those with other characterized IGF-binding proteins. Importantly, we have ascertained that both human and bovine VN also specifically bind IGF-II. These results are particularly relevant in the light of the recent report that the urokinase-type plasminogen activator receptor, a protein that also binds VN, has been shown to associate with the cation-independent mannose-6-phosphate/IGF-II receptor and suggest a possible role for IGF-II in cell adhesion and invasion.

We have previously reported the presence of a 70 kDa insulin-like growth factor (IGF)-II-specific binding protein in chicken serum using Western ligand blotting approaches. In order to ascertain the identity of this 70 kDa IGF-II binding species, the protein has been purified from chicken serum using a combination of ion-exchange and gel-permeation chromatography. Interestingly, amino acid sequencing of the purified protein revealed that it has the same N-terminal sequence as chicken vitronectin (VN). The protein has the ability to specifically bind IGF-II and not IGF-I as determined by ligand blotting, cross-linking and competitive binding assay approaches. In addition, the protein binds 125I-des(1-6)-IGF-II, suggesting that the interaction with IGF-II is different to those with other characterized IGF-binding proteins. Importantly, we have ascertained that both human and bovine VN also specifically bind IGF-II. These results are particularly relevant in the light of the recent report that the urokinase-type plasminogen activator receptor, a protein that also binds VN, has been shown to associate with the cation-independent mannose-6-phosphate/IGF-II receptor and suggest a possible role for IGF-II in cell adhesion and invasion.