Spectrophotometric Assay Using o-Phthaldialdehyde for Determination of Proteolysis in Milk and Isolated Milk Proteins

Church, Frank C.; Swaisgood, Harold E.; Porter, David H.; Catignani, George L.

doi:10.3168/jds.s0022-0302(83)81926-2

Cited by 1,325 publications

(716 citation statements)

References 28 publications

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“…Measurement of protein solubility and peptide content The soluble contents of skin and bone gelatins and their hydrolysates were determined by the Folin-Lowry method (Lowry et al 1951) and their peptide contents measured by method of Church et al (1983) using o-phthaldialdehyde (OPA) spectrophotometric assay. Each hydrolysate, containing 5-100 μg protein, was mixed with OPA reagent and the absorbance measured at 340 nm.…”

Section: Methodsmentioning

confidence: 99%

ACE inhibitory activity of pangasius catfish (Pangasius sutchi) skin and bone gelatin hydrolysate

et al. 2012

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Skin and bone gelatins of pangasius catfish (Pangasius sutchi) were hydrolyzed with alcalase to isolate Angiotensin Converting Enzyme (ACE) inhibitory peptides. Samples with the highest degree of hydrolysis (DH) were separated into different fractions with molecular weight cutoff (MWCO) sizes of 10, 3 and 1 kDa, respectively and assayed for ACE inhibitory activity. Skin and bone gelatins had highest DH of 64.87 and 68.48 % after 2 and 1 h incubation, respectively. Results from this study indicated that by decreasing the molecular weight of fractions, ACE inhibitory activity was increased. Therefore, F 3 permeates (MWCO< 1 kDa) of skin (IC 50 03.2 μg/ml) and bone (IC 50 01.3 μg/ml) gelatins possessed higher ACE inhibitory activity compared to their untreated gelatins and corresponding hydrolyzed fractions. In this study, the major amino acids were Glycine followed by Proline with an increased amount of hydrophobic amino acid content in F 3 permeates of skin (4.01 %) and bone (5.79 %) gelatin. Digestion stability against gastrointestinal proteases did not show any remarkable change on ACE inhibition potency of these permeates. It was concluded that alcalase hydrolysis of P. sutchi by-products could be utilized as a part of functional food or ingredients of a formulated drug in order to control high blood pressure.

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Section: Methodsmentioning

confidence: 99%

ACE inhibitory activity of pangasius catfish (Pangasius sutchi) skin and bone gelatin hydrolysate

et al. 2012

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“…The amount of free amino groups of casein, casein hydrolysate or the modified hydrolysate on protein or peptide basis was measured by o-phthaldialdehyde (OPA) assay (Church et al 1983;Spellman et al 2003) with some modifications. OPA reagent was prepared by combing following reagents and diluting them to a final volume of 100 mL with water: 75 mL 0.2 mol⋅L −1 sodium borate buffer (pH 9.5), 5 mL 400 g SDS⋅L −1 , 80 mg OPA (in 1 mL methanol) and 0.4 mL β-mercaptoenthanol.…”

Section: Analysis Of Protease Activity and Protein Or Peptide Contentmentioning

confidence: 99%

Optimization of some conditions of Neutrase-catalyzed plastein reaction to mediate ACE-inhibitory activity in vitro of casein hydrolysate prepared by Neutrase

Kong

Zhao

2011

J Food Sci Technol

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Casein hydrolysate was prepared by hydrolyzing casein with Neutrase and then modified by a Neutrasecatalyzed plastein reaction. The prepared hydrolysate had a degree of hydrolysis of 13.0% and exhibited ACE inhibition in vitro with an IC 50 value of 40.4 μg⋅mL −1 . With the decreased amount of free amino groups of the modified hydrolysate as the response, some conditions of the plastein reaction including substrate concentration, enzyme to substrate ratio, reaction temperature and time were studied by single factor experiments and response surface methodology, and optimized finally as 62% (w/w), 3.0 kU⋅g −1 peptides, 30°C and 6.3 h, respectively. The maximum decreased amount of free amino groups of the modified hydrolysate prepared under these optimized conditions was 210.0 μmol⋅g −1 peptides, while corresponding IC 50 value was lowered to 14.7 μg⋅mL −1 . The present result indicates that Neutrase-catalyzed plastein reaction was capable of enhancing ACE-inhibitory activity in vitro of casein hydrolysate, and also highlights the importance of a forthcoming study to investigate the peptide compositions of the modified hydrolysate and the role of protease used in the plastein reaction.

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“…The degree of hydrolysis (OH) of each fraction was evaluated in triplicate by measuring the amount of c-arnino nitrogen with the o-phthaldialdehyde spectrophotometric method (OPA) using t-leucine as standard (Church et al, 1983). Total content of peptide bonds used for calculation of OH was 8.2 meqv g-' protein (Adler-Nissen, 1977).…”

Section: Chemical Analysismentioning

confidence: 99%

Comparison of bifidobacterial growth-promoting activity of ultrafiltered casein hydrolyzate fractions

Proulx

Ward²,

Gauthier

et al. 1994

Lait

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Summary -Growth rates and acid production of the most common dairy-related bifidobacteria (Bifidobacterium infantis, B breve and B longum) were determined in the presence of casein hydrolyzates produced by the action of three proteolytic enzymes (alcalase, chymotrypsin and trypsin). Casein hydrolyzates were fractionated with a two-step ultrafiltration process to study the effect of molecular mass of peptides on bifidobacterial growth. The retentate of the second ultrafiltration (nominal molecular eut-off of membranes was 1000 Da) was called mixture of polypeptides (MP) and the permeate fraction which was mainly composed of free amino acids and small peptides was called AA. These hydrolyzate fractions were characterized by a very different concentration of some amino acids (glu, tyr, phe). Among MP and AA casein fractions, trypsin MP fraction at a final concentration of 2% in synthetic medium (Garches medium) exhibited a higher growth-promoting activity on the three bifidobacterial species tested. However, addition of alcalase AA fraction at a final concentration of 1 or 2% repressed the growth and acid production of B breve and B longum.

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Spectrophotometric Assay Using o-Phthaldialdehyde for Determination of Proteolysis in Milk and Isolated Milk Proteins

Cited by 1,325 publications

References 28 publications

ACE inhibitory activity of pangasius catfish (Pangasius sutchi) skin and bone gelatin hydrolysate

ACE inhibitory activity of pangasius catfish (Pangasius sutchi) skin and bone gelatin hydrolysate

Optimization of some conditions of Neutrase-catalyzed plastein reaction to mediate ACE-inhibitory activity in vitro of casein hydrolysate prepared by Neutrase

Comparison of bifidobacterial growth-promoting activity of ultrafiltered casein hydrolyzate fractions

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