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Screening of soy protein-derived hypotriglyceridemic di-peptides in vitro and in vivo.

Inoue N, Nagao K, Sakata K, Yamano N, Gunawardena PE, Han SY, Matsui T, Nakamori T, Furuta H, Takamatsu K, Yanagita T - Lipids Health Dis (2011)

Bottom Line: In the third experiment, we found that Fraction-C (Frc-C) peptides, fractionated from hydrophilic peptides by gel permeation chromatography-high performance liquid chromatography, significantly reduced TG synthesis and apolipoprotein B (apoB) secretion in HepG2 cells.In the fourth experiment, we found that the fraction with 0.1% trifluoroacetic acid, isolated from Frc-C peptides by octadecylsilyl column chromatography, showed hypolipidemic effects in HepG2 cells.In the final experiment, we found that 3 di-peptides, Lys-Ala, Val-Lys, and Ser-Tyr, reduced TG synthesis, and Ser-Tyr additionally reduced apoB secretion in HepG2 cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Applied Biochemistry and Food Science, Saga University, Saga 840-8502, Japan.

ABSTRACT

Background: Soy protein and soy peptides have attracted considerable attention because of their potentially beneficial biological properties, including antihypertensive, anticarcinogenic, and hypolipidemic effects. Although soy protein isolate contains several bioactive peptides that have distinct physiological activities in lipid metabolism, it is not clear which peptide sequences are responsible for the triglyceride (TG)-lowering effects. In the present study, we investigated the effects of soy protein-derived peptides on lipid metabolism, especially TG metabolism, in HepG2 cells and obese Otsuka Long-Evans Tokushima fatty (OLETF) rats.

Results: In the first experiment, we found that soy crude peptide (SCP)-LD3, which was prepared by hydrolyze of soy protein isolate with endo-type protease, showed hypolipidemic effects in HepG2 cells and OLETF rats. In the second experiment, we found that hydrophilic fraction, separated from SCP-LD3 with hydrophobic synthetic absorbent, revealed lipid-lowering effects in HepG2 cells and OLETF rats. In the third experiment, we found that Fraction-C (Frc-C) peptides, fractionated from hydrophilic peptides by gel permeation chromatography-high performance liquid chromatography, significantly reduced TG synthesis and apolipoprotein B (apoB) secretion in HepG2 cells. In the fourth experiment, we found that the fraction with 0.1% trifluoroacetic acid, isolated from Frc-C peptides by octadecylsilyl column chromatography, showed hypolipidemic effects in HepG2 cells. In the final experiment, we found that 3 di-peptides, Lys-Ala, Val-Lys, and Ser-Tyr, reduced TG synthesis, and Ser-Tyr additionally reduced apoB secretion in HepG2 cells.

Conclusion: Novel active peptides with TG-lowering effects from soy protein have been isolated.

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Related in: MedlinePlus

Calibration of Superdex Peptide HR10/30 gel permeation chromatography column by using standard peptides and the elution profile of the hydrophilic fraction of soy crude peptide-LD3.
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Figure 1: Calibration of Superdex Peptide HR10/30 gel permeation chromatography column by using standard peptides and the elution profile of the hydrophilic fraction of soy crude peptide-LD3.

Mentions: Preparation of soy crude peptides (SCP)-LD3 was performed by hydrolyze of soy protein isolate with endo-type protease derived from Bacillus sp. and fractionation with centrifugation [27]. SCP-LD3 was separated into 2 fractions (hydrophobic fraction and hydrophilic fraction) on the basis of hydrophobicity by using hydrophobic synthetic absorbent (DIAION HP21; Mitsubishi Chemical Corporation, Tokyo, Japan). The amino acid composition of SCP-LD3, hydrophobic fraction, and hydrophilic fraction was analyzed with an automatic amino acid analyzer; results are shown in Table 1. Hydrophilic fraction was further fractionated by gel permeation chromatography-high performance liquid chromatography (GPC-HPLC, Bio-Rad Superdex Peptide HR10/30 column, 30% CH3CN/0.1% trifluoroacetic acid (TFA), 0.3 mL/min), and di- or tri-peptide fractions were collected at an elution time ranging from 45 to 75 min (Figure 1; Fraction A, Frc-A, 45-55 min; Fraction B, Frc-B, 55-65 min; Fraction C, Frc-C, 65-75 min). Then, 2 peptide fractions (0% CH3CN/0.1% TFA and 20% CH3CN/0.1% TFA fractions) were obtained by octadecylsilyl (ODS) open-column chromatography (20 × 60 mm) from Frc-C peptides. Finally, 0% CH3CN/0.1% TFA fraction peptides were separated by reversed-phase HPLC (COSMOSIL 5C18-AR-II column; Nakalai tesque, Kyoto, Japan), and 7 di-peptides (Figure 2; Ala-Leu, Ala-Tyr, Ala-Val, Leu-Val, Lys-Ala, Ser-Tyr, and Val-Lys) were identified. These di-peptides, except Leu-Val, could be synthesized using a Fmoc solid-phase synthesis method according to the manufacturer's instructions (Kokusan Chemicals, Osaka, Japan). The sequence of the di-peptides was confirmed on a PPSQ-21 amino acid sequencer (Shimadzu Co., Kyoto, Japan).


Screening of soy protein-derived hypotriglyceridemic di-peptides in vitro and in vivo.

Inoue N, Nagao K, Sakata K, Yamano N, Gunawardena PE, Han SY, Matsui T, Nakamori T, Furuta H, Takamatsu K, Yanagita T - Lipids Health Dis (2011)

Calibration of Superdex Peptide HR10/30 gel permeation chromatography column by using standard peptides and the elution profile of the hydrophilic fraction of soy crude peptide-LD3.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3116501&req=5

Figure 1: Calibration of Superdex Peptide HR10/30 gel permeation chromatography column by using standard peptides and the elution profile of the hydrophilic fraction of soy crude peptide-LD3.
Mentions: Preparation of soy crude peptides (SCP)-LD3 was performed by hydrolyze of soy protein isolate with endo-type protease derived from Bacillus sp. and fractionation with centrifugation [27]. SCP-LD3 was separated into 2 fractions (hydrophobic fraction and hydrophilic fraction) on the basis of hydrophobicity by using hydrophobic synthetic absorbent (DIAION HP21; Mitsubishi Chemical Corporation, Tokyo, Japan). The amino acid composition of SCP-LD3, hydrophobic fraction, and hydrophilic fraction was analyzed with an automatic amino acid analyzer; results are shown in Table 1. Hydrophilic fraction was further fractionated by gel permeation chromatography-high performance liquid chromatography (GPC-HPLC, Bio-Rad Superdex Peptide HR10/30 column, 30% CH3CN/0.1% trifluoroacetic acid (TFA), 0.3 mL/min), and di- or tri-peptide fractions were collected at an elution time ranging from 45 to 75 min (Figure 1; Fraction A, Frc-A, 45-55 min; Fraction B, Frc-B, 55-65 min; Fraction C, Frc-C, 65-75 min). Then, 2 peptide fractions (0% CH3CN/0.1% TFA and 20% CH3CN/0.1% TFA fractions) were obtained by octadecylsilyl (ODS) open-column chromatography (20 × 60 mm) from Frc-C peptides. Finally, 0% CH3CN/0.1% TFA fraction peptides were separated by reversed-phase HPLC (COSMOSIL 5C18-AR-II column; Nakalai tesque, Kyoto, Japan), and 7 di-peptides (Figure 2; Ala-Leu, Ala-Tyr, Ala-Val, Leu-Val, Lys-Ala, Ser-Tyr, and Val-Lys) were identified. These di-peptides, except Leu-Val, could be synthesized using a Fmoc solid-phase synthesis method according to the manufacturer's instructions (Kokusan Chemicals, Osaka, Japan). The sequence of the di-peptides was confirmed on a PPSQ-21 amino acid sequencer (Shimadzu Co., Kyoto, Japan).

Bottom Line: In the third experiment, we found that Fraction-C (Frc-C) peptides, fractionated from hydrophilic peptides by gel permeation chromatography-high performance liquid chromatography, significantly reduced TG synthesis and apolipoprotein B (apoB) secretion in HepG2 cells.In the fourth experiment, we found that the fraction with 0.1% trifluoroacetic acid, isolated from Frc-C peptides by octadecylsilyl column chromatography, showed hypolipidemic effects in HepG2 cells.In the final experiment, we found that 3 di-peptides, Lys-Ala, Val-Lys, and Ser-Tyr, reduced TG synthesis, and Ser-Tyr additionally reduced apoB secretion in HepG2 cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Applied Biochemistry and Food Science, Saga University, Saga 840-8502, Japan.

ABSTRACT

Background: Soy protein and soy peptides have attracted considerable attention because of their potentially beneficial biological properties, including antihypertensive, anticarcinogenic, and hypolipidemic effects. Although soy protein isolate contains several bioactive peptides that have distinct physiological activities in lipid metabolism, it is not clear which peptide sequences are responsible for the triglyceride (TG)-lowering effects. In the present study, we investigated the effects of soy protein-derived peptides on lipid metabolism, especially TG metabolism, in HepG2 cells and obese Otsuka Long-Evans Tokushima fatty (OLETF) rats.

Results: In the first experiment, we found that soy crude peptide (SCP)-LD3, which was prepared by hydrolyze of soy protein isolate with endo-type protease, showed hypolipidemic effects in HepG2 cells and OLETF rats. In the second experiment, we found that hydrophilic fraction, separated from SCP-LD3 with hydrophobic synthetic absorbent, revealed lipid-lowering effects in HepG2 cells and OLETF rats. In the third experiment, we found that Fraction-C (Frc-C) peptides, fractionated from hydrophilic peptides by gel permeation chromatography-high performance liquid chromatography, significantly reduced TG synthesis and apolipoprotein B (apoB) secretion in HepG2 cells. In the fourth experiment, we found that the fraction with 0.1% trifluoroacetic acid, isolated from Frc-C peptides by octadecylsilyl column chromatography, showed hypolipidemic effects in HepG2 cells. In the final experiment, we found that 3 di-peptides, Lys-Ala, Val-Lys, and Ser-Tyr, reduced TG synthesis, and Ser-Tyr additionally reduced apoB secretion in HepG2 cells.

Conclusion: Novel active peptides with TG-lowering effects from soy protein have been isolated.

Show MeSH
Related in: MedlinePlus