In many studies, green tea epigallocatechin-3-gallate (EGCG) has already shown its therapeutic effects in colorectal cancer cells (CRC). However, its mechanism of actions in CRC is poorly elucidated. Hence, this study attempts to elucidate the mechanism of actions of green tea ECGG via iron chelation activity in CRC. In order to investigate this property, HT-29 cell lines (CRC) were treated with EGCG for 24 h, 48 h, and 72 h. From western blot analysis, EGCG had upregulated transferrin receptor (TfR) protein and downregulated Ferritin-H (FtH) protein indicating that iron chelation activity has occurred in CRC. Meanwhile, the molecular docking study demonstrated that EGCG is able to strongly interact the ferritin protein with a high binding affinity (-7.3 kcal/mol) via strong hydrogen bindings to glutamic acid 64 and lysine 71; two moderate hydrogen bindings to asparagine 74 and a hydrophobic interaction to the hydrophobic pocket of lysine 71. The strong interaction predicted between EGCG to ferritin may lead to inhibition of ferritin by EGCG, thus supporting the downregulation of FtH observed in in vitro studies. Molecular docking study of TfR to EGCG cannot be modulated based on the in vitro results. In conclusion, EGCG possesses iron chelator property in CRC and this potential could be further exploited for CRC treatment.
This study was conducted to evaluate the kinetic characteristics of proteolytic activity of proteases on Channa striatus protein fractions. Degree of hydrolysis (DH), amino acid composition and kinetic parameters of sarcoplasmic and myofibrillar proteins were investigated when incubated with proteinase K and thermolysin, separately. After 30 min incubation with proteases, a decrease in DH of sarcoplasmic protein was observed whereas, hydrolysis of myofibrillar protein with proteases took 2 h with an increase in DH. The major amino acids were glutamic acid (16.6%) in thermolysin- myofibrillar hydrolysate followed by aspartic acid (11.1%) in sarcoplasmic protein fraction with no enzyme treatment and lysine (10%) in thermolysin-myofibrillar hydrolysate. The apparent Michaelis constant of proteinase K was lower than thermolysin for both sarcoplasmic and myofibrillar proteins. However, rate of turnover and enzyme efficiency suggested that sarcoplasmic and myofibrillar proteins are suitable substrates for proteinase K and thermolysin hydrolytic reaction, respectively.
A study to quantify the free glutamic acid content of six processed foods, 44 dishes and 26 condiments available in Malaysia was performed using high-performance liquid chromatography with a fluorescence detector (HPLC-FRD). Recovery tests were carried out with spiked samples at levels from 6 to 31 mg g(-1). High recovery in different matrices was achieved ranging from 88% +/- 13% to 102% +/- 5.12%, with an average of 97% +/- 8.92%. Results from the study revealed that the average free glutamic acid content ranged from 0.34 +/- 0.20 to 4.63 +/- 0.41 mg g(-1) in processed foods, while in prepared dishes it was as low as 0.24 +/- 0.15 mg g(-1) in roti canai (puffed bread served with curry or dhal) to 8.16 +/- 1.99 mg g(-1) in dim sum (a small casing of dough, usually filled with minced meat, seafood, and vegetables, either steamed or fried). Relatively, the content of free glutamic acid was found to be higher in condiments at 0.28 +/- 0 mg g(-1) in mayonnaise to 170.90 +/- 6.40 mg g(-1) in chicken stock powder.
In photodynamic therapy (PDT), the low absorptivity of photosensitizers in an aqueous environment reduces singlet oxygen generation efficiency and thereby decreases photosensitizing efficacy in biological conditions. To circumvent this problem, we designed a phthalocyanine-poly-L-glutamic acid conjugate (1-PG) made from a new phthalocyanine (Pc 1) monofunctionalized to allow adequate conjugation to PGA. The resulting 1-PG conjugate retained high absorptivity in the near-infrared (NIR) region at its λmax675nm in an aqueous environment. The 1-PG conjugate demonstrated good singlet oxygen generation efficiency, increased uptake by 4 T1 breast cancer cells via clathrin-mediated endocytosis, and enhanced photocytotoxic efficacy. The conjugate also displayed a high light-dark toxicity ratio, approximately 1.5-fold greater than zinc phthalocyanine at higher concentration (10 μM), an important feature for the reduction of dark toxicity and unwanted side effects. These results suggest that the 1-PG conjugate could be a useful alternative for deep tissue treatment with enhanced anti-cancer (PDT) efficacy.
The habenula, located on the dorsal thalamic surface, is an emotional and reward processing center. As in the mammalian brain, the zebrafish habenula is divided into dorsal (dHb) and ventral (vHb) subdivisions that project to the interpeduncular nucleus and median raphe (MR) respectively. Previously, we have shown that kisspeptin 1 (Kiss1) expressing in the vHb, regulates the serotonin (5-HT) system in the MR. However, the connectivity between the Kiss1 neurons and the 5-HT system remains unknown. To resolve this issue, we generated a specific antibody against zebrafish Kiss1 receptor (Kiss-R1); using this primary antibody we found intense immunohistochemical labeling in the ventro-anterior corner of the MR (vaMR) but not in 5-HT neurons, suggesting the potential involvement of interneurons in 5-HT modulation by Kiss1. Double-fluorescence labeling showed that the majority of habenular Kiss1 neurons are glutamatergic. In the MR region, Kiss1 fibers were mainly seen in close association with glutamatergic neurons and only scarcely within GABAergic and 5-HT neurons. Our findings indicate that the habenular Kiss1 neurons potentially modulate the 5-HT system primarily through glutamatergic neurotransmission via as yet uncharacterized interneurons. The neuropeptide kisspeptin (Kiss1) play a key role in vertebrate reproduction. We have previously shown modulatory role of habenular Kiss1 in the raphe serotonin (5-HT) systems. This study proposed that the habenular Kiss1 neurons modulate the 5-HT system primarily through glutamatergic neurotransmission, which provides an important insight for understanding of the modulation of 5-HT system by the habenula-raphe pathway.
L-Glutamate plays a crucial role in neuronal cell death, which is known to be associated with various neurodegenerative diseases, such as Alzheimer's, Parkinson's, and Huntington's diseases. In this study, we investigated the protective effects of biochanin A, a phytoestrogen compound found mainly in Trifolium pratense, against L-glutamate-induced cytotoxicity in a PC12 cell line. Exposure of the cells to 10 mM L-glutamate was found to significantly increase cell viability loss and apoptosis, whereas pretreatment with various concentrations of biochanin A attenuated the cytotoxic effects of L-glutamate. Specifically, the pretreatment led to not only decreases in the release of lactate dehydrogenase, the number of apoptotic cells, and the activity of caspase-3 but also an increase in the total glutathione level in the L-glutamate-treated PC12 cells. These results indicate that biochanin A may be able to exert neuroprotective effects against L-glutamate-induced cytotoxicity. Furthermore, our findings also imply that biochanin A may act as an antiapoptotic agent in order to perform its protective function.
Traumatic brain injury (TBI) is known to inflict significant morbidity and mortality worldwide. In severe TBI cases, the resulting physical and cognitive impairments incur high management and rehabilitation costs that crucially involve monitoring intracranial pressure (ICP) and improving brain oxygenation. Normobaric Hyperoxia Treatment (NBOT) is a therapeutic strategy to improve brain oxygen metabolism and to decrease ICP by reducing tissue swelling and deactivating toxin. NBOT is administered by increasing the inspired oxygen concentration to 100% in normal atmospheric pressure. Previous studies involving NBOT had explored its effectiveness to salvage the TBI-related cognitive and motor deficits. However, the focus of these studies has frequently been on the cortical lesions despite the known facts that TBI often inflicts tissue damage to the subcortical areas such as the basal ganglia. There are growing evidence to support recent functional theories that implicate a pivotal role of the basal ganglia in regulating normal movements and cognition through dopamine (DA) and glutamate interaction. Thus, tissue damages leading to TBI-related motor and cognitive deficits may involve the different affected brain regions. This minireview attempts to highlight the key processes involved in the pathophysiology of severe TBI and offers insights into the role of NBOT by exploring its potential effects on the cerebral energy metabolism and gene expression patterns of dopamine receptor in a mouse model.
Phoenix dactylifera L (Date palm) is one of the oldest known fruit crops in the world, and
the consumption of date fruits is no longer restricted to the Middle Eastern countries. Date
palm kernels are waste products of date fruit industry which are normally being discarded.
Based on their dietary fiber content; date palm kernels (DPK) have been proposed to be used
as fiber-based food supplement, caffeine free coffee alternative and animal feed ingredient.
Hence, utilization of such waste is highly desirable for the date industry. To accommodate these
benefits, and subsequent to some uses associated with DPK, this study sought to investigate the
biochemical and nutritional values of the Barhi date palm kernels (BDPK) grown in Iraq. The
results show that BDPK is an excellent source of dietary fiber (66.24 g/100g). Glutamic acid
was found to be the predominant amino acid, (0.674 g/100g), followed by Arginine and aspartic
acid (0.437 g/100g and 0.320 g/100g, respectively). Potassium was the most occurring mineral
in BDPK (2.39 g/kg), and the main sugars were sucrose and fructose (0.548 g/100g and 0.249
g/100g, respectively). Gas-liquid chromatography revealed that the main unsaturated fatty acid
(USFA) was oleic acid (40.927 mg/100g), while the main saturated fatty acid (SFA) were lauric
acid (20.270 mg/100g) and myristic acid (12.288 mg/100g). Furthermore, the BDPK depicted
considerable concentrations of vitamins, in which vitamin B5 (40.4 mg/100g) showed the
highest value. The results obtained indicate a strong potential for BDPK to be used in human
nutrition, cosmetics, and pharmaceutical applications and may provide an important economic
advantage through increasing the utilization of BDKP while also additive value will be added
to the residue.
Fish protein hydrolysate was prepared from tilapia muscle using commercial Alcalase enzyme. Optimization of enzymatic hydrolysis process for preparing tilapia muscle protein hydrolysates (TMPH) was performed by employing central composite design (CCD) method of response surface methodology (RSM). O-phtaldialdehyde (OPA) method was employed to calculate the degree of hydrolysis (DH), which is the key parameter for monitoring the reaction of protein hydrolysis. The suggested model equation was proposed based on the effects of pH, temperature, substrate concentration and enzyme concentration on the DH. Optimum enzymatic hydrolysis conditions using Alcalase enzyme were obtained at pH7.5, temperature of 50oC, substrate concentration of 2.5% and enzyme concentration of 4.0%. Under these conditions, the highest value of the DH was achieved at 25.16% after hydrolysing at 120 min. The TMPH was further assessed for their nutritional value with respect to chemical and amino acid compositions. Molecular weight distributions of TMPH were characterized by SDS-PAGE. TMPH contains moderate amount of protein (28.14%) and good nutritive value with respect to the higher total amino acid composition (267.57 mg/g). Glutamic acid, aspartic acid and lysine were the most abundant amino acids present in TMPH with values 42.68, 29.16 and 26.21 mg/g, respectively. Protein hydrolysates from tilapia muscle containing a desirable peptide with low molecular weight which may potentially to be used as functional food products.
Response surface methodology was applied to study the optimization of palm kernel cake protein (PKCP) hexametaphosphate-assisted extraction. The optimum PKCP yield (28.37%) when extracted using 1.50% sodium hexametaphosphate (SHMP) of pH 10, at 50 °C, and the 1:70 (w/v) ratio of cake-to-solvent was significantly (P acid (16.86 g/100 g protein) and followed by arginine (10.78 g/100 g protein). With respect to the 1991 standard of the FAO/WHO for preschool children, PKCP's essential amino acid profile showed deficiencies. Therefore, it can be used as a complementary protein source by supplementing with a tryptophan-rich source, as this was the limiting amino acid.
Dominant strains of lactic acid bacteria (LAB) isolated from honey bees were evaluated for their γ-aminobutyric acid (GABA)-producing ability. Out of 24 strains, strain Taj-Apis362 showed the highest GABA-producing ability (1.76 mM) in MRS broth containing 50 mM initial glutamic acid cultured for 60 h. Effects of fermentation parameters, including initial glutamic acid level, culture temperature, initial pH and incubation time on GABA production were investigated via a single parameter optimization strategy. The optimal fermentation condition for GABA production was modeled using response surface methodology (RSM). The results showed that the culture temperature was the most significant factor for GABA production. The optimum conditions for maximum GABA production by Lactobacillus plantarum Taj-Apis362 were an initial glutamic acid concentration of 497.97 mM, culture temperature of 36 °C, initial pH of 5.31 and incubation time of 60 h, which produced 7.15 mM of GABA. The value is comparable with the predicted value of 7.21 mM.
The anticonvulsive potential of proteins extracted from Orthosiphon stamineus leaves (OSLP) has never been elucidated in zebrafish (Danio rerio). This study thus aims to elucidate the anticonvulsive potential of OSLP in pentylenetetrazol (PTZ)-induced seizure model. Physical changes (seizure score and seizure onset time, behavior, locomotor) and neurotransmitter analysis were elucidated to assess the pharmacological activity. The protective mechanism of OSLP on brain was also studied using mass spectrometry-based label-free proteomic quantification (LFQ) and bioinformatics. OSLP was found to be safe up to 800 µg/kg and pre-treatment with OSLP (800 µg/kg, i.p., 30 min) decreased the frequency of convulsive activities (lower seizure score and prolonged seizure onset time), improved locomotor behaviors (reduced erratic swimming movements and bottom-dwelling habit), and lowered the excitatory neurotransmitter (glutamate). Pre-treatment with OSLP increased protein Complexin 2 (Cplx 2) expression in the zebrafish brain. Cplx2 is an important regulator in the trans-SNARE complex which is required during the vesicle priming phase in the calcium-dependent synaptic vesicle exocytosis. Findings in this study collectively suggests that OSLP could be regulating the release of neurotransmitters via calcium-dependent synaptic vesicle exocytosis mediated by the "Synaptic Vesicle Cycle" pathway. OSLP's anticonvulsive actions could be acting differently from diazepam (DZP) and with that, it might not produce the similar cognitive insults such as DZP.
Panic disorder (PD) being one of the most intensively investigated anxiety disorders is considered a heterogeneous psychiatric disease which has difficulties with early diagnosis. The disorder is recurrent and usually associated with low remission rates and high rates of relapse which may exacerbated social and quality of life, causes unnecessary cost and increased risk for complication and suicide. Current pharmacotherapy for PD are available but these drugs have slow therapeutic onset, several side effects and most patients do not fully respond to these standard pharmacological treatments. Ongoing investigations indicate the need for new and promising agents for the treatment of PD. This article will cover the importance of immediate and proper treatment, the gap in the current management of PD with special emphasis on pharmacotherapy, and evidence regarding the novel anti-panic drugs including the drugs in developments such as metabotropic glutamate (mGlu 2/3) agonist and levetiracetam. Preliminary results suggest the anti-panic properties and the efficacy of duloxetine, reboxetine, mirtazapine, nefazodone, risperidone and inositol as a monotherapy drug. Apart for their effectiveness, the aforementioned compounds were generally well tolerated compared to the standard available pharmacotherapy drugs, indicating their potential therapeutic usefulness for ambivalent and hypervigilance patient. Further strong clinical trials will provide an ample support to these novel compounds as an alternative monotherapy for PD treatment-resistant patient.
Background: This study is the first phylogenetic genotype analysis of Giardia lamblia in Iran. The main objective was to determine genotyping and identify the sub-assemblages of Giardia lamblia isolates involved in the transmission of giardiasis in Fars Province, south of Iran, in 2012.
Methods: Forty G. lamblia isolates were collected from the patient's fecal samples with gastrointestinal discomfort referred to the health centers and hospitals in Shiraz, Fars Province, south of Iran. Purification of G. lamblia cysts from fecal samples and DNA extraction were performed using monolayer of sucrose density gradient and Phenol-Chloroform-Isoamylalcohol (PCI) respectively. Semi-nested PCR and sequence analysis were then performed using the primers (GDHeF, GDHiF, and GDHiR) which amplified a 432-bp fragment of Giardia glutamate dehydrogenase (gdh) gene. Phylogenetic analysis was carried out using a neighbor-joining tree composed of the nucleotide sequences of G. lamblia isolates obtained in this study and the known sequences isolates published in GenBank.
Results: G. lamblia sub-assemblage AII was the most prevalent genotype with 80% of the cases and 20% of the cases belong to sub-assemblage BIII and BIV based on the DNA sequence of the gdh. G. lamblia isolates at Fars Province were widely distributed within assemblage A cluster (sub-assemblage AII) and the remaining isolates were dispersed throughout the assemblage B cluster (sub-assemblage BIII and BIV).
Conclusion: PCR Sequencing and phylogenetic analysis was a proper molecular method for genotyping and discriminating of the of G. lamblia sub-assemblages in fecal samples, using the glutamate dehydrogenase gene that suggests a human contamination origin of giardiasis.
The seeds of Terminalia catappa from Malaysia were analyzed for their physicochemical properties. The following values were obtained: moisture 6.23 ± 0.09 %, ash 3.78 ± 0.04 %, lipid 54.68 ± 0.14 %, protein 17.66 ± 0.13 %, total dietary fibre 9.97 ± 0.08 %, carbohydrate 7.68 ± 0.06 %, reducing sugar 1.36 ± 0.16 %, starch 1.22 ± 0.15 %, caloric value 593.48 ± 0.24 %. Studies were also conducted on amino acid profile and free fatty acid composition of the seed oil. Results revealed that glutamic acid was the major essential amino acid while methionine and lysine were the limiting amino acids. The major saturated fatty acid was palmitic acid, while the main unsaturated fatty acid was oleic acid followed by linoleic acid. In addition, the seed was rich in sucrose and had trace amount of glucose and fructose. Briefly, the seed was high in proteins and lipids which are beneficial to human.
The physicochemical properties of silver catfish frame hydrolysate powder at three different degree of hydrolysis, DH43%, DH 55% and DH 68% were studied. The hydrolysates powder were obtained by hydrolysis using Alcalase®, centrifugation and spray drying of the supernatant. The study found that preparation of these hydrolysates affected the protein, ash and fat content as well as amino acid composition. As for essential amino acids, their values were generally considered as adequate as compared to the suggested essential amino acids profile of FAO/WHO. The results showed that SFHs were rich in lysine and glutamate. Hydrolysate at DH 68% exhibited better peptide solubility and water holding capacity. As degree of hydrolysis increased, emulsifying capacity and foaming capacity of the hydrolysate decreased. It was also found that the lightness in hydrolysate powder decreased with increase in degree of hydrolysis. This study shows that silver catfish frame hydrolysate has good solubility, good foaming properties and light colour profile, thus having high potential as food ingredient.
The exact etiology of Parkinson's disease (PD) remains obscure, although many cellular mechanisms including α-synuclein aggregation, oxidative damage, excessive neuroinflammation, and dopaminergic neuronal apoptosis are implicated in its pathogenesis. There is still no disease-modifying treatment for PD and the gold standard therapy, chronic use of levodopa is usually accompanied by severe side effects, mainly levodopa-induced dyskinesia (LID). Hence, the elucidation of the precise underlying molecular mechanisms is of paramount importance. Fyn is a tyrosine phospho-transferase of the Src family nonreceptor kinases that is highly implicated in immune regulation, cell proliferation and normal brain development. Accumulating preclinical evidence highlights the emerging role of Fyn in key aspects of PD and LID pathogenesis: it may regulate α-synuclein phosphorylation, oxidative stress-induced dopaminergic neuronal death, enhanced neuroinflammation and glutamate excitotoxicity by mediating key signaling pathways, such as BDNF/TrkB, PKCδ, MAPK, AMPK, NF-κB, Nrf2, and NMDAR axes. These findings suggest that therapeutic targeting of Fyn or Fyn-related pathways may represent a novel approach in PD treatment. Saracatinib, a nonselective Fyn inhibitor, has already been tested in clinical trials for Alzheimer's disease, and novel selective Fyn inhibitors are under investigation. In this comprehensive review, we discuss recent evidence on the role of Fyn in the pathogenesis of PD and LID and provide insights on additional Fyn-related molecular mechanisms to be explored in PD and LID pathology that could aid in the development of future Fyn-targeted therapeutic approaches.
The perception of pain caused by inflammation serves as a warning sign to avoid further injury. The generation and transmission of pain impulses involves various pathways and receptors. Cardamonin isolated from Boesenbergia rotunda (L.) Mansf. has been reported to exert antinociceptive effects in thermal and mechanical pain models; however, the precise mechanism has yet to be examined. The present study investigated the possible mechanisms involved in the antinociceptive activity of cardamonin on protein kinase C, N-methyl-d-aspartate (NMDA) and non-NMDA glutamate receptors, l-arginine/cyclic guanosine monophosphate (cGMP) mechanism, as well as the ATP-sensitive potassium (K+) channel. Cardamonin was administered to the animals intra-peritoneally. Present findings showed that cardamonin significantly inhibited pain elicited by intraplantar injection of phorbol 12-myristate 13-acetate (PMA, a protein kinase C activator) with calculated mean ED50 of 2.0 mg/kg (0.9-4.5 mg/kg). The study presented that pre-treatment with MK-801 (NMDA receptor antagonist) and NBQX (non-NMDA receptor antagonist) significantly modulates the antinociceptive activity of cardamonin at 3 mg/kg when tested with glutamate-induced paw licking test. Pre-treatment with l-arginine (a nitric oxide precursor), ODQ (selective inhibitor of soluble guanylyl cyclase) and glibenclamide (ATP-sensitive K+ channel inhibitor) significantly enhanced the antinociception produced by cardamonin. In conclusion, the present findings showed that the antinociceptive activity of cardamonin might involve the modulation of PKC activity, NMDA and non-NMDA glutamate receptors, l-arginine/nitric oxide/cGMP pathway and ATP-sensitive K+ channel.
Oligopeptidase B (OPB) is a member of the prolyl oligopeptidase (POP) family of serine proteases. OPB in trypanosomes is an important virulence factor and potential pharmaceutical target. Characteristic structural features of POP family members include lack of a propeptide and presence of a β-propeller domain (PD), although the role of the β-PD has yet to be fully understood. In this work, residues Glu(172), Glu(490), Glu(524) and Arg(689) in Trypanosoma brucei OPB (Tb OPB), which are predicted to form inter-domain salt bridges, were substituted for Gln and Ala, respectively. These mutants were evaluated in terms of catalytic properties and stability. A negative effect on kcat/Km was obtained following mutation of Glu(172) or Arg(689). In contrast, the E490Q mutant exhibited markedly decreased thermal stability, although this mutation had less effect on catalytic properties compared to the E172Q and R689A mutants. Trypsin digestion showed that the boundary regions between the β-PD and catalytic domains (CDs) of the E490Q mutant are unfolded with heat treatment. These results indicated that Glu(490) in the CD plays a role in stabilization of Tb OPB, whereas Glu(172) in the β-PD is critical for the catalytic activity of Tb OPB.