AIM： To isolate and identify the soybean conglycinin peptides that selectively stimulates the growth of bifidobacteria in vitro, and to investigate the effect of soybean conglycinin peptides on intestinal ecosystem i...AIM： To isolate and identify the soybean conglycinin peptides that selectively stimulates the growth of bifidobacteria in vitro, and to investigate the effect of soybean conglycinin peptides on intestinal ecosystem in vivo. METHODS： Soybean conglycinin was purified from soybean seeds by gel filtration （Sepharose-CL-6B）. These proteins were submitted to hydrolysis by pepsin. Several growth-stimulating peptides for bifidobacteria were isolated chromatographically from pepsin hydrolysis of soybean conglycinin and identified by means of matrixassisted laser desorption ionization time of flight mass spectrometry （MALDI-TOF-MS）. Parallel to in vitro study, in vivo experiments with soybean conglycinin peptides were performed in mice. Ninety male KM mice were randomly assigned into five groups of 16 mice each, and each group was administered for 21d intragastrically with physiological saline （control）, conglycinin, pepsin-treated conglycinin （PTC）, the most active fraction which isolated from pepsin-treated conglycinin （P2-PTC） and HCl-full hydrolysis of conglycinin （HCl-FHC）, respectively. Intestinal microflora were evaluated by standard microbiologic methods and biochemical assays of cecal content samples after treatment. RESULTS： The results showed that the peptides which were isolated from soybean conglycinin could stimulate the growth of bifidobacteria in vitro, and the molecular mass of purified peptides with MALDI-TOF-MS ranged from 693.32 to 1829.55. Compared with control group, in vivo experiments showed that P2-PTC group decreased cecal pH （7.08±0.08 vs7.21±0.09, P〈0.05） and enterococcicounts （5.38±0.26 log10CFU/g vs 5.78±0.19 log10CFU/g, P〈0.05）, significantly increased sIgA level （172.08±35.40 ng/g vs 118.27±33.93 ng/g, P〈0.01） and β-galactosidase activity （1.28±0.23 U/g vs 1.82±0.58 U/g, P〈0.05） CONCLUSION： The results have shown that conglycinin is good source for enzyme-mediated production of peptides which stimulate the growth of bifidobacteria. These peptides are inactive within the sequence of the parent protein but can be released during enzymatic hydrolysis, and in vivo experiments demonstrate that conglycinin peptides may be beneficial for improving gastrointestinal health.展开更多
β-Conglycinin,the main component of 7S globulin in soybean protein,is also a key soybean antigen protein that causes allergic reactions.Extrusion technologies have received considerable attention as amethod for modif...β-Conglycinin,the main component of 7S globulin in soybean protein,is also a key soybean antigen protein that causes allergic reactions.Extrusion technologies have received considerable attention as amethod for modifying soybean protein allergens.This study investigated the changes inβ-conglycinin structure and antigenicity upon extrusion.Isoelectric precipitation,ammoniumsulfate precipitation,and sepharose CL-6B gel filtration were used to isolate and purifyβ-conglycinin from soybean powder,and single-factor and orthogonal tests were used to study the effects of water content,extrusion temperature,screw rotation speed,and feeding speed on the antigenicity ofβ-conglycinin after extrusion.Fourier transforminfrared spectrometry(FTIR)was then employed to analyze the structure ofβ-conglycinin after extrusion under the optimal conditions determined by the orthogonal test.The results showed that extrusion significantly reduced the antigenicity ofβ-conglycinin(P<0.05),and the degree of influence of the factors studied may be ordered as extrusion temperature>feeding speed>screw rotation speed>water content.The optimal parameters were temperature at 130°C,screwrotation speed at 140 r/min,and feeding speed at 35 g/min.Under these conditions,the contents ofα-helix,β-pleated sheet,andβ-turn structures inβ-conglycinin were significantly reduced(P<0.05),while the contents of random coils were significantly increased(P<0.05).The peak absorption intensity of amides I,II,and III also decreased.Taken together,the findings suggest that extrusion could be an effective method for reducing the antigenicity ofβ-conglycinin.展开更多
[Objective] The paper was to investigate the protective effects of different concentrations of Bacillus subtilis peptidoglycan(PG) on β-conglycinin-induced inflammatory injury in intestinal epithelial cells of juveni...[Objective] The paper was to investigate the protective effects of different concentrations of Bacillus subtilis peptidoglycan(PG) on β-conglycinin-induced inflammatory injury in intestinal epithelial cells of juvenile carp(Cyprinus carpio).[Method] In 24-cell microplates, the intestinal epithelial cells(IECs) of juvenile carp were primarily cultured for 72 h at 26°C and 6% CO2, and then the IECs were randomly divided into6 groups with 4 replicates per group. One of the six groups was set as negative control group, and the other groups were all supplemented with 1.0 mg/mL β-conglycinin in culture medium to establish inflammatory injury. At 24 h post induction, the culture media were changed into B. subtilis PG culture media with the concentrations of 0(positive control group), 0.15, 0.30, 0.45 and 0.60 mg/mL, respectively. The samples were collected to measure the antioxidant and anti-inflammatory indices at 12, 24 and 36 h post culture.[Result]β-conglycinin exposure significantly decreased the activity of ASA, AHR, SOD, CAT, GPx, and increased the PC content and the mRNA expression of inflammatory cytokines(IL-1β, IL-8, TNF-α1,IL-10 and TGF-β). At 12, 24 and 36 h post PG treatment, the activities of ASA, AHR, SOD, CAT, GPx and the content of PC in cells decreased in a dose-dependent manner;the mRNA levels of IL-1β, IL-8 and TNF-α1 were down-regulated and those of IL-10 and TGF-β were up-regulated.[Conclusion] Different concentrations of B. subtilis PG could protect IECs oxidative damage induced by β-conglycinin and improve the antioxidant capacity of IECs. High concentration of PG could improve the anti-inflammatory ability of IECs by inhibiting inflammatory factors and promoting the gene expression of anti-inflammatory cytokines.展开更多
By now, the digestive stability experiments provided by most authoritative organizations are in vitro tests. Evaluating the protein digestive stability with in vivo models should be more objective. The present study a...By now, the digestive stability experiments provided by most authoritative organizations are in vitro tests. Evaluating the protein digestive stability with in vivo models should be more objective. The present study aimed to verify the in vivo digestibility of soybean β-conglycinin β-subunit in Wuzhishan （WZS） minipigs. Three minipigs were surgically fitted with O-stomach and T-ileum cannulae and fed with soybean meals. According to SDS-PAGE, the 50 kD fraction of soybean β-conglycinin β-subunit persisted in the gastric fluid until 6 h after feeding, which was detected at 3 h and clearly visible at 4-6 h in the intestinal fluid. Western blot with anti-β-conglycinin β-subunit McAb confirmed it.展开更多
The experiment was conducted to study the effects and possible mechanism of GLP-2 on proliferation,metabolism and apoptosis of cultured enterocytes from a 28-d weaned piglet injured by exposure to β-conglycinin.A cel...The experiment was conducted to study the effects and possible mechanism of GLP-2 on proliferation,metabolism and apoptosis of cultured enterocytes from a 28-d weaned piglet injured by exposure to β-conglycinin.A cell damage model was established to investigate cell proliferation, metabolism and apoptosis by exposing primary cell cultures of intestinal epithelial cells(IEC) to 1.2 and 2.4 mg/mL β-conglycinin.A 2×3 factorial experiment was then used to study the effect of different GLP-2 concentrations of(1×10<sup>-9</sup>,1×10<sup>-8</sup> and 1×10<sup>-7</sup>mol/L),in combination with the two concentrations ofβ-conglycinin.Cells exposed to the allergenβ-conglycinin had decreased(P【0.05) MTT OD;decreased (P【0.01) protein retention and total protein content of cells;increased(P【0.01) LDH and caspase-3 activities and decreased(P【0.05) Na<sup>+</sup>,K<sup>+</sup>-ATPase activity.When GLP-2 was used in combination withβ-conglycinin,MTT OD,protein retention,total protein content and Na<sup>+</sup>,K<sup>+</sup>-ATPase activity significantly increased(P【0.05);LDH activity gradually decreased(P【0.05 or P【0.01) and Caspase-3 activity significantly decreased(P【0.01) with increasing concentrations of GLP-2.The results indicated thatβ-conglycinin had adverse effects on proliferation and integrity of IEC in vitro.GLP-2 relieved or prevented the adverse effects ofβ-conglycinin on proliferation and integrity of IEC by regulating Na<sup>+</sup>,K<sup>+</sup>- ATPase and Caspase-3 activities,and consequently affecting cell metabolism.展开更多
基金Supported by the National Key Basic Research Development Program of China, 973 Program, No. 2004CB117505
文摘AIM： To isolate and identify the soybean conglycinin peptides that selectively stimulates the growth of bifidobacteria in vitro, and to investigate the effect of soybean conglycinin peptides on intestinal ecosystem in vivo. METHODS： Soybean conglycinin was purified from soybean seeds by gel filtration （Sepharose-CL-6B）. These proteins were submitted to hydrolysis by pepsin. Several growth-stimulating peptides for bifidobacteria were isolated chromatographically from pepsin hydrolysis of soybean conglycinin and identified by means of matrixassisted laser desorption ionization time of flight mass spectrometry （MALDI-TOF-MS）. Parallel to in vitro study, in vivo experiments with soybean conglycinin peptides were performed in mice. Ninety male KM mice were randomly assigned into five groups of 16 mice each, and each group was administered for 21d intragastrically with physiological saline （control）, conglycinin, pepsin-treated conglycinin （PTC）, the most active fraction which isolated from pepsin-treated conglycinin （P2-PTC） and HCl-full hydrolysis of conglycinin （HCl-FHC）, respectively. Intestinal microflora were evaluated by standard microbiologic methods and biochemical assays of cecal content samples after treatment. RESULTS： The results showed that the peptides which were isolated from soybean conglycinin could stimulate the growth of bifidobacteria in vitro, and the molecular mass of purified peptides with MALDI-TOF-MS ranged from 693.32 to 1829.55. Compared with control group, in vivo experiments showed that P2-PTC group decreased cecal pH （7.08±0.08 vs7.21±0.09, P〈0.05） and enterococcicounts （5.38±0.26 log10CFU/g vs 5.78±0.19 log10CFU/g, P〈0.05）, significantly increased sIgA level （172.08±35.40 ng/g vs 118.27±33.93 ng/g, P〈0.01） and β-galactosidase activity （1.28±0.23 U/g vs 1.82±0.58 U/g, P〈0.05） CONCLUSION： The results have shown that conglycinin is good source for enzyme-mediated production of peptides which stimulate the growth of bifidobacteria. These peptides are inactive within the sequence of the parent protein but can be released during enzymatic hydrolysis, and in vivo experiments demonstrate that conglycinin peptides may be beneficial for improving gastrointestinal health.
基金NSFC-Joint Research Fund of Henan (U1404323), Grain & Corn Engineering Technology Research CenterState Administration of Grain (GA2017004)Science and Technology Research Project of Henan (172102110205 and 182102310676) for providing funds
文摘β-Conglycinin,the main component of 7S globulin in soybean protein,is also a key soybean antigen protein that causes allergic reactions.Extrusion technologies have received considerable attention as amethod for modifying soybean protein allergens.This study investigated the changes inβ-conglycinin structure and antigenicity upon extrusion.Isoelectric precipitation,ammoniumsulfate precipitation,and sepharose CL-6B gel filtration were used to isolate and purifyβ-conglycinin from soybean powder,and single-factor and orthogonal tests were used to study the effects of water content,extrusion temperature,screw rotation speed,and feeding speed on the antigenicity ofβ-conglycinin after extrusion.Fourier transforminfrared spectrometry(FTIR)was then employed to analyze the structure ofβ-conglycinin after extrusion under the optimal conditions determined by the orthogonal test.The results showed that extrusion significantly reduced the antigenicity ofβ-conglycinin(P<0.05),and the degree of influence of the factors studied may be ordered as extrusion temperature>feeding speed>screw rotation speed>water content.The optimal parameters were temperature at 130°C,screwrotation speed at 140 r/min,and feeding speed at 35 g/min.Under these conditions,the contents ofα-helix,β-pleated sheet,andβ-turn structures inβ-conglycinin were significantly reduced(P<0.05),while the contents of random coils were significantly increased(P<0.05).The peak absorption intensity of amides I,II,and III also decreased.Taken together,the findings suggest that extrusion could be an effective method for reducing the antigenicity ofβ-conglycinin.
基金Supported by Science and Technology Research Project of Henan Province(172102110205)
文摘[Objective] The paper was to investigate the protective effects of different concentrations of Bacillus subtilis peptidoglycan(PG) on β-conglycinin-induced inflammatory injury in intestinal epithelial cells of juvenile carp(Cyprinus carpio).[Method] In 24-cell microplates, the intestinal epithelial cells(IECs) of juvenile carp were primarily cultured for 72 h at 26°C and 6% CO2, and then the IECs were randomly divided into6 groups with 4 replicates per group. One of the six groups was set as negative control group, and the other groups were all supplemented with 1.0 mg/mL β-conglycinin in culture medium to establish inflammatory injury. At 24 h post induction, the culture media were changed into B. subtilis PG culture media with the concentrations of 0(positive control group), 0.15, 0.30, 0.45 and 0.60 mg/mL, respectively. The samples were collected to measure the antioxidant and anti-inflammatory indices at 12, 24 and 36 h post culture.[Result]β-conglycinin exposure significantly decreased the activity of ASA, AHR, SOD, CAT, GPx, and increased the PC content and the mRNA expression of inflammatory cytokines(IL-1β, IL-8, TNF-α1,IL-10 and TGF-β). At 12, 24 and 36 h post PG treatment, the activities of ASA, AHR, SOD, CAT, GPx and the content of PC in cells decreased in a dose-dependent manner;the mRNA levels of IL-1β, IL-8 and TNF-α1 were down-regulated and those of IL-10 and TGF-β were up-regulated.[Conclusion] Different concentrations of B. subtilis PG could protect IECs oxidative damage induced by β-conglycinin and improve the antioxidant capacity of IECs. High concentration of PG could improve the anti-inflammatory ability of IECs by inhibiting inflammatory factors and promoting the gene expression of anti-inflammatory cytokines.
基金supported by the Key Science Project ‘973’ Award(2007CB109207) of the National Science and Technology Committee of Chinathe Key Science Project on GMO Award(2014ZX08011005)of the Ministry of Agriculture of China
文摘By now, the digestive stability experiments provided by most authoritative organizations are in vitro tests. Evaluating the protein digestive stability with in vivo models should be more objective. The present study aimed to verify the in vivo digestibility of soybean β-conglycinin β-subunit in Wuzhishan （WZS） minipigs. Three minipigs were surgically fitted with O-stomach and T-ileum cannulae and fed with soybean meals. According to SDS-PAGE, the 50 kD fraction of soybean β-conglycinin β-subunit persisted in the gastric fluid until 6 h after feeding, which was detected at 3 h and clearly visible at 4-6 h in the intestinal fluid. Western blot with anti-β-conglycinin β-subunit McAb confirmed it.
基金supported by Program for Changjiang Scholars and Innovative Reseach Team in University (IRTO 555)Applied Basic Research(045Y029-031) of Sichuan Province,People's Republic of China
文摘The experiment was conducted to study the effects and possible mechanism of GLP-2 on proliferation,metabolism and apoptosis of cultured enterocytes from a 28-d weaned piglet injured by exposure to β-conglycinin.A cell damage model was established to investigate cell proliferation, metabolism and apoptosis by exposing primary cell cultures of intestinal epithelial cells(IEC) to 1.2 and 2.4 mg/mL β-conglycinin.A 2×3 factorial experiment was then used to study the effect of different GLP-2 concentrations of(1×10<sup>-9</sup>,1×10<sup>-8</sup> and 1×10<sup>-7</sup>mol/L),in combination with the two concentrations ofβ-conglycinin.Cells exposed to the allergenβ-conglycinin had decreased(P【0.05) MTT OD;decreased (P【0.01) protein retention and total protein content of cells;increased(P【0.01) LDH and caspase-3 activities and decreased(P【0.05) Na<sup>+</sup>,K<sup>+</sup>-ATPase activity.When GLP-2 was used in combination withβ-conglycinin,MTT OD,protein retention,total protein content and Na<sup>+</sup>,K<sup>+</sup>-ATPase activity significantly increased(P【0.05);LDH activity gradually decreased(P【0.05 or P【0.01) and Caspase-3 activity significantly decreased(P【0.01) with increasing concentrations of GLP-2.The results indicated thatβ-conglycinin had adverse effects on proliferation and integrity of IEC in vitro.GLP-2 relieved or prevented the adverse effects ofβ-conglycinin on proliferation and integrity of IEC by regulating Na<sup>+</sup>,K<sup>+</sup>- ATPase and Caspase-3 activities,and consequently affecting cell metabolism.
