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Mariela Anahí Bruno - One of the best experts on this subject based on the ideXlab platform.
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Peptidases from Maclura pomifera for Preparation of Food Protein Hydrolysates: Purification by Single-Step Chromatography and Characterization of Pomiferin I
Applied Biochemistry and Biotechnology, 2020Co-Authors: Andrea Milagros Reyes Jara, Constanza Silvina Liggieri, María Alicia Corrons, Lucía Salese, Mariela Anahí BrunoAbstract:Our objective was to isolate peptidases from the latex of Maclura pomifera fruits and use them to hydrolyze food proteins, as well as to purify and characterize the main peptidase. Two partially purified proteolytic extracts were prepared by ethanol (EE) and acetone (AE) precipitation from an aqueous suspension of exuded fruit latex. EE was used to hydrolyze food proteins with a ratio of 0.19 caseinolytic units (Ucas) per mg of substrate. Different values of hydrolysis degree were observed for hydrolysates of egg white, soy protein isolate, and casein at 180 min (9.3%, 31.1%, and 29.1%, respectively). AE was employed to purify a peptidase which exhibited an isoelectric point (pI) of 8.70 and whose abundance in AE was 28.3%. This enzyme was purified to homogeneity using a single-step procedure by cation-exchange chromatography, achieving an 8.1-fold purification and a yield of 16.7%. The peptidase was named pomiferin I and showed a molecular mass of 63,177.77 Da. Kinetic constants (K_M 0.84 mM, V_max 27.50 uM s^−1, k_cat 72.37 s^−1, and k_cat/K_M 86.15 mM^−1 s^−1) were determined employing N-α-carbobenzoxy-L-alanyl-p-nitrophenyl ester as substrate. Analysis by PMF showed only partial homology of pomiferin I with a serine peptidase from a species of the same family.
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Preparation of soy protein hydrolysates with antioxidant activity by using peptidases from latex of Maclura pomifera fruits.
Food Chemistry, 2018Co-Authors: Andrea Milagros Reyes Jara, Constanza Silvina Liggieri, Mariela Anahí BrunoAbstract:Abstract A partially purified proteolytic extract prepared from Maclura pomifera latex was employed in hydrolyzing a soybean-protein isolate (4.2 mg/mL). The hydrolysis-product formation, monitored by tricine–sodium-dodecyl-sulfate–polyacrylamyde-gel electrophoresis and reverse-phase high-performance liquid chromatography, indicated that after 10 min of reaction the main soybean proteins disappeared. The maximum degree of hydrolysis was 36.2% after a 180-min digestion. The 90-min hydrolysate presented an IC50 of 31.6 ± 0.2 µg/mL, and a trolox equivalent antioxidant capacity of 157.6 and 176.9 µmoles TE per g of peptide determined by two different methods. Analysis by matrix-assisted–laser-desorption-ionization–time-of-flight mass spectrometry (MALDI-TOF MS), followed by the application of bioinformatics tools, enabled the deduction of fourteen theoretical peptide sequences containing antioxidant amino acids at >60%, none of which sequences had been previously reported as antioxidants. Finally, we consider that this 90-min hydrolysate would constitute a promising ingredient in the manufacture of functional foods.
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ACE-inhibitory peptides from bovine caseins released with peptidases from Maclura pomifera latex.
Food Research International, 2017Co-Authors: María Alicia Corrons, Sebastián A. Trejo, Constanza Silvina Liggieri, Mariela Anahí BrunoAbstract:Abstract In work reported here, a proteolytic extract prepared from Maclura pomifera latex was employed to hydrolyze bovine caseins. Densitograms of Tricine–sodium-dodecyl-sulfate–polyacrylamide-gel electrophoresis (SDS-PAGE) indicated that the caseins were considerably degraded after a 10-min reaction. The degree of hydrolysis determined by the 2,4,6-trinitrobenzenesulfonic-acid method was 17.1 ± 0.7% after 180 min of digestion. The concentration of small peptides increased with hydrolysis time, and analysis by reverse-phase high-performance liquid chromatography (RP HPLC) and mass spectrometry, revealed a virtually unchanged peptide profile. These results suggested that those proteases were highly specific, as only certain peptide bonds were cleaved. The hydrolysate of 180 min displayed the highest inhibition of angiotensin-converting enzyme (ACE) showing an IC 50 of 1.72 ± 0.25 mg/mL, and the analysis of the peptide fractionation in this hydrolysate by RP HPLC exhibited two peaks responsible for that activity. Fragmentation analysis through the use of iterated matrix-assisted–laser-desorption-ionization–time-of-flight mass spectrometry (MALDI-TOF/TOF MS/MS) with the aid of bioinformatics tools enabled us to deduce two peptide sequences—one, YQEPVLGPVRGPFPIIV, having been previously reported as an ACE-inhibitor; the other, RFFVAPFPE, as yet undescribed. The presence of bioactive peptides in these casein hydrolysates argues for their potential use in the development of functional foods.
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Application of peptidases from Maclura pomifera fruit for the production of active biopeptides from whey protein
LWT - Food Science and Technology, 2015Co-Authors: Juan Ignacio Bertucci, Constanza Silvina Liggieri, Maria Laura Colombo, Sandra Elizabeth Vairo Cavalli, Mariela Anahí BrunoAbstract:A crude extract containing serine peptidases, was prepared from latex of Maclura pomifera fruits. Peptidases were isolated by precipitation with one volume of ethanol with a yield of 5.4 ± 0.4 Ucas per milligram of protein. This extract was used for hydrolysis of bovine whey proteins at 45 °C and pH 6.5. Proteolytic activity was 99% inactivated after 5 min of heat treatment (100 °C). Major whey proteins degradation profile was analysed by tricine SDS-PAGE. After 180 min of hydrolysis alpha-lactalbumin (α-LA) and beta-lactoglobulin (β-LG) were almost completely degraded. Hydrolysis degree was 31.3 ± 1.7% at 180 min of reaction and the peptides produced that were smaller than 3 kDa were analysed by reversed-phase high-performance liquid chromatography (RP-HPLC). Angiotensin-converting enzyme (ACE) inhibitory activity and antioxidant capacity were detected in the hydrolysates and IC50 values for 180 min of hydrolysis were 0.53 ± 0.02 and 4.44 ± 0.44 mg/ml, respectively. One peptide sequence deduced from peptide masses in the 180 min filtered hydrolysate, coincided with an ACE-inhibitory peptide reported by other author. The results support the conclusion that, by the presence of ACE-inhibitory and antioxidant peptides, it would be possible to use these whey protein hydrolysates for functional food manufacturing.
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Milk clotting activity and production of bioactive peptides from whey using Maclura pomifera proteases
LWT - Food Science and Technology, 2012Co-Authors: María Alicia Corrons, Laura M. I. López, Constanza Silvina Liggieri, Juan Ignacio Bertucci, Mariela Anahí BrunoAbstract:Abstract A crude extract named pomiferin was obtained from latex of fruits of Maclura pomifera (Raf.) Schneid. (Moraceae), containing serine endopeptidases. The caseinolytic activity was 14.1 ± 0.8 U cas /mL and the protein concentration was 1.5 ± 0.1 mg/mL. Isoelectrofocusing of pomiferin followed by zymogram analysis showed several bands (pI 3.7–9.2) of which those of pI ≥ 6.0 had a high proteolytic activity. The highest esterase activity was achieved on Ala, Gly and Leu derivatives of N-α-CBZ-p-nitrophenyl amino acid esters. In milk clotting assays pomiferin showed a clotting strength similar to that of chymosin standard (50 IMCU/mL), and clots presented stable consistency over time. A linear dependence was presented between the clotting time and the enzyme dilution. Clotting activity had increased significantly until 25 mmol/L of CaCl 2 content. Wheys obtained using pomiferin and chymosin as clotting agents showed specific peptide profiles on tricine SDS-PAGE gels. Antioxidant activity and ACE-inhibitory activity of filtered whey obtained with pomiferin were 57.4 ± 0.4% and 11 ± 2%, respectively. Both activities could be attributed to small peptides generated by action of proteases on milk proteins during clotting, therefore this whey could be used as ingredient in the manufacture of functional foods.
Vitenko V.a. - One of the best experts on this subject based on the ideXlab platform.
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Підсумки інтродукції Maclura рomifera (Rafin.) Schneid в умовах Правобережного Лісостепу України
M.M. Gryshko National Botanical Garden of the NAS of Ukraine, 2006Co-Authors: Vitenko V.a., Shlapak V.p.Abstract:The results of Maclura pomifera (Rafin.) Schneid introduction in the conditions of the Right-Bank Forest-Steppe zone of Ukraine are given. This species is determined to be perspective for using it in the ornamental horticulture of this region.Наведено результати вивчення біолого-екологічних особливостей інтродукції Maclura pomifera (Rafin.) Schneid в умовах Правобережного Лісостепу України. Встановлено, що вона є перспективною рослиною для використання в декоративному садівництві цього регіону
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Насіннєве розмноження Maclura pomifera (Raf.) Schneid. в умовах Правобережного Лісостепу України
M.M. Gryshko National Botanical Garden of the NAS of Ukraine, 2002Co-Authors: Vitenko V.a.Abstract:Investigations on seed propagation of Maclura pomifera (Raf.) Schneid in conditions of Right-Bank of the Forest-Steppe of Ukraine were done. Duration of germination, medium germination of seed, optimal depth of sowing were determined.Досліджено насіннєве розмноження маклюри в умовах Правобережного Лісостепу України. Встановлено тривалість періоду проростання, середню схожість насіння та оптимальну глибину його посіву
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Оцінка успішності інтродукції Maclura pomifera (Raf.) Schneid. в умовах Правобережного Лісостепу України
M.M. Gryshko National Botanical Garden of the NAS of Ukraine, 2001Co-Authors: Vitenko V.a.Abstract:It was evaluated the effectiveness of the introduction of Maclura pomifera (Raf.) Schneid. in the conditions of Righе-Bank Forest-Steppe. It was established that Maclura is a plant with good prospects for introduction in these conditions, the rate of acclimatization in this region is good, and the terms of acclimatizations are normal.Дана оцінка успішності інтродукції Maclura pomifera (Raf.) Schneid. в умовах Правобережного Лісостепу. Встановлено, що маклюра є перспективною рослиною для інтродукції в цих умовах, ступінь її акліматизації в цьому регіоні добрий, швидкість акліматизації нормальна
G. Delle Monache - One of the best experts on this subject based on the ideXlab platform.
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Maclura pomifera (Osage Orange): In Vitro Culture and the Formation of Flavonoids and Other Secondary Metabolites
Medicinal and Aromatic Plants VIII, 1995Co-Authors: B. Botta, G. Delle MonacheAbstract:Maclura pomifera (Raf.) Schneid. (syn Maclura aurantiaca or Toxylon pomiferum) is the only species of the genus Maclura, belonging to the family Moraceae. The plant is a hardwood tree native to the southwestern United States (Texas and Arkansas), but dispersed also in Europe (Fig. 1A). The tree, which may attain the height of 20 m, has a trunk covered by a furrowed orangecolored bark and thorny branches, surmounted by a long, luxuriant, dishevelled hair.
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Cell Suspension Cultures of Maclura pomifera: Optimization of Growth and Metabolite Production
Journal of Plant Physiology, 1991Co-Authors: Gabriella Pasqua, Barbara Monacelli, A. Cuteri, O. Finocchiaro, Bruno Botta, Alberto Vitali, G. Delle MonacheAbstract:Summary Explants of stem of Maclura pomifera were cultured in media with varied concentrations of hormones. The obtained calli were separated on the basis of colour into two lines, A and B, which gave different metabolite accumulations. The major components of the extracts were A-ring prenylated flavanones, isoflavones and flavones. Cell suspension cultures of lines A and B showed the same difference together with a considerable increase in flavonoid production.
R. L. Rhykerd - One of the best experts on this subject based on the ideXlab platform.
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Impact of selected cultural practices on seedling growth of Osage orange (Maclura pomifera).
Transactions of the Illinois State Academy of Science, 2010Co-Authors: J. D. Rozum, D. E. Kopsell, Gary R. Bachman, B. R. Wiegand, John C. Sedbrook, R. L. RhykerdAbstract:Osage orange (Maclura pomifera [Raf.] C.K. Schneid.) is a perennial tree species whose fruit shows potential as a biofuel energy source. However, limited data exists describing the best management practices for seedling production. Research objectives included determining the effects of varying shade percentages on Osage orange seedling growth in a controlled greenhouse environment and the effect from being intercropped with soybean (Glycine max L. Merr.) in a field situation. In two separate greenhouse trials, Osage orange seedlings were subjected to a no shade control treatment and shading cloth treatments of 30%, 50%, and 70%. In a field experiment, Osage orange seedlings were intercropped and surrounded by two, one, or no rows of soybeans. In both greenhouse trials, height, fresh and dry leaf weight, dry shoot weight, fresh and dry root weight, and leaf area were significant (P
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impact of selected cultural practices on seedling growth of osage orange Maclura pomifera
Transactions of the Illinois State Academy of Science, 2010Co-Authors: J. D. Rozum, D. E. Kopsell, Gary R. Bachman, B. R. Wiegand, John C. Sedbrook, R. L. RhykerdAbstract:Osage orange (Maclura pomifera [Raf.] C.K. Schneid.) is a perennial tree species whose fruit shows potential as a biofuel energy source. However, limited data exists describing the best management practices for seedling production. Research objectives included determining the effects of varying shade percentages on Osage orange seedling growth in a controlled greenhouse environment and the effect from being intercropped with soybean (Glycine max L. Merr.) in a field situation. In two separate greenhouse trials, Osage orange seedlings were subjected to a no shade control treatment and shading cloth treatments of 30%, 50%, and 70%. In a field experiment, Osage orange seedlings were intercropped and surrounded by two, one, or no rows of soybeans. In both greenhouse trials, height, fresh and dry leaf weight, dry shoot weight, fresh and dry root weight, and leaf area were significant (P<0.05). Fresh shoot weight was also significant in the 120 day greenhouse trial. Generally, height, leaf and shoot weights, and leaf area increased under increasing shade treatments. Contrarily, fresh and dry root weights decreased significantly under increasing shade treatments. In the field trial, Osage orange grown independently or surrounded by one soybean row had significantly (P<0.05) higher biomass compared to seedlings intercropped with soybean rows on each side. Although contrary to greenhouse trial results, competition effects such as nutrient and water availability from intercropping in the field, not shading, were likely greater factors in Osage orange seedling biomass. Results show Osage orange can tolerate increased shade without detriment to overall biomass, therefore showing promise for intercropping systems. However, water and nutrient competition should be considered.
Laura M. I. López - One of the best experts on this subject based on the ideXlab platform.
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A Bowman-Birk protease inhibitor purified, cloned, sequenced and characterized from the seeds of Maclura pomifera (Raf.) Schneid.
Planta, 2016Co-Authors: Martín Indarte, Cristian M. Lazza, Diego M. Assis, Néstor O. Caffini, Maria A. Juliano, Francesc X. Avilés, Xavier Daura, Laura M. I. López, Sebastián A. TrejoAbstract:Main conclusion A new BBI-type protease inhibitor with remarkable structural characteristics was purified, cloned, and sequenced from seeds of Maclura pomifera , a dicotyledonous plant belonging to the Moraceae family. In this work, we report a Bowman–Birk inhibitor (BBI) isolated, purified, cloned, and characterized from Maclura pomifera seeds (MpBBI), the first of this type from a species belonging to Moraceae family. MpBBI was purified to homogeneity by RP-HPLC, total RNA was extracted from seeds of M. pomifera, and the 3′RACE-PCR method was applied to obtain the cDNA, which was cloned and sequenced. Peptide mass fingerprinting (PMF) analysis showed correspondence between the in silico-translated protein and MpBBI, confirming that it corresponds to a new plant protease inhibitor. The obtained cDNA encoded a polypeptide of 65 residues and possesses 10 cysteine residues, with molecular mass of 7379.27, pI 6.10, and extinction molar coefficient of 9105 M−1 cm−1. MpBBI inhibits strongly trypsin with K i in the 10−10 M range and was stable in a wide array of pH and extreme temperatures. MpBBI comparative modeling was applied to gain insight into its 3D structure and highlighted some distinguishing features: (1) two non-identical loops, (2) loop 1 (CEEESRC) is completely different from any known BBI, and (3) the amount of disulphide bonds is also different from any reported BBI from dicot plants.
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Milk clotting activity and production of bioactive peptides from whey using Maclura pomifera proteases
LWT - Food Science and Technology, 2012Co-Authors: María Alicia Corrons, Laura M. I. López, Constanza Silvina Liggieri, Juan Ignacio Bertucci, Mariela Anahí BrunoAbstract:Abstract A crude extract named pomiferin was obtained from latex of fruits of Maclura pomifera (Raf.) Schneid. (Moraceae), containing serine endopeptidases. The caseinolytic activity was 14.1 ± 0.8 U cas /mL and the protein concentration was 1.5 ± 0.1 mg/mL. Isoelectrofocusing of pomiferin followed by zymogram analysis showed several bands (pI 3.7–9.2) of which those of pI ≥ 6.0 had a high proteolytic activity. The highest esterase activity was achieved on Ala, Gly and Leu derivatives of N-α-CBZ-p-nitrophenyl amino acid esters. In milk clotting assays pomiferin showed a clotting strength similar to that of chymosin standard (50 IMCU/mL), and clots presented stable consistency over time. A linear dependence was presented between the clotting time and the enzyme dilution. Clotting activity had increased significantly until 25 mmol/L of CaCl 2 content. Wheys obtained using pomiferin and chymosin as clotting agents showed specific peptide profiles on tricine SDS-PAGE gels. Antioxidant activity and ACE-inhibitory activity of filtered whey obtained with pomiferin were 57.4 ± 0.4% and 11 ± 2%, respectively. Both activities could be attributed to small peptides generated by action of proteases on milk proteins during clotting, therefore this whey could be used as ingredient in the manufacture of functional foods.
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Isolation and partial characterization of serine proteinases present in the latex of Maclura pomifera (osage orange) fruits
Acta Alimentaria, 1993Co-Authors: Laura M. I. López, Néstor O. Caffini, Claudia L. Natalucci, E. CurottoAbstract:Latex provided by superficial incisions of Maclura pomifera fruits contains several serine proteinases. Maximal activity of crude preparations is reached at alkaline pH (more than 80% of maximal activity between pH 9.2 and 10.8 on casein and between pH 8.0 and 9.9 on azocoll), but when ionic strength is higher than 0.15 M an abrupt falling of enzyme activity is noted. Thermal stability of crude preparations is a remarkable fact, as enzyme activity is high even after 25 min at 65 °C, but decreases in the case of the main purified proteolytic fraction (III). Acetone fractionation followed by ion-exchange chromatography (DEAE- and CM-Sepharose CL-6B) affords four active fractions with closely related molecular sizes (63-71 kD, SDS-PAGE)
