Multienzyme-Mediated Dual-Channel Magnetic Relaxation Switching Taste Biosensor (D-MRSTB) for Simultaneous Detection of Umami Compounds and Synergistic Enhancement in Food.

Umami substances play a significant role in the evaluation of food quality, and their synergistic enhancement is of great importance in improving and intensifying food flavors and tastes. Current biosensors available for umami detection still confront challenges in simultaneous quantification of multiple umami substances and umami intensities. In this study, an innovative dual-channel magnetic relaxation switching taste biosensor (D-MRSTB) was developed for the quantitative detection of representative umami substances. The multienzyme signal of D-MRSTB specifically catalyzes the umami substances of interest to generate hydrogen peroxide (H2O2), which is then used to oxidate Fe2+ to Fe3+. Such a valence-state transition of paramagnetic ions was utilized as a magnetic relaxation signaling switch to influence the transverse magnetic relaxation time (T2) within the reaction milieu, thus achieving simultaneous detection of monosodium glutamate (MSG) and inosine 5'-monophosphate (IMP). The biosensor showed good linearity (R2 > 0.99) in the concentration range of 50-1000 and 10-1000 µmol/L, with limits of detection (LOD) of 0.61 and 0.09 µmol/L for MSG and IMP, respectively. Furthermore, the biosensor accurately characterized the synergistic effect of the mixed solution of IMP and MSG, where ΔT2 showed a good linear relationship with the equivalent umami concentration (EUC) of the mixed solution (R2 = 0.998). Moreover, the D-MRSTB successfully achieved the quantitative detection of umami compounds in real samples. This sensing technology provides a powerful tool for achieving the detection of synergistic enhancement among umami compounds and demonstrates its potential for application in the food industry.

Preparation of sunflower seed-derived umami protein hydrolysates and their synergistic effect with monosodium glutamate and disodium inosine-5'-monophosphate.

Sunflower seeds are rich in protein and can be an excellent raw material for the production of umami peptides. In this study, sunflower seed meal, which was defatted at a low temperature, was taken as the raw material, and proteins were separated, followed by hydrolyzation for 4 h by flavourzyme® to obtain hydrolysates with strong umami intensity. These hydrolysates were deamidated using glutaminase to increase the umami intensity. The highest umami value of 11.48 was recorded for hydrolysates deamidated for 6 h, and the umami intensity was determined. The umami hydrolysates mixed with 8.92 mmol IMP + 8.02 mmol MSG showed the highest umami value of 25.21. Different concentrations of ethanol were used for further separation of hydrolysates, and the highest umami value of 13.54 was observed for 20% ethanol fraction. The results of this study provide utilization method for sunflower seed meal protein and a theoretical basis for the preparation of umami peptides. PRACTICAL APPLICATION: A large number of sunflower seed meals after oil production are used as feed for livestock and poultry. Sunflower seed meal is rich in protein, and umami amino acid composition in sunflower seed meal is up to 25%-30%, which is potentially an excellent raw material for the production of umami peptides. The umami flavor and synergistic effect of obtained hydrolysates, with MSG and IMP, were analyzed in the present study. We intend to provide a novel way for utilization of protein from sunflower seed meal along with a theoretical basis for the preparation of umami peptides.

The pursuit of new alternative ways to eradicate Helicobacter pylori continues: Detailed characterization of interactions in the adenylosuccinate synthetase active site.

Purine nucleotide synthesis is realised only through the salvage pathway in pathogenic bacterium Helicobacter pylori. Therefore, the enzymes of this pathway, among them also the adenylosuccinate synthetase (AdSS), present potential new drug targets. This paper describes characterization of His6-tagged AdSS from H. pylori. Thorough analysis of 3D-structures of fully ligated AdSS (in a complex with guanosine diphosphate, 6-phosphoryl-inosine monophosphate, hadacidin and Mg2+) and AdSS in a complex with inosine monophosphate (IMP) only, enabled identification of active site interactions crucial for ligand binding and enzyme activity. Combination of experimental and molecular dynamics (MD) simulations data, particularly emphasized the importance of hydrogen bond Arg135-IMP for enzyme dimerization and active site formation. The synergistic effect of substrates (IMP and guanosine triphosphate) binding was suggested by MD simulations. Several flexible elements of the structure (loops) are stabilized by the presence of IMP alone, however loops comprising residues 287-293 and 40-44 occupy different positions in two solved H. pylori AdSS structures. MD simulations discovered the hydrogen bond network that stabilizes the closed conformation of the residues 40-50 loop, only in the presence of IMP. Presented findings provide a solid basis for the design of new AdSS inhibitors as potential drugs against H. pylori.

Taste Characteristics of Various Amino Acid Derivatives.

Amino acids contribute to the taste of foods. Previous studies on the taste of amino acids focused mainly on α-amino acids, and therefore, the taste characteristics of amino acid derivatives remain unclear. In the present study, we targeted 6 different amino acid derivatives, ß-alanine, citrulline, creatine, γ-aminobutyric acid, taurine, and ornithine, and evaluated their taste characteristics in a human sensory study. All tested amino acid derivatives showed multiple taste qualities; no derivatives had only a single taste quality. However, their taste intensities were relatively weak even at high concentrations. Given that the interactions between amino acid derivatives and nucleotide result in taste enhancements, we investigated the effect of inosine 5'-monophosphate (IMP) on the taste characteristics and found that the taste intensity of ornithine increased in the presence of IMP. This finding will be useful for understanding the role of amino acid derivatives as taste substances in daily foods.

Small Molecule CD38 Inhibitors: Synthesis of 8-Amino-N1-inosine 5'-monophosphate, Analogues and Early Structure-Activity Relationship.

Although a monoclonal antibody targeting the multifunctional ectoenzyme CD38 is an FDA-approved drug, few small molecule inhibitors exist for this enzyme that catalyzes inter alia the formation and metabolism of the N1-ribosylated, Ca2+-mobilizing, second messenger cyclic adenosine 5'-diphosphoribose (cADPR). N1-Inosine 5'-monophosphate (N1-IMP) is a fragment directly related to cADPR. 8-Substituted-N1-IMP derivatives, prepared by degradation of cyclic parent compounds, inhibit CD38-mediated cADPR hydrolysis more efficiently than related cyclic analogues, making them attractive for inhibitor development. We report a total synthesis of the N1-IMP scaffold from adenine and a small initial compound series that facilitated early delineation of structure-activity parameters, with analogues evaluated for inhibition of CD38-mediated hydrolysis of cADPR. The 5'-phosphate group proved essential for useful activity, but substitution of this group by a sulfonamide bioisostere was not fruitful. 8-NH2-N1-IMP is the most potent inhibitor (IC50 = 7.6 µM) and importantly HPLC studies showed this ligand to be cleaved at high CD38 concentrations, confirming its access to the CD38 catalytic machinery and demonstrating the potential of our fragment approach.

Condition-dependent adenosine monophosphate decomposition pathways in striated adductor muscle from Japanese scallop (Patinopecten yessoensis).

The purpose of this study was to confirm inosine monophosphate (IMP) generation and to clarify the decomposition pathway of adenosine monophosphate (AMP) by investigating the properties of AMP, IMP, and adenosine (AdR) decomposition enzymes in Japanese scallop (Patinopecten yessoensis). The results showed that IMP accumulated due to AMP decomposition via endogenous enzymes in scallops stored at both 4 °C and 20 °C. The AMP decomposition rate was highest in the supernatant of homogenized scallop adductor muscle, followed by the suspended solution and precipitate, while IMP could not be decomposed in scallop. The results indicated that the activity of AdR deaminase was very high, and this enzyme was involved in an intracellular process in scallop. Moreover, 1 min of heating exerted little influence on the AMP and AdR decomposition rates, while 5 min of heating induced enzyme denaturation. The IMP generation rate increased dramatically in scallop crude enzyme solution containing 5 mM ethylenediaminetetraacetic acid (EDTA). This suggests that the major pathway of AMP decomposition might change with variations in metal ion concentrations in Japanese scallop. PRACTICAL APPLICATION: IMP generation in Japanese scallop (Patinopecten yessoensis) caused by endogenous enzymes was confirmed. IMP is very important for the umami taste (a pleasant savory taste) of aquatic products. As IMP accumulation might be achieved by changing the concentration of divalent metal ions and no IMP 5'-nucleotidase activity was detected in scallop, a suitable process to produce good flavor scallops with high IMP contents might be developed.

Mutation S115T in IMP-Type Metallo-ß-Lactamases Compensates for Decreased Expression Levels Caused by Mutation S119G.

(1) Background: Metallo-ß-lactamases (MBLs) have raised concerns due to their ability to inactivate carbapenems and newer generation cephalosporins and the absence of clinically available MBL inhibitors. Their genes are often transferred horizontally, and the number of MBL variants has grown exponentially, with many newer variants showing enhanced enzyme activity or stability. In this study, we investigated a closely related group of variants from the IMP family that all contain the combination of mutations S115T and S119G relative to IMP-1. (2) Methods: The effects of each individual mutation and their combination in the IMP-1 sequence background in comparison to IMP-1 were investigated. Their ability to confer resistance and their in-cell expression levels were determined. All enzymes were purified, and their secondary structure and thermal stability were determined with circular dichroism. Their Zn(II) content and kinetic constants with a panel of ß-lactam antibiotics were determined. (3) Results: All four enzymes were viable and conferred resistance to all antibiotics tested except aztreonam. However, the single-mutant enzymes were slightly deficient, IMP-1S115T due to decreased enzyme activity and IMP-1-S119G due to decreased thermal stability and expression, while the double mutant did not show these defects. (4) Conclusions: These observations suggest that S119G was acquired due to its increased enzyme activity and S115T to suppress the thermal stability and expression defect introduced by S119G.

Structure of Arabidopsis thaliana N6-methyl-AMP deaminase ADAL with bound GMP and IMP and implications for N6-methyl-AMP recognition and processing.

Arabidopsis thaliana aminohydrolase (AtADAL) has been shown to be involved in the metabolism of N6-methyl-AMP, a proposed intermediate during m6A-modified RNA metabolism, which can be subsequently incorporated into newly synthesized RNA by Pol II. It has been proposed that AtADAL will prevent N6-methyl-AMP reuse and catabolize it to inosine monophosphate (IMP). Here, we have solved the crystal structures of AtADAL in the apo form and in complex with GMP and IMP in the presence of Zn2+. We have identified the substrate-binding pocket of AtADAL and compared it with that for adenosine deaminase (ADA), adenine deaminase (ADE) and AMP deaminase (AMPD) from multiple species. The comparisons reveal that plant ADAL1 may have the potential ability to catalyze different alkyl-group substituted substrates.

Crystal structures of Trypanosoma brucei hypoxanthine - guanine - xanthine phosphoribosyltransferase in complex with IMP, GMP and XMP.

The 6-oxopurine phosphoribosyltransferases (PRTs) are drug targets for the treatment of parasitic diseases. This is due to the fact that parasites are auxotrophic for the 6-oxopurine bases relying on salvage enzymes for the synthesis of their 6-oxopurine nucleoside monophosphates. In Trypanosoma brucei, the parasite that is the aetiological agent for sleeping sickness, there are three 6-oxopurine PRT isoforms. Two are specific for hypoxanthine and guanine, whilst the third, characterized here, uses all three naturally occurring bases with similar efficiency. Here, we have determined crystal structures for TbrHGXPRT in complex with GMP, XMP and IMP to investigate the structural basis for substrate specificity. The results show that Y201 and E208, not commonly observed within the purine binding pocket of 6-oxopurine PRTs, contribute to the versatility of this enzyme. The structures further show that a nearby water can act as an adaptor to facilitate the binding of XMP and GMP. When GMP binds, a water can accept a proton from the 2-amino group but when XMP binds, the equivalent water can donate its proton to the 2-oxo group. However, when IMP is bound, no water molecule is observed at that location. DATABASE: Coordinates and structure factors were submitted to the Protein Data Bank and have accession codes of 6MXB, 6MXC, 6MXD and 6MXG for the TbrHGXPRT.XMP complex, TbrHGXPRT.GMP complex, TbrHGXPRT.IMP complex, and TbrHGPRT.XMP complex, respectively.

Application of a humidity-mediated method to remove residual solvent from crystal lattice.

We previously reported a humidity-mediated method to effectively remove methanol from the crystal lattice of 3',5'-cyclic monophosphate sodium (cAMPNa) methanol trihydrate, converting it to the pentahydrate without changing its inherent orthorhombic packing mode, and preserving its stability. In this paper, we expand this approach to the removal of residual solvents from l-lysine l-glutamate salt and inosine-5'-monophosphate, and contrast the humidity-mediated method with a solvent-mediated method and a conventional drying method. The packing density of the products obtained from the humidity-mediated method were ∼60% higher than those of the products obtained from the solvent-mediated method, and their stability is ∼5-10% higher than those obtained from the solvent-mediated and traditional drying methods. Furthermore, the humidity-mediated method can remove residual methanol more completely. Therefore, the humidity-mediated method can be regarded as a simple and effective route to eliminate residual solvent from crystal lattice for some crystal products, especially residual methanol.

Enterococcus faecium NCIMB 10415 supplementation improves the meat quality and antioxidant capacity of muscle of broilers.

The present study was conducted to investigate the effects of Enterococcus faecium (E. faecium) on the meat quality and antioxidant capacity of muscle in broilers. A total of 600 Arbor Acre broiler chickens (1-day-old, male) were randomly divided into five treatments with six replicates (20 chickens per replicate) for each treatment. The five treatments were the control treatment (CON, basal diet), antibiotic treatment (ANT, basal diet supplemented with 0.1% chlortetracycline) and E. faecium-supplemented treatments (LEF, MEF and HEF, basal diet supplemented with 50, 100 and 200 mg/kg of E. faecium respectively). The experiment lasted 42 days in two periods of 21 days. Results showed that there were no differences in breast meat quality among different treatments (p > 0.05). Compared with the CON and ANT treatments, the yellowness of thigh meat in E. faecium-supplemented treatments was significantly increased (p < 0.05); the shear force of thigh meat in the LEF and MEF treatments was lower than that of the CON treatment (p < 0.05). In addition, the concentration of the inosine monophosphate (IMP) in the breast and thigh meat of the MEF treatment was significantly higher than that of the other treatments (p < 0.05). At 21 days, the total antioxidant capability (T-AOC) level and glutathione peroxidase (GSH-Px) activity of breast meat and superoxide dismutase (SOD) in the thigh meat of the MEF treatment were greatly increased (p < 0.05). At 42 days of age, the catalase (CAT), GSH-Px and T-AOC activities in the breast meat of the MEF treatment were increased (p < 0.05) and the CAT activity of thigh meat in the LEF and MEF treatments was increased (p < 0.05). In conclusion, E. faecium supplementation increased the meat quality of the thigh muscle, increased the IMP content and the activities of CAT, SOD, T-AOC, and GSH-Px of muscle in broilers. Supplementation with 100 mg/kg E. faecium had the greatest effects.

Genomic characterization of IMP and VIM carbapenemase-encoding transferable plasmids of Enterobacteriaceae.

Objectives: IMP and VIM carbapenemases, in association with class 1 integrons, have spread globally among multiple Enterobacteriaceae species. We characterized IMP- and VIM-encoding transferable plasmids of clinical Enterobacteriaceae collected from two global surveillance programmes. Methods: We performed conjugation and transformation experiments for 38 IMP and 89 VIM producers. Plasmids, obtained from transconjugants or transformants, were sequenced with Illumina next-generation sequencing and analysed for replicon types and antimicrobial resistance genes. Results: A total of 41 transconjugants (blaIMP, n = 22; blaVIM, n = 19) and 10 transformants (blaIMP, n = 1; blaVIM, n = 9) were obtained. Broad-host-range IncL/M, IncC and IncN plasmids were associated with blaIMP and blaVIM, contained various integrons and showed inter-species and international distribution. Narrow-host-range IncFII(K) plasmids were limited to Klebsiella pneumoniae with blaIMP-26 from the Philippines, while IncR and IncHI1B-IncFIB(Mar) plasmids were restricted to K. pneumoniae with blaVIM from Greece and Spain. IncA-like hybrid plasmids were detected in Enterobacter xiangfangensis from Italy and K. pneumoniae from Spain. Transferable plasmids were major contributors of antimicrobial resistance genes among Enterobacteriaceae with blaIMP and blaVIM. Conclusions: This is the first report, to the best of our knowledge, describing blaIMP on IncFII(K) plasmids and blaVIM on IncL/M, IncN2, IncHI1B-IncFIB(Mar) and IncX3-IncC-like plasmids and showed that broad-host-range and narrow-host-range plasmids have contributed to the global spread of blaIMP and blaVIM among different species. This study highlights the importance of molecular analysis of plasmids in providing insight into horizontal spread of these carbapenemases.

Thermal behavior of inosine 5'-monophosphate in acidic form and as alkali and alkaline earth salts.

Inosine 5'-monophosphate in acidic form and its lithium, potassium, magnesium, calcium, strontium and barium were prepared from the sodium salt, characterized by elemental analysis and Fourier transform infrared spectroscopy and submitted to thermogravimetry (TG), differential thermal analysis (DTA), differential scanning calorimetry (DSC) and thermogravimetry coupled to infrared spectroscopy (TG-FTIR) of the volatile products evolved during heating. All the salts were hydrated containing from 4 to 7.5 H2O. After dehydration these salts decomposed releasing the nitrogenous base followed by the ribose group, and producing pyrophosphates as final residue. Evolved Gas Analysis (EGA) reveled the release of water, isocyanic acid and hydrocyanic acid during decomposition of the organic moiety. It was observed only water loss up to 200 °C. At temperatures above 200 °C, the nucleotides were unstable and decomposed, implying that foods containing those additives should be processed below this temperature. Finally, a general mechanism for the decomposition of the inosinates was proposed.

Measuring deaminated nucleotide surveillance enzyme ITPA activity with an ATP-releasing nucleotide chimera.

Nucleotide quality surveillance enzymes play important roles in human health, by detecting damaged molecules in the nucleotide pool and deactivating them before they are incorporated into chromosomal DNA or adversely affect metabolism. In particular, deamination of adenine moiety in (deoxy)nucleoside triphosphates, resulting in formation of (d)ITP, can be deleterious, leading to DNA damage, mutagenesis and other harmful cellular effects. The 21.5 kDa human enzyme that mitigates this damage by conversion of (d)ITP to monophosphate, ITPA, has been proposed as a possible therapeutic and diagnostic target for multiple diseases. Measuring the activity of this enzyme is useful both in basic research and in clinical applications involving this pathway, but current methods are nonselective and are not applicable to measurement of the enzyme from cells or tissues. Here, we describe the design and synthesis of an ITPA-specific chimeric dinucleotide (DIAL) that replaces the pyrophosphate leaving group of the native substrate with adenosine triphosphate, enabling sensitive detection via luciferase luminescence signaling. The probe is shown to function sensitively and selectively to quantify enzyme activity in vitro, and can be used to measure the activity of ITPA in bacterial, yeast and human cell lysates.

GMP and IMP Are Competitive Inhibitors of CMY-10, an Extended-Spectrum Class C ß-Lactamase.

Nucleotides were effective in inhibiting the class C ß-lactamase CMY-10. IMP was the most potent competitive inhibitor, with a Ki value of 16.2 µM. The crystal structure of CMY-10 complexed with GMP or IMP revealed that nucleotides fit into the R2 subsite of the active site with a unique vertical binding mode where the phosphate group at one terminus is deeply bound in the subsite and the base at the other terminus faces the solvent.

Substrate and Cofactor Dynamics on Guanosine Monophosphate Reductase Probed by High Resolution Field Cycling 31P NMR Relaxometry.

Guanosine-5'-monophosphate reductase (GMPR) catalyzes the reduction of GMP to IMP and ammonia with concomitant oxidation of NADPH. Here we investigated the structure and dynamics of enzyme-bound substrates and cofactors by measuring 31P relaxation rates over a large magnetic field range using high resolution field cycling NMR relaxometry. Surprisingly, these experiments reveal differences in the low field relaxation profiles for the monophosphate of GMP compared with IMP in their respective NADP+ complexes. These complexes undergo partial reactions that mimic different steps in the overall catalytic cycle. The relaxation profiles indicate that the substrate monophosphates have distinct interactions in E·IMP·NADP+ and E·GMP·NADP+ complexes. These findings were not anticipated by x-ray crystal structures, which show identical interactions for the monophosphates of GMP and IMP in several inert complexes. In addition, the motion of the cofactor is enhanced in the E·GMP·NADP+ complex. Last, the motions of the substrate and cofactor are coordinately regulated; the cofactor has faster local motions than GMP in the deamination complex but is more constrained than IMP in that complex, leading to hydride transfer. These results show that field cycling can be used to investigate the dynamics of protein-bound ligands and provide new insights into how portions of the substrate remote from the site of chemical transformation promote catalysis.

Fully automatic input function determination program for simple noninvasive (123)I-IMP microsphere cerebral blood flow quantification method.

We recently developed a simple noninvasive (123)I-IMP microsphere (SIMS) method using chest dynamic planar images and brain single photon emission computed tomography. The SIMS method is an automatic analysis method, except for the process of setting the region of interest (ROI) of the input function. If a fully automatic ROI setting algorithm can be developed to determine the input function for the SIMS method, repeatability and reproducibility of the analysis of regional cerebral blood flow (rCBF) of the SIMS method can be guaranteed. The purpose of this study is to develop a fully automatic input function determination program for the SIMS method and to confirm the clinical usefulness of this program. The automatic input function determination program consists of two ROI setting programs for the PA and lung regions, and it is developed using the image phase analysis of a chest RI angiogram. To confirm the clinical usefulness of this program, the rCBF in 34 patients measured using the automatic method were compared with the values obtained through the manual setting method. Input functions by the automatic and manual methods were approximately equal. A good correlation was observed between the rCBF values obtained by the automatic method and those obtained by the manual setting method (r=0.96, p<0.01). Further, the total time taken for the automatic SIMS analysis is 1-2min as compared to 20-30min for the current analysis, and therefore, this technique contributes to the improvement of the throughput of nuclear medical examinations.

Role of W181 in modulating kinetic properties of Plasmodium falciparum hypoxanthine guanine xanthine phosphoribosyltransferase.

Hypoxanthine-guanine-xanthine phosphoribosyltransference (HGXPRT), a key enzyme in the purine salvage pathway of the malarial parasite, Plasmodium falciparum (Pf), catalyses the conversion of hypoxanthine, guanine, and xanthine to their corresponding mononucleotides; IMP, GMP, and XMP, respectively. Out of the five active site loops (I, II, III, III', and IV) in PfHGXPRT, loop III' facilitates the closure of the hood over the core domain which is the penultimate step during enzymatic catalysis. PfHGXPRT mutants were constructed wherein Trp 181 in loop III' was substituted with Ser, Thr, Tyr, and Phe. The mutants (W181S, W181Y and W181F), when examined for xanthine phosphoribosylation activity, showed an increase in Km for PRPP by 2.1-3.4 fold under unactivated condition and a decrease in catalytic efficiency by more than 5-fold under activated condition as compared to that of the wild-type enzyme. The W181T mutant showed 10-fold reduced xanthine phosphoribosylation activity. Furthermore, molecular dynamics simulations of WT and in silico W181S/Y/F/T PfHGXPRT mutants bound to IMP.PPi.Mg2+ have been carried out to address the effect of the mutation of W181 on the overall dynamics of the systems and identify local changes in loop III'. Dynamic cross-correlation analyses show a communication between loop III' and the substrate binding site. Differential cross-correlation maps indicate altered communication among different regions in the mutants. Changes in the local contacts and hydrogen bonding between residue 181 with the nearby residues cause altered substrate affinity and catalytic efficiency of the mutant enzymes. Proteins 2016; 84:1658-1669. © 2016 Wiley Periodicals, Inc.

Overproduction, Purification and Characterization of Adenylate Deaminase from Aspergillus oryzae.

Adenylate deaminase (AMPD, EC 3.5.4.6) is an aminohydrolase that widely used in the food and medicine industries. In this study, the gene encoding Aspergillus oryzae AMPD was cloned and expressed in Escherichia coli. Induction with 0.75 mM isopropyl ß-D-l-thiogalactopyranoside resulted in an enzyme activity of 1773.9 U/mL. Recombinant AMPD was purified to electrophoretic homogeneity using nickel affinity chromatography, and its molecular weight was calculated as 78.6 kDa. Purified AMPD exhibited maximal activity at 35 °C, pH 6.0 and 30 mM K+, with apparent K m and V max values of 2.7 × 10-4 M and 77.5 µmol/mg/min under these conditions. HPLC revealed that recombinant AMPD could effectively catalyse the synthesis of inosine-5'-monophosphate (IMP) with minimal by-products, indicating high specificity and suggesting that it could prove useful for IMP production.

A novel multi-biofunctional protein from brown rice hydrolysed by endo/endo-exoproteases.

Brown rice, which is a less allergenic food grain and contains essential amino acids, was hydrolysed by bromelain and PROTEASE FP51® to improve its functionalities and taste for food applications. The hydrolysate prepared by bromelain (eb-RPH) had high protein solubility, surface hydrophobicity, low molecular weight peptides, hydrophobic amino acids (leucine, valine and glycine) and flavor amino acids (glutamic acid and aspartic acid). The eb-RPH exhibited higher 1,1-diphenyl-2-picrylhydrazyl (DPPH˙) and 2,2'-azino-bis 3-ethylbenzthiazoline-6-sulfonic (ABTS˙(+)) radical-scavenging activities of 76.62% and 52.96%, respectively, and possessed a better foaming capacity (221.76%) and emulsifying capacity (32.34%) than the hydrolysate prepared by PROTEASE FP51® (ep-RPH) did. The eb-RPH gave the desired taste, which is attributed to volatile flavor compounds (benzaldehyde, benzeneacetaldehyde and 2-acetyl-1-pyrroline) and non-volatile flavor compounds, such as monosodium glutamate, 5'-guanosine monophosphate and 5'-inosine monophosphate (0.07, 0.03 and 0.05 mg mL(-1), respectively). Brown rice peptides generated by bromelain were novel bioactive peptides with multifunctional properties.