Purification and Characterization of an Opioid Antagonist from a Peptic Digest of Bovine κ-Casein

Yoshikawa, Masaaki; Tani, Fumito; Ashikaga, Tadaaki; Yoshimura, Takashi; Chiba, Hideo

doi:10.1080/00021369.1986.10867863

Cited by 12 publications

(15 citation statements)

References 2 publications

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“…1433 Many biologically active peptides have been discovered, some of which are parts of the amino acid sequence in dietary proteins such as casein. 1 ) Dipeptides and tripeptides are assimilated by the brush-border membrane of the intestinal tract more quickly than free amino acids, for which the absorption mechanism is different. 2 ) In this sense, peptides are superior to the free amino acids with which they are constructed.…”

Section: Continuous Synthesis Of a Tripeptide By Successive Condensatmentioning

confidence: 99%

Continuous Synthesis of a Tripeptide by Successive Condensation and Transesterification Catalyzed by Two Immobilized Proteinases in Organic Solvent

Kimura¹,

Yoshida²,

Muraya³

et al. 1990

Agricultural and Biological Chemistry

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The tripeptide Z-GlyPheLeuNH2 was continuously synthesized in a high yield from three amino acid derivatives, Z-Gly, PheOMe, and LeuNH2, by immobilized thermolysin (IMT) and immobilized alpha-chymotrypsin (IMC) in an organic solvent, ethyl acetate. The optimal conditions for the synthesis of Z-GlyPheOMe were established theoretically. The yield of Z-GlyPheOMe with IMT in ethyl acetate saturated with buffer was more than 88% after continuous synthesis for 116 hr. The optimal conditions for the synthesis of Z-GlyPheLeuNH2 from Z-GlyPheOMe and LeuNH2 by IMC through transesterification was established in batch reaction experiments. When the concentration of water in the reaction solution was 17-20 microliters/ml, the activity of IMC was highest. The equilibrium between the water concentration in the reaction solution and that in the resin used for enzyme immobilization depended on the resin and was not affected by the presence of the enzyme immobilized. Z-GlyPheLeuNH2 was synthesized from Z-GlyPheOMe and LeuNH2 with a yield of 100%, by continuous reaction for 160 hr. The reactor for synthesis of this tripeptide was efficient and stable because of the use of transesterification and the choice of an appropriate organic solvent. The series plug-flow reactor was successfully operated for 220 hr with a yield of more than 80%. The residual activity of IMT was 94% and that of IMC was 100%.

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Section: Continuous Synthesis Of a Tripeptide By Successive Condensatmentioning

confidence: 99%

Continuous Synthesis of a Tripeptide by Successive Condensation and Transesterification Catalyzed by Two Immobilized Proteinases in Organic Solvent

Kimura¹,

Yoshida²,

Muraya³

et al. 1990

Agricultural and Biological Chemistry

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“…A less recognized and more diversified (in terms of chemical structure) group of opioid peptides, the MOP receptor antagonistic peptides, named casoxins, consist of 4-10 amino acid fragments of bovine k-casein (f25-34, f33-38, f35-38, f35-41, f58-61); (Chiba et al, 1989;Yoshikawa et al, 1986). Except for casoxin B (f58-61), the possibility of releasing the other casoxins from the native protein by trypsin and/or pepsin has been shown, but only in vitro.…”

Section: Introductionmentioning

confidence: 99%

Transport of bovine milk-derived opioid peptides across a Caco-2 monolayer

Sienkiewicz-Szłapka

Jarmołowska

Krawczuk

et al. 2009

International Dairy Journal

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“…The μ-specific opioid antagonist, casoxin 4, one of the smallest and most potent opioid antagonist peptides yet described, is a synthetic tetrapeptide fragment of casoxin 6 24,25. The potency of casoxin 4 to antagonize morphine inhibition of electrically driven contractions is shown in Figure 3.…”

Section: Resultsmentioning

confidence: 99%

Effects of casoxin 4 on morphine inhibition of small animal intestinal contractility and gut transit in the mouse

Patten

Head²,

Abeywardena³

2011

CEG

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Background and aims:Chronic opioid analgesia has the debilitating side-effect of constipation in human patients. The major aims of this study were to: 1) characterize the opioid-specific antagonism of morphine-induced inhibition of electrically driven contraction of the small intestine of mice, rats, and guinea pigs; and 2) test if the oral delivery of small milk-derived opioid antagonist peptides could block morphine-induced inhibition of intestinal transit in mice.Methods:Mouse, rat, and guinea pig intact ileal sections were electrically stimulated to contract and inhibited with morphine in vitro. Morphine inhibition was then blocked by opioid subtype antagonists in the mouse and guinea pig. Using a polymeric dye, Poly R-478, the opioid antagonists casoxin 4 and lactoferroxin A were tested orally for blocking activity of morphine inhibition of gut transit in vivo by single or double gavage techniques.Results:The guinea pig tissue was more sensitive to morphine inhibition compared with the mouse or the rat (IC50 [half maximal inhibitory concentration] values as nmol/L ± SEM were 34 ± 3, 230 ± 13, and 310 ± 14 respectively) (P < 0.01). The inhibitory influence of opioid agonists (IC50) in electrically driven ileal mouse preparations were DADLE ([D-Ala2, D-Leu5]-enkephalin) ≥ met-enkephalin ≥ dynorphin A ≥ DAMGO ([D-Ala2, N-Me-Phe4, Glyol5]-enkephalin) > morphine > morphiceptin as nmol/L 13.9, 17.3, 19.5, 23.3, 230, and 403 respectively. The mouse demonstrated predominantly κ- and δ-opioid receptor activity with a smaller μ-opioid receptor component. Both mouse and guinea pig tissue were sensitive to casoxin 4 antagonism of morphine inhibition of contraction. In contrast to naloxone, relatively high oral doses of the μ-opioid receptor antagonists, casoxin 4 and lactoferroxin A, applied before and after morphine injection were unable to antagonize morphine inhibition of gut transit.Conclusions:Casoxin 4 reverses morphine-induced inhibition of contraction in mice and guinea pigs in vitro but fails to influence morphine inhibition of mouse small intestinal transit by the oral route.

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Purification and Characterization of an Opioid Antagonist from a Peptic Digest of Bovine κ-Casein

Cited by 12 publications

References 2 publications

Continuous Synthesis of a Tripeptide by Successive Condensation and Transesterification Catalyzed by Two Immobilized Proteinases in Organic Solvent

Continuous Synthesis of a Tripeptide by Successive Condensation and Transesterification Catalyzed by Two Immobilized Proteinases in Organic Solvent

Transport of bovine milk-derived opioid peptides across a Caco-2 monolayer

Effects of casoxin 4 on morphine inhibition of small animal intestinal contractility and gut transit in the mouse

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