Rami Althnaibat | ALES Graduate Seminar

A graduate exam seminar is a presentation of the student’s final research project for their degree.
This is an ALES PhD Final Exam Seminar by Rami Althnaibat. This seminar is open to the general public to attend.

https://ualberta-ca.zoom.us/j/98177681878

Thesis Topic: Identification of bioactive peptides from camel milk whey protein with potential health benefits through control of oxidative stress, glucose release after starch digestion, and pathogen adhesion

PhD with Drs. Michael Gaenzle and Heather Bruce.

Seminar Abstract:

Camel milk cheese is popular in many countries and the production of camel milk cheese has increased, as has its by-product whey. The prevalence of non-communicable diseases, mainly diet-related chronic diseases like diabetes mellitus (DM), has increased worldwide. On other hand, the enterotoxigenic Escherichia coli (ETEC) is a major cause of childhood diarrhea and diarrhea in piglets and calves. This research aimed to investigate the potential antioxidant activities, starch digestion inhibitory activities, and antiadhesive activity against ETEC of bioactive peptides from camel milk whey protein (CMWP). CMWP was hydrolyzed by flavourzyme, neutrase, alcalase, or a mixture of neutrase and flavourzyme. The antioxidant activities of the hydrolysates were determined using 2, 2′-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging, ferrous ion (Fe²⁺) chelating activity, superoxide (O^.2-) free radical scavenging, and measurement of reducing power. The antioxidant activity of flavourzyme hydrolysates (FH) at 1.5 h hydrolysis was highest and pepsin digested FH effectively to obtain peptides with high antioxidant activity, whereas trypsin digestion negatively affected antioxidant activities. The peptic hydrolysates were characterized by one sharp main peak of molecular weight (MW (6.83KDa)); however, further hydrolysis by trypsin led to hydrolysis of the peptides to amino acids and very short peptides. The low-medium sized peptides exhibited the highest antioxidant activities. Starch digestion inhibitory activity of bioactive peptides derived from CMWP was investigated using a digestibility assay that included addition of pepsin, pancreatic enzymes, and brush border enzymes. The hydrolysates from the flavourzyme hydrolysis were fractionated either by hydrophobic interaction chromatography (HIC), or by cation exchange chromatography (CEX) followed by HIC. The successive chromatographic separation aiming to produce positively charged peptide with hydrophobic amino acids improved the starch digestion inhibition. Fractions that inhibited starch digestion the most were selected for peptide sequencing by LC-MS/MS. Among the sequenced peptides, six short peptides were chosen for peptide synthesis by Canada Peptide (Quebec, Canada). The original unfractionated hydrolyzed whey showed about 16.5% inhibition of starch digestion. However, the fractionation on HIC column alone or on CEX and HIC boosted the inhibitory activity by about 63% and 116%, respectively. LALDIEIATYR and VLDELTLAR had the same activities as the entire fraction. Sweet whey from camel milk contains ~5.1% glycomacropeptide (GMP).  The GMP from camel milk was purified by ion exchange chromatography and ultra-filtration, and the purity of camel GMP determined by SDS-PAGE and mass spectrometry. The anti-adhesion activity of camel GMP against ETEC was determined using an hemagglutination assay and by enzyme-linked immunosorbent assay (ELISA).  The monosaccharide content of GMP from Bactrian camels and dromedaries was about twice as high when compared to bovine GMP. Glycans from camels included fucose and N-acetylglucosamine, which were absent in bovine GMP. GMP from both camel’s species prevented ETEC adhesion to porcine blood cells at a concentration of 0.24 g/L to 0.28 g/L respectively, a concentration that is about 20-fold lower when compared to bovine. This increased activity likely relates to the increased glycosylation and the density of glycan spacing, and / or to differences in the glycan composition. In conclusion, bioactive peptides derived from CMWP have potential inhibitory effect on oxidative stress, starch digestion, and pathogenic bacteria adhesion.