EFFECT OF HESPERETIN TREATMENT ON BLOOD GLUCOSE LEVEL, SPERMATOZOA QUALITY, AND SPERMATOZOA QUANTITY IN ALLOXAN-INDUCED DIABETIC MICE

Male infertility has occurred rapidly in the last few decades, primarily in developing countries. An antioxidant, hesperetin is a flavonoid that is found in abundance in orange peels. The aims of this research were to determine the effect of hesperetin on blood sugar levels, spermatozoaquality, and spermatozoa quantity. The research structure included induction of diabetes mellitus and treatment for 8 weeks, followed bydetermination of blood sugar levels, spermatozoa quality, and spermatozoa quantity. Hesperetin has the ability to restore blood sugar levels, spermatozoa quantity, seminiferous tubules diameter, and testicular weight, volume, and germinal epithelial layer thickness with significant difference from the normal control group. Hesperetin did not fully restore spermatozoa motility, viability, and morphology with significant difference from the normal control group, nor from the positive control group. However, overall, hesperetin decreased blood glucose levels, increased spermatozoa quantity, and improved the spermatozoa quality in alloxan-induced diabetes mellitus mice. Dose-dependent activity was observed with the optimum dose at 200 mg/kg body weight

Neuroprotective activities of acai berries (Euterpe sp.) against monosodium glutamate induced toxicity in neural cells

Although L-glutamate (L-Glu) is not an essential amino acid, it is a vital excitatory neurotransmitter that plays a critical role in brain function. An excess of L-Glu levels over 1 mM within excitatory synaptic cleft has been linked to neurotoxicity in neurodegenerative diseases (NDDs) and stroke. Monosodium glutamate (MSG) is a highly used food enhancer and food additive worldwide, and excessive consumption of it can cause extreme levels of L-Glu to build up in the cerebrospinal fluid (CSF), leading to neuronal death. The native South American palm berry known as the acai berry (Euterpe sp.) is a potential nutraceutical that contains several bio-active phytochemicals with multi�pharmacological effects. Several studies in recent years have shown that acai berries and their bioactive contents can relieve inflammation, act as antioxidants, prevent carcinogenesis, and protect the nervous system. The first thesis aim was to conduct a systematic review of existing literature to examine how L-Glu affects neuronal viability and its implications for NDDs. A systematic review of 71 studies that met eligibility criteria found that exposure to L-Glu in vitro or in vivo is associated with multiple pathogenic mechanisms that affect neuronal viability, including oxidative stress, antioxidant defence decline, neuroinflammation, neurotransmitter levels dyshomeostasis, aberrant protein aggregation, excitotoxicity, mitochondrial malfunction, calcium level dyshomeostasis, and abnormalities in neuronal histology, cognitive function, and animal behaviour. Thus, identifying and understanding these diverse mechanisms might enable the design of more effective and efficient agents targeting multiple L-Glu-based pathways for NDDs. Moreover, the following research project aimed to evaluate L-Glu (0.137 - 100 mM), acai aqueous and ethanolic extracts (0.001 µg/mL - 1000 µg/mL) toxicity and investigate the neuroprotection effects of acai berry extracts. The evaluation was carried out using various methods, including dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays to assess cell viability, and to assess mitochondrial function the adenosine 5′-triphosphate (ATP) bioluminescent, and VII mitochondrial membrane potential (MMP) measurements as well as oxidative stress measurement using the 2,7-dichlorodihydrofluorescein diacetate (DCFHDA) assay. A whole-cell patch-clamp assay was conducted to investigate the effects of L-Glu and acai berry extracts on NMDARs. This study investigated human neuroblastoma cells SH�SY5Y and differentiated human rhabdomyosarcoma cells TE671. The impact of acai berry extracts and L-Glu toxicity was also investigated in differentiated SH-SY5Y cells and human cortical neural progenitor cells (ReNcell CX) via MTT assay. L-Glu caused a significant reduction in cell viability (p < 0.01 - p < 0.0001), ATP levels (p < 0.05 - p < 0.0001), and MMP level (p < 0.05 - p < 0.0001) and increased ROS production (p < 0.05 - p < 0.0001) in human undifferentiated SH-SY5Y and differentiated TE671 cells. Whole-cell patch-clamp recordings showed that L-Glu and glycine (Gly) administration did not activate currents in SH-SY5Y cells, while activated currents were observed in differentiated TE671 cells. Although acai berry extracts alone had some adverse effects high concentration at 1-1000 µg/mL reduced cell viability, ATP, and MMP level and increased ROS production in undifferentiated SH-SY5Y and differentiated TE671 cells. However, the co-application of acai berry extracts to L-Glu provided neuroprotection against L-Glu with sustained cell viability, decreased LDH production, restored ATP levels, preserved MMP levels and reduced ROS levels. Moreover, the results of this thesis showed novel results that the acai berry aqueous extract significant inhibits (0.001, 100, 1000 µg/mL, p < 0.001, p < 0.0001, and p < 0.0001, respectively) L-Glu + Gly�activated currents in differentiated TE671 cells in a concentration-dependent manner. Furthermore, differentiated SH-SY5Y cells showed no significant reduction in viability after exposure to L-Glu (0.137 - 100 mM), but ReNcell CX cells experienced significant (p < 0.0001) toxicity impacts after exposure to L-Glu at concentration range 0.03 - 100 mM. In ReNcell CX cells, aqueous acai berry extract (0.0001 - 100 µg/mL) co-applied to neurons exhibited significant (p < 0.05 - p < 0.0001) neuroprotection against L-Glu. Thus, this study showed that acai berries contain nutraceuticals with antioxidant and anti�excitotoxicity properties, making them a potentially beneficial dietary component to prevent pathological deficits caused by excessive levels of L-Glu. VIII The third project in this dissertation is designed to evaluate the potential nutraceutical benefits of acai berry extracts in vitro by examining their ability to inhibit cholinesterase enzymes (ChE) and scavenging free radicals by using 2,2-diphenyl-1-picryl-hydrazyl�hydrate (DPPH) and 2,2′-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) assays. Furthermore, several antioxidant effects were assessed, including hydrogen peroxide (H2O2) or hydroxyl radical (OH• ) scavenging, nitric oxide radical (NO• ) scavenging, lipid peroxidation (LPO) inhibition, and ferric ion reduction (Fe+3). Total polyphenols (TPC) and flavonoids (TFC) were determined in both acai extracts. In addition, acai berry extracts were fractionated and analyzed by liquid chromatography mass spectrometry (LC-MS) to identify phytochemicals that may possess anti�cholinesterase and antioxidant properties. Acai berry extracts showed novel action in their ability to inhibit acetyl- and butyryl-cholinesterase with estimated IC50 of 0.001 µg/mL, and 6.378 mg/mL, respectively. Moreover, both acai extracts exhibited effective concentration-dependent antioxidant and free radical scavenging properties. The acai ethanolic extract showed the most potent antioxidant capability and exhibited the highest phenolic and flavonoid contents 101.39 ± 4.61 milligram gallic acid equivalents/gram of acai berry extracts (mg GAE/g) and 11.78 ±1.42 milligram quercetin equivalents/gram of extract (mg QUER E/g), respectively. Fractionation and analysis of acai berry extracts with LC-MS identified several phytochemicals that may have provided antioxidants and anti-cholinesterase effects. Therefore, acai berry extracts could be a potentially dietary supplements that reduce the pathogenic impairments seen in AD. Taken together, the data in this thesis suggest that excessive L-Glu intake has adverse effects, including neuronal tissue degradation, similar pathomechanisms found in NDDs and stroke. Moreover, this thesis reveals that acai berry extracts may include neuroprotective compounds and could be exploited to develop drugs to treat NDDs and strokes, which are linked to excessive buildup and toxicity of L-Glu. In addition, acai extracts may present a novel approach for prospective AD therapy if further fractionated due to their novel anti-cholinesterase and antioxidant properties. However, it is still necessary to validate these findings using human primary cell lines, 3D neuronal models, and in vivo models to explore the influences of L-Glu and acai berry extracts exposure in humans

Neuroprotective activities of acai berries (Euterpe sp.) against monosodium glutamate induced toxicity in neural cells

Although L-glutamate (L-Glu) is not an essential amino acid, it is a vital excitatory neurotransmitter that plays a critical role in brain function. An excess of L-Glu levels over 1 mM within excitatory synaptic cleft has been linked to neurotoxicity in neurodegenerative diseases (NDDs) and stroke. Monosodium glutamate (MSG) is a highly used food enhancer and food additive worldwide, and excessive consumption of it can cause extreme levels of L-Glu to build up in the cerebrospinal fluid (CSF), leading to neuronal death. The native South American palm berry known as the acai berry (Euterpe sp.) is a potential nutraceutical that contains several bio-active phytochemicals with multi�pharmacological effects. Several studies in recent years have shown that acai berries and their bioactive contents can relieve inflammation, act as antioxidants, prevent carcinogenesis, and protect the nervous system. The first thesis aim was to conduct a systematic review of existing literature to examine how L-Glu affects neuronal viability and its implications for NDDs. A systematic review of 71 studies that met eligibility criteria found that exposure to L-Glu in vitro or in vivo is associated with multiple pathogenic mechanisms that affect neuronal viability, including oxidative stress, antioxidant defence decline, neuroinflammation, neurotransmitter levels dyshomeostasis, aberrant protein aggregation, excitotoxicity, mitochondrial malfunction, calcium level dyshomeostasis, and abnormalities in neuronal histology, cognitive function, and animal behaviour. Thus, identifying and understanding these diverse mechanisms might enable the design of more effective and efficient agents targeting multiple L-Glu-based pathways for NDDs. Moreover, the following research project aimed to evaluate L-Glu (0.137 - 100 mM), acai aqueous and ethanolic extracts (0.001 µg/mL - 1000 µg/mL) toxicity and investigate the neuroprotection effects of acai berry extracts. The evaluation was carried out using various methods, including dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays to assess cell viability, and to assess mitochondrial function the adenosine 5′-triphosphate (ATP) bioluminescent, and VII mitochondrial membrane potential (MMP) measurements as well as oxidative stress measurement using the 2,7-dichlorodihydrofluorescein diacetate (DCFHDA) assay. A whole-cell patch-clamp assay was conducted to investigate the effects of L-Glu and acai berry extracts on NMDARs. This study investigated human neuroblastoma cells SH�SY5Y and differentiated human rhabdomyosarcoma cells TE671. The impact of acai berry extracts and L-Glu toxicity was also investigated in differentiated SH-SY5Y cells and human cortical neural progenitor cells (ReNcell CX) via MTT assay. L-Glu caused a significant reduction in cell viability (p < 0.01 - p < 0.0001), ATP levels (p < 0.05 - p < 0.0001), and MMP level (p < 0.05 - p < 0.0001) and increased ROS production (p < 0.05 - p < 0.0001) in human undifferentiated SH-SY5Y and differentiated TE671 cells. Whole-cell patch-clamp recordings showed that L-Glu and glycine (Gly) administration did not activate currents in SH-SY5Y cells, while activated currents were observed in differentiated TE671 cells. Although acai berry extracts alone had some adverse effects high concentration at 1-1000 µg/mL reduced cell viability, ATP, and MMP level and increased ROS production in undifferentiated SH-SY5Y and differentiated TE671 cells. However, the co-application of acai berry extracts to L-Glu provided neuroprotection against L-Glu with sustained cell viability, decreased LDH production, restored ATP levels, preserved MMP levels and reduced ROS levels. Moreover, the results of this thesis showed novel results that the acai berry aqueous extract significant inhibits (0.001, 100, 1000 µg/mL, p < 0.001, p < 0.0001, and p < 0.0001, respectively) L-Glu + Gly�activated currents in differentiated TE671 cells in a concentration-dependent manner. Furthermore, differentiated SH-SY5Y cells showed no significant reduction in viability after exposure to L-Glu (0.137 - 100 mM), but ReNcell CX cells experienced significant (p < 0.0001) toxicity impacts after exposure to L-Glu at concentration range 0.03 - 100 mM. In ReNcell CX cells, aqueous acai berry extract (0.0001 - 100 µg/mL) co-applied to neurons exhibited significant (p < 0.05 - p < 0.0001) neuroprotection against L-Glu. Thus, this study showed that acai berries contain nutraceuticals with antioxidant and anti�excitotoxicity properties, making them a potentially beneficial dietary component to prevent pathological deficits caused by excessive levels of L-Glu. VIII The third project in this dissertation is designed to evaluate the potential nutraceutical benefits of acai berry extracts in vitro by examining their ability to inhibit cholinesterase enzymes (ChE) and scavenging free radicals by using 2,2-diphenyl-1-picryl-hydrazyl�hydrate (DPPH) and 2,2′-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) assays. Furthermore, several antioxidant effects were assessed, including hydrogen peroxide (H2O2) or hydroxyl radical (OH• ) scavenging, nitric oxide radical (NO• ) scavenging, lipid peroxidation (LPO) inhibition, and ferric ion reduction (Fe+3). Total polyphenols (TPC) and flavonoids (TFC) were determined in both acai extracts. In addition, acai berry extracts were fractionated and analyzed by liquid chromatography mass spectrometry (LC-MS) to identify phytochemicals that may possess anti�cholinesterase and antioxidant properties. Acai berry extracts showed novel action in their ability to inhibit acetyl- and butyryl-cholinesterase with estimated IC50 of 0.001 µg/mL, and 6.378 mg/mL, respectively. Moreover, both acai extracts exhibited effective concentration-dependent antioxidant and free radical scavenging properties. The acai ethanolic extract showed the most potent antioxidant capability and exhibited the highest phenolic and flavonoid contents 101.39 ± 4.61 milligram gallic acid equivalents/gram of acai berry extracts (mg GAE/g) and 11.78 ±1.42 milligram quercetin equivalents/gram of extract (mg QUER E/g), respectively. Fractionation and analysis of acai berry extracts with LC-MS identified several phytochemicals that may have provided antioxidants and anti-cholinesterase effects. Therefore, acai berry extracts could be a potentially dietary supplements that reduce the pathogenic impairments seen in AD. Taken together, the data in this thesis suggest that excessive L-Glu intake has adverse effects, including neuronal tissue degradation, similar pathomechanisms found in NDDs and stroke. Moreover, this thesis reveals that acai berry extracts may include neuroprotective compounds and could be exploited to develop drugs to treat NDDs and strokes, which are linked to excessive buildup and toxicity of L-Glu. In addition, acai extracts may present a novel approach for prospective AD therapy if further fractionated due to their novel anti-cholinesterase and antioxidant properties. However, it is still necessary to validate these findings using human primary cell lines, 3D neuronal models, and in vivo models to explore the influences of L-Glu and acai berry extracts exposure in humans

The influence of elicitor Cu2+ and Fe 2+ to asiaticoside level of pennywort (Centella asiatica L. urban) callus

Pennywort (Centella asiatica L. Urban is a medicinal plant used in traditional herbal medicine industry in Indonesia. Production of bioactive compounds through tissue culture can be enhanced by elicitation. This study was an experimental study using completely randomized design.The treatment used is various concentration of metal ions Fe2+ on callus subculture medium namely 0, 90, 100, and 110 mM. with 4 replications to obtain 16 trials. Cu2+ ion concentration treatments used consisted of 4 levels: Cu0 = control (0 mM), Cu1=30 mM , Cu2 = 35 mM, Cu3 =40 mM with 4 replications to obtain 16 trials Elicitor Cu2+ with 40μM concentration resulted in the high levels production of secondary metabolites asiaticoside 4.1595 g / 100g. Elicitor Fe2+ with a concentration of 100 mM asiaticoside produce secondary metabolite 3.836 g / 100g. Elicitor Cu2 + produces higher levels asiaticoside compared with Fe2+

Antioxidant Effects of Marigold (Calendula officinalis) Flower Extract on the Oxidative Balance of Bovine Spermatozoa

The aim of our study was evaluation of potential antioxidant effects of marigold (Calendula officinalis) extract and assessment of its in vitro impact on the selected quality parameters of bovine spermatozoa. Marigold is medicinal herb from the family Asteraceae native to southern Europe, and it commonly used in pharmacology and medicine. Its well-known positive properties include antioxidant, antibacterial, antiflammatory, antiviral, antifungal, antihelmintic and wound-healing activities. In this study, the flower extracts were subjected to high performance liquid chromatography (HPLC), which identified especially phenolic acids (rosmarinic and chlorogenic acids) and polyphenols (rutin, kaempferol, resveratrol, quercetin and apigenin). These substances are known for their antioxidant activity and protective effects against oxidative stress. For our experiments, 10 samples of semen from sexually mature Holstein bulls were collected on a single day by using an artificial vagina, diluted in physiological saline solution and exposed to solutions with different concentration of marigold flower extract (75, 150 and 300 µg/mL). Selected quality parameters (motility, mitochondrial activity, production of reactive oxygen species – ROS, protein oxidation and lipid peroxidation – LPO) were analyzed after 0, 2 and 24 hours of in vitro culture. The motility evaluation was performed by using the computer-assisted sperm analysis (CASA) method. This method revealed that 75 and 150 µg/mL extract had positive effects and increased the motility (P<0.01) and mitochondrial activity (P<0.0001) of bovine spermatozoa compared to the control group following 2 and 24 hours. This phenomenon was observed also in case of ROS production, protein oxidation and LPO. Marigold extract concentrations of 75 and 150 µg/mL decreased the levels of ROS, protein oxidation and damage to the membranes caused by LPO compared to the control group (P<0.05; P<0.01) at time 2 and 24 hours. At the same time, 300 µg/mL extract exhibited positive, although less significant, effects compared to 75 and 150 µg/mL extracts. The data acquired from our study confirm that 75 and 150 µg/mL of marigold flower extract have positive effects on the motility and mitochondrial activity of bovine spermatozoa, and decrease ROS generation, LPO and protein oxidation in spermatozoa. Based on our results, the flower extract from marigold could be used for protection against oxidative stress in in vitro cultures of male gametes

Food science sourcebook

2 v. : ill. ; 26 cm2nd edition.Rev. ed. of: Source book for food scientists. c1978"An AVI book."Pt. 1. Terms and descriptions -- pt. 2. Food composition,properties, and general data