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Antioxidant activities of Ptychopetalum olacoides (“muirapuama”) in mice brain
Abstract
Ptychopetalum olacoides (PO) roots are used by Amazonian peoples to prepare traditional remedies for treating various central nervous system conditions in which free radicals are likely to be implicated. Following the identification of PO ethanol extract (POEE) free-radical scavenging properties in vitro, the aim of this study was to verify the in vivo antioxidant effect of POEE. Aging mice (14 months) were treated (i.p.) with saline, DMSO (20%) or POEE (100mg/kg body wt.), and the hippocampi, cerebral cortex, striata, hypothalamus and cerebellum dissected out 60 min later to measure antioxidant enzyme activities, free-radical production and damage to macromolecules. POEE administration reduced free-radical production in the hypothalamus, lead to significant decrease in lipid peroxidation in the cerebral cortex, striatum and hypothalamus, as well as in the carbonyl content in cerebellum and striatum. In terms of antioxidant enzymes, catalase activity was increased in the cortex, striatum, cerebellum and hippocampus, while glutathione peroxidase activity was increased in the hippocampus. This study suggests that POEE contains compounds able to improve the cellular antioxidant network efficacy in the brain, ultimately reducing the damage caused by oxidative stress.
... In this study, we also observed that these behavioural deficits were considerably reversed with Muira puama. Several studies have reported the ability of MP to ameliorate central nervous system disorders like anxiety, memory loss, depression and neuronal injury [22,23,24]. These effects have been linked to its ability to inhibit acetylcholinesterase, enhance antioxidant capacity, and modulate neurotransmitters [22,23,24]. ...
... Several studies have reported the ability of MP to ameliorate central nervous system disorders like anxiety, memory loss, depression and neuronal injury [22,23,24]. These effects have been linked to its ability to inhibit acetylcholinesterase, enhance antioxidant capacity, and modulate neurotransmitters [22,23,24]. The antioxidant properties of Muira puama which were also observed in this study have been previously reported. ...
... Muira puama was also observed to have reversed some of the neurodegenerative changes due to AlCl₃. The neuroprotective effects of MP have been attributed to its potent antioxidant and anti-inflammatory potential as well as its ability to modulate brain neurotransmitters [24,32]. ...
Muira puama is a plant renowned for its potent antioxidant and neuroprotective properties. The effects of ethanol extract of Muira puama on aluminium chloride-induced changes in rat behaviour and cerebral cortex histomorphology was examined in this study. Fifty male rats weighing 120-150 g each were divided into six groups: control (normal saline), Muira puama extract groups (25 mg/kg and 50 mg/kg), AlCl3 group (100 mg/kg), and AlCl3 combined with Muira puama extract groups (25 mg/kg and 50 mg/kg respectively). All treatments were administered orally for 21 days. Behaviours were assessed after the final dose of treatment. The result showed that the administration of AlCl3 was associated with decreased weight gain, behavioural alterations, and spatial working memory deficits. However, co-treatment with Muira puama was associated with increased food intake, reversal of weight loss, enhanced locomotion/self-grooming, and reduction of anxiety. Additionally, spatial working memory scores were significantly improved with Muira puama. Biochemical analysis showed reduced levels of malondialdehyde and tumour necrosis factor-alpha, and increased levels of interleukin 10 and total antioxidant capacity. Histological examination revealed neuronal preservation with Muira puama, suggesting protective effects on the cerebral cortex. These findings suggest that Muira puama extract has the ability to mitigate aluminium chloride-induced changes in rats. However, more studies would be needed to determine its suitability for use in humans.
... Additionally, we have recently reported marked free radicals scavenging activity of PO ethanol extract (POEE) in several in vitro assays (Siqueira et al., 2002), as well as a significant inhibition of acethylcholinesterase in different brain regions (Siqueira et al., 2003). Relevant to this study, it was also found that the acute (ip) administration of POEE in mice leads to significant reduction on free radical generation, lipid peroxidation and protein-bound carbonyl content, and increased activities of the antioxidant enzymes catalase and glutathione peroxidase, pertinent indexes of oxidative status in diverse brain structures (Siqueira et al., 2004). ...
... It has been shown that catalases (both, mitochondrial and cytosolic) enhance respiration through complexes I and II (Rodriguez et al., 2000). We here observed that POEE seems to increase respiratory chain activity, which might be related to POEE-induced increase in brain catalase activity (Siqueira et al., 2004). Furthermore, the superoxide anion scavenging porperties of POEE (Siqueira et al., 2002) is of relevance, since mitochondria complexes I and II and aconitase, iron-sulfur cluster-containing enzymes, may suffer free radical inactivation (Melov et al., 1999). ...
... Moreover, POEE peroxyl scavenger action can be regarded as a ''chain-breaking antioxidant'' property useful in protecting membranes lipids (Siqueira et al., 2002). It is important to note that the administration of POEE to mice leads to significant changes on indexes of oxidative status in brain structures, including increases in catalase and glutathione peroxidase activities and reduction of free radical levels, lipid peroxidation and protein-bound carbonyl content (Siqueira et al., 2004). Consistently, by protecting submitochondrial particles from free radical attack POEE facilitates the maintenance of mitochondria respiratory processes under ischemic conditions. ...
... The formula is usually drunk daily before meals, an usual dose roughly equivalent to 60 ml of a ''garrafada''. We have shown that a standardized ethanol extract of P. olacoides (POEE) is promnesic (da Silva et al. 2004) counteracts several types of amnesia (da Silva et al. 2008), and has neuroprotective (Siqueira et al. 2004), antioxidant (Siqueira et al. 2007), and antidepressant properties (Piato et al. 2008Piato et al. , 2009). Given the traditional uses of PO, the antidepressant effects identified for POEE, and the relationship between stress and depression (Sapolsky et al. 2000), the present study aimed to evaluate whether POEE counteracts stress-induced effects. ...
... Ground P. olacoides roots (2.5 kg) were extracted with ethanol (12 l) in Soxhlet (40 h), and evaporated under reduced pressure to yield POEE (6 g of extract per 100 g of drug). The HPLC of the extract used in this study was previously published at Siqueira et al. (2007); the HPLC analysis was conducted using HP 1100 system equipped with photodiode array detector [Agilent Technologies]; the extract was analyzed on a Zorbax extended C18 column [250 Â 4.6 mm] with the MeOH-H2O gradient [10:90–100:0]; solvents were used in HPLC grade with high purity. The flow rate was 1 ml/min; the UV traces were measured at 210 and 254 nm and UV spectra were recorded between 200 and 500 nm. ...
... When mice are exposed to a hypoxic environment, brain noradrenaline is significantly decreased (Georgiev et al. 1995), and a state of oxidative stress is established (Morin et al. 2001 ). POEEinduced resistance to hypoxia may therefore be at least partially explained by its capacity to potentiate noradrenergic activity by beta receptors (Siqueira et al. 1998; Piato et al. 2008), combined with its antioxidant properties (Siqueira et al. 2007). The data here reported are consistent with the neuroprotective effects of POEE found with hippocampal slices submitted to oxygen and glucose deprivation (Siqueira et al. 2004). ...
... Coherent with users' claims, this research group has established promnesic (da Silva et al. 2004 Silva et al. , 2008), anti-amnesic (da Silva et al. 2009) neuroprotective (Siqueira et al. 2004) and antidepressant (Piato et al. 2008 ) properties for a standardized ethanol extract (POEE) of Marapuama. As neurochemical correlates of these properties, antioxidant (Siqueira et al. 2007) and anticholinesterasic effects (Siqueira et al. 2003) have also been identified. A huge impediment to the development of drugs for the treatment of CNS diseases is the blood–brain barrier (BBB) (Pardridge 2009), and the extent to which a drug can readily penetrate the BBB determines its bioavailability within the CNS (Anekonda and Reddy 2005). ...
... Goeldi Museum, identified by Nelson Rosa), in accordance with national guidelines as they relate to the UN Convention on Biodiversity. POEE preparation and its HPLC (HP 1100/photodiode array detector , Agilent Technologies, with a Zorbax extended C18 column [250 mm × 4.6 mm, MeOH–H 2 O gradient 10:90–100:0] ) fingerprinting are detailed at Siqueira et al. (2007). Due to patent PI0307647-4 INPI/Br and US61/297,442 issues, the nature of active compounds and/or detailed extract composition cannot be currently disclosed. ...
... The data in line with those arguing on the advantages in developing anticholinesterase compounds with region-and isoform-specific activity to improve on currently available treatments for cognitive deficits in general, and AD in particular (Zhao and Tang 2002). Several aspects of POEE effects suggest its potential in the treatment of neurodegenerative disorders associated with cholinergic deficits, including: the ability to reverse amnesias (by ageing, scopolamine or MK-801) (da Silva et al. 2004da Silva et al. , 2009 ), the characteristics of its anticholinesterasic properties (selectivity for central cholinesterases, isoform selectivity and an apparent regional selectivity for brain areas most salient for cognition), as well as marked antioxidant (Siqueira et al. 2007) and neuroprotective (Siqueira et al. 2004) properties. Given the huge impediment in drug access to the CNS, the body of experimental evidence generated with this extract, combined with the agreeing claims by native users, reinforces the need to fully characterize the safety and efficacy of extract and/or its isolated compounds by adequate clinical assess- ments. ...
The goal of acetylcholinesterase inhibitors (AChEIs) used to treat Alzheimer's patients is an improvement in cholinergic transmission. While currently available AChEIs have limited success, a huge impediment to the development of newer ones is access to the relevant brain areas. Promnesic, anti-amnesic and AChEI properties were identified in a standardized ethanol extract from Ptychopetalum olacoides (POEE), a medicinal plant favored by the elderly in Amazon communities. The purpose of this study was to provide conclusive evidence that orally given POEE induces AChE inhibition in brain areas relevant to cognition. Histochemistry experiments confirmed that the anticholinesterase compound(s) present in POEE are orally bioavailable, inducing meaningful AChE inhibition in the hippocampus CA1 (∼33%) and CA3 (∼20%), and striatum (∼17%). Ellman's colorimetric analysis revealed that G1 and G4 AChE isoforms activities were markedly inhibited (66 and 72%, respectively) in hippocampus and frontal cortex (50 and 63%, respectively), while G4 appeared to be selectively inhibited (72%) in the striatum. Western blotting showed that POEE did not induce significant changes in the AChE immunocontent suggesting that its synthesis is not extensively modified. This study provides definitive proof of meaningful anticholinesterase activity compatible with the observed promnesic and anti-amnesic effects of POEE in mice, reaffirming the potential of this extract for treating neurodegenerative conditions where a hypofunctioning cholinergic neurotransmission is prominent. Adequate assessment of the safety and efficacy of this extract and/or its isolated active compound(s) are warranted.
... The formula is usually drunk daily before meals, an usual dose roughly equivalent to 60 ml of a ''garrafada''. We have shown that a standardized ethanol extract of P. olacoides (POEE) is promnesic (da Silva et al. 2004) counteracts several types of amnesia (da Silva et al. 2008), and has neuroprotective (Siqueira et al. 2004), antioxidant (Siqueira et al. 2007), and antidepressant properties (Piato et al. 2008Piato et al. , 2009). Given the traditional uses of PO, the antidepressant effects identified for POEE, and the relationship between stress and depression (Sapolsky et al. 2000), the present study aimed to evaluate whether POEE counteracts stress-induced effects. ...
... Ground P. olacoides roots (2.5 kg) were extracted with ethanol (12 l) in Soxhlet (40 h), and evaporated under reduced pressure to yield POEE (6 g of extract per 100 g of drug). The HPLC of the extract used in this study was previously published at Siqueira et al. (2007); the HPLC analysis was conducted using HP 1100 system equipped with photodiode array detector [Agilent Technologies]; the extract was analyzed on a Zorbax extended C18 column [250 Â 4.6 mm] with the MeOH-H2O gradient [10:90–100:0]; solvents were used in HPLC grade with high purity. The flow rate was 1 ml/min; the UV traces were measured at 210 and 254 nm and UV spectra were recorded between 200 and 500 nm. ...
... When mice are exposed to a hypoxic environment, brain noradrenaline is significantly decreased (Georgiev et al. 1995), and a state of oxidative stress is established (Morin et al. 2001 ). POEEinduced resistance to hypoxia may therefore be at least partially explained by its capacity to potentiate noradrenergic activity by beta receptors (Siqueira et al. 1998; Piato et al. 2008), combined with its antioxidant properties (Siqueira et al. 2007). The data here reported are consistent with the neuroprotective effects of POEE found with hippocampal slices submitted to oxygen and glucose deprivation (Siqueira et al. 2004). ...
With the recognition that high levels of sustained stress are associated with the natural course of countless illnesses, effective anti-stress agents have gained importance. Improved endurance to particularly stressful periods is one of the medicinal claims for Marapuama (Ptychopetalum olacoides Bentham, PO), a popular Amazonian herbal. The purpose of this study was to evaluate if PO possesses anti-stress properties. To this end, an extract from PO (POEE) was evaluated on anxiety and glucose levels in mice submitted to the unpredictable chronic mild stress (UCMS) paradigm. POEE did not present anxiolytic effects, but was able to prevent (p<0.01) the UCMS-induced anxiety as assessed by the light/dark test (time spent in the lit area, POEE 100 and 300mg/kg 235.9+/-20.6s and 250.4+/-17.4s, respectively, compared to DMSO 104.7+/-24.4s). Likewise, although POEE did not induce noticeable effects on glycemia, it effectively (p<0.01) prevented the UCMS-induced hyperglycemia (POEE 100 and 300mg/kg 106.4+/-6.7mg/dl and 107.3+/-3.3mg/dl, respectively, compared to DMSO 134.6+/-5.9mg/dl). Additionally, POEE (50-200mg/kg i.p. and 800mg/kg p.o.) significantly (p<0.01 and p<0.05, respectively) increased the time to hypoxia-induced convulsion (by 38%, 51%, 59% and 27%, respectively for i.p. and p.o. treatments). The data indicate that POEE counteracts some of the effects brought about by chronic stress. This study combined with the identified antioxidant and neuroprotective properties, as well as the claimed benefits associated with stressful periods suggest that Ptychopetalum olacoides (Marapuama) might possess adaptogen-like properties.
... Alcoholic infusions of Ptychopetalum olacoides Bentham (PO, Olacaceae), known as " Marapuama, " are consumed in the Amazon region for the treatment of central nervous system (CNS) conditions, and/or during particularly stressful periods (Elisabetsky and Siqueira 1998). We reported that P. olacoides standardized ethanol extract (POEE), a standardized ethanol extract obtained from P. olacoides, possesses various CNS-relevant properties, including antioxidant (Siqueira et al. 2007) and neuroprotective (Siqueira et al. 2004) activities. Regarding cognition, we showed that POEE facilitates short-term and long-term memories (inhibitory avoidance and object recognition) in adult and aged mice (da Silva et al. 2004). ...
... Regarding cognition, we showed that POEE facilitates short-term and long-term memories (inhibitory avoidance and object recognition) in adult and aged mice (da Silva et al. 2004). As a likely neurochemical correlate of POEE promnesic effects, we also reported in vitro and in vivo acetylcholinesterase inhibition for the same extract (Siqueira et al. 2003Siqueira et al. , 2007). The purpose of this study was to investigate if the POEE promnesic effects can offset scop-and MK-801-induced amnesias in mice. ...
... Preparation of Extract Roots of P. olacoides Bentham (Olacaceae) were collected in Pará (Brazil) and identified by Nelson Rosa (MPEG 108.036 voucher at the Goeldi Museum Herbarium). P. olacoides standardized ethanol extract (POEE) was prepared and characterized as detailed elsewhere (Elisabetsky and Siqueira 1998 ); analytical highperformance liquid chromatography fingerprinting can be seen in Siqueira et al. (2007). Drugs Dimethyl sulfoxide (DMSO) was acquired from Delaware (Brazil). ...
Traditional remedies prepared from Ptychopetalum olacoides (PO) are used throughout the Amazon to alleviate age-related conditions. These formulas are mainly used by elders, and alleged effects may be related to the anticholinesterase properties identified in a standardized ethanol extract of this species [P. olacoides standardized ethanol extract (POEE)].
To further characterize the potential of this extract for developing drugs useful to treat cognitive deficits, the effects of POEE on scopolamine (scop)- and MK801-induced amnesias (acquisition, consolidation, and retrieval) in mice were investigated.
Scop (3.0 mg/kg, ip) significantly impaired memory (all three phases) in the step-down inhibitory avoidance protocol. As expected, MK801 (0.1 mg/kg, ip) was amnesic regarding acquisition and consolidation, but not retrieval. POEE (100 mg/kg, ip) reversed the scop-induced impairment in all three phases of long-term and short memories, whereas only the memory consolidation deficit was reversed with MK801-induced amnesia.
This study complements previously reported promnesic properties of this plant extract and suggests that POEE may be further developed for treating conditions associated with cognitive deficits, especially those linked with cholinergic malfunction.
... Additionally, we have recently reported marked free radicals scavenging activity of PO ethanol extract (POEE) in several in vitro assays (Siqueira et al., 2002), as well as a significant inhibition of acethylcholinesterase in different brain regions (Siqueira et al., 2003). Relevant to this study, it was also found that the acute (ip) administration of POEE in mice leads to significant reduction on free radical generation, lipid peroxidation and protein-bound carbonyl content, and increased activities of the antioxidant enzymes catalase and glutathione peroxidase, pertinent indexes of oxidative status in diverse brain structures (Siqueira et al., 2004). ...
... It has been shown that catalases (both, mitochondrial and cytosolic) enhance respiration through complexes I and II (Rodriguez et al., 2000). We here observed that POEE seems to increase respiratory chain activity, which might be related to POEE-induced increase in brain catalase activity (Siqueira et al., 2004). Furthermore, the superoxide anion scavenging porperties of POEE (Siqueira et al., 2002) is of relevance, since mitochondria complexes I and II and aconitase, iron-sulfur cluster-containing enzymes, may suffer free radical inactivation (Melov et al., 1999). ...
... Despite the understanding of molecular mechanisms involved with the proposed neuroprotective action of POEE is incomplete, we suggest that its antioxidant properties (Siqueira et al., 2002;Siqueira et al., 2004) plays a significant role given that POEE reduced OGD-induced increase in the free radical content. The detailed mechanism by which POEE reduced OGD-induced free radical production remains a subject for further investigation. ...
Alcoholic infusions of Ptychopetalum olacoides Bentham (PO, Olacaceae) are used in traditional medicine by patients presenting age associated symptoms and those recovering from stroke. The aim of this study is to evaluate the neuroprotective properties of PO ethanol extract (POEE) using hippocampal slices from Wistar rats exposed to oxygen and glucose deprivation (OGD, followed by reoxygenation). Mitochondrial activity, an index of cell viability, was assessed by the MTT assay; in addition, the free radicals content was estimated by the use of dichlorofluorescein diacetate as probe. The OGD ischemic condition significantly impaired cellular viability, and increased free radicals generation. In non-OGD slices, incubation with POEE (0.6 microg/ml) increased (approximately 40%) mitochondrial activity, without affecting free radicals levels. In comparison to OGD controls, slices incubated with POEE (0.6 microg/ml) during and after OGD exposure had significantly increased cellular viability. In addition, at this same concentration, POEE prevented the increase of free radicals content induced by OGD. In view of the fact that respiratory chain inhibition and increased generation of free radicals are major consequences of the ischemic injury, this study suggests that Ptychopetalum olacoides contains useful neuroprotective compounds and, therefore, deserves further scrutiny.
... 3 Previous pharmacological studies have shown that the EtOH extract of P olacoides produces a series of beneficial effects on the central nervous system, consisting of neuroprotective, anti-stress, antidepressant, antioxidative, adaptogen-like activities, and inhibitory effects on acetylcholinesterase in mice. [4][5][6][7][8][9][10] As part of our ongoing research for natural products with neurotrophic activities, we have continued to investigate phytochemical constituents of the MeOH extract of the bark of P olacoides, which exhibits neurite outgrowthpromoting activity in NGF-mediated PC12 cells. [11][12][13] Further research of this plant led to the isolation of 4 new clerodane-type diterpenoids, named ptycholide V (1), 7α,20-dihydroxykolavelool (2), ptycholide VI (3), and ptycholide VII (4) (Figure 1). ...
... The extraction procedure has been reported previously. 13 The extract was separated by CC on silica gel using a linear gradient solvent system (100% n-hexane to 100% EtOAc) to give 15 fractions (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15) Screening for Neurite Outgrowth-Promoting Activity PC12 (phenochromocytoma) cells were cultured in a 24-well plate at density of 8 × 10 3 cells/mL in DMEM + 10% HS, 5% FBS, 100 IU/mL penicillin, and 100 μg/mL streptomycin at 37°C under a humidified atmosphere of 95% air and 5% CO 2 for 24 h. The culture medium was then changed to DMEM + 2% HS, 1% FBS, 100 IU/mL penicillin, and 100 μg/mL streptomycin. ...
A phytochemical component investigation of the bark of Ptychopetalum olacoides led to the isolation of 4 new clerodane-type diterpenoids, namely, ptycholide V (1), 7α,20-dihydroxykolavelool (2), ptycholide VI (3), and ptycholide VII (4). Their structures were elucidated by extensive spectroscopic data and comparison of NMR data with that obtained for known compounds.
... Marapuama (Ptychopetalum olacoides) is one of the most popular Amazonian herbs used in the treatment of central nervous system-related ailment and/or during highly stressful periods [116]. The root and bark of the plant are used in South America as a tea, mainly against sexual impotence, debilities, fatigue etc. ...
... Subtantial evidence from the literature indicate that P. olacoides act as a scavenger of nitrogen oxides as well as superoxide generated from the xanthine dehydrogenase/xanthine oxidase (XD/XO) system in vitro. The same authors showed that the administration of P. olacoides (100 mg/kg) significantly decreased free radical production and lipid peroxidation in different brain areas of mice in relation to the control [116,120]. P. olacoides has also been demonstrated to inhibit acethylcholinesterase activity in cognitiverelevant brain region areas of mice [121]. This evidence suggests that the plant can be used for treating neurodegenerative conditions where a hypofunctioning cholinergic neurotransmission is prominent, like Alzheimer disease and perhaps brain I/R [122]. ...
Brain ischemia-reperfusion injury is a complex pathological condition that involves a cascade of events like excitotoxicity, peri-infarct depolarization, oxidative/nitrosative stress, inflammation, apoptosis, necrosis and autophagic degeneration. Since the past decades, researchers have provided significant information about the therapeutic potential and possible mechanism of action of plant extracts and derived chemicals to target multiple pathways of the ischemic cascade. Here, we summarized experimental, clinical and epidemiological therapeutic interventions of plants extracts and phytochemicals against brain ischemia-reperfusion injury. Whether oxidative stress is the cause or consequence of brain ischemia is open for debate but details about oxidative burden and inflammation following brain ischemia are also described. Furthermore, the antioxidant mechanism of these extracts/phytochemicals was reviewed. Although these plant extracts and phytochemicals showed the ability to act on the multiple steps of the ischemic cascade in in vitro and in vivo models of brain ischemia, further investigations are needed for their validation and for the development of new drugs.
... The traditional uses of P. olacoides have been extensively exploited by herbal industries in several American and European countries, and the native herb or its extracts are constituents of various phytomedicines supplied in the form of pills, tinctures, tablets, multivitamin supplements or compound extracts (Silva et al., 2002;Siqueira et al., 2003Siqueira et al., , 2007. The plant is also a component of the medication Catuama®, which is widely used in several parts of Brazil as a general tonic and to alleviate physical and mental fatigue (Antunes et al., 2001). ...
... The plant is also a component of the medication Catuama®, which is widely used in several parts of Brazil as a general tonic and to alleviate physical and mental fatigue (Antunes et al., 2001). The pharmacological properties of such tonics have been elucidated and discussed (Siqueira et al., 2003), while a number of experimental and clinical studies carried out on extracts of P. olacoides have validated their medicinal applications (Drewes et al., 2003;Siqueira et al., 2003Siqueira et al., , 2007Silva et al 2007). In contrast, little is known concerning the phytochemistry of the species. ...
Ptychopetalum olacoides Benth., Olacaceae, popularly known as marapuama or muirapuama or miriantã, is a species native to the Amazonian region of Brazil. Extracts of the bark of the plant have been used traditionally for its stimulating and aphrodisiac properties and currently commercialised by the herbal industry as constituents in a wide range of phytomedicines. Fractionation by open column chromatography followed by preparative HPLC-UV/PAD of the stem bark and of three commercial extracts of P. olacoides allowed the isolation of three components that were common to all extracts analysed, and these were identified by NMR to be vanillic acid, protocatechuic acid and theobromine. Vanillic acid, which has been proposed as a phytochemical marker for P. olacoides, was employed as an external standard in the development and validation of a rapid qualitative and quantitative HPLC assay for the analyte. The recoveries values of the developed method were 99.02% and the LOD and LOQ values were 0.033 and 0.11 mg.L-1, respectively. The described method may be applied to the standardisation of herbs, extracts or phytomedicines commercialised as marapuama.
... A voucher specimen (GM108.036) is deposited at the Goeldi Museum Herbarium. POEE (ethanol extract) preparation and its HPLC (HP 1100/photodiode array detector, Agilent Technologies, with a Zorbax extended C18 column [250 mm × 4.6 mm, MeOH-H 2 O gradient 10:90-100:0]) fingerprinting are detailed at Siqueira et al. (2007). The nature of the active compounds and/or detailed extract composition cannot be disclosed due to patent issues (PI0307647-4, INPI/Br; US61297442). ...
... Unfortunately, due to the importance of using the very same batch of extract with which promnesic (da Silva et al., 2004) and anti-amnesic (da Silva et al., 2004) effects were shown in mouse models less directly linked to AD, complementary experiments (longer treatment and/or wider dose range) were here precluded.Given the multifactorial nature of neurodegeneration in itself (Pimplikar, 2009), and the complex interplay of factors relevant to AD susceptibility and course, the benefits of multi-functional drugs have been advocated (Youdim and Buccafusco, 2005). In this context, POEE, here shown to mitigate A consequences in mice, has also been shown to be an effective brain antioxidant (Siqueira et al., 2007), to increase hippocampal cell viability after hypoxia (Siqueira et al., 2004), and to possess relevant AChE inhibitory properties in cognition-important areas (Siqueira et al., 2003;Figueiró et al., in press). ...
Alzheimer's disease (AD) is expected to affect more than 22 million people worldwide by 2025, causing devastating suffering and enormous costs to families and society. AD is a multifactorial disease, with a complex pathological mosaic. In rodents, AD-like dementia can be induced by cerebral microinjection of Aβ peptide, leading to amyloid deposits, amnesia and various features of neurodegeneration. Marapuama (Ptychopetalum olacoides) is regarded as a "brain tonic" in the Amazon region and shows a nootropic profile in rodents.
Because a specific extract (POEE) of Marapuama was shown to possess promnesic and anti-amnesic properties, the aim of this study was to verify if POEE is also effective against Aβ(1-42)-induced cognitive deficit in mice. Additionally, Aβ deposits (Congo red), GFAP immunoreactivity (immunohistochemistry), and neurodegenerative changes in the hippocampal pyramidal layer (Nissl) were examined as measures of Aβ(1-42)-induced neurodegeneration.
CF1 mice were subjected to the experimental Alzheimer model with the Aβ(1-42) i.c.v. administration. The effects of POEE 800 mg/kg were evaluated over 14 consecutive days of treatment.
The data show that 14 days of oral treatment with POEE (800 mg/kg) was effective in preventing Aβ-induced cognitive impairment, without altering the levels of BDNF and with parallel reductions in Aβ deposits and astrogliosis. CA1 hippocampus loss induced by Aβ(1-42) was also diminished in POEE-treated mice.
This study offers evidence of functional and neuroprotective effects of two weeks treatment with a Ptychopetalum olacoides extract against Aβ peptide-induced neurotoxicity in mice. Given the multifactorial nature of neurodegeneration, the considerable potential for an AChE inhibitor displaying associated neuroprotective properties such as here reported warrants further clinic evaluation.
... The plant material is usually prepared in cachaç a (a distilled spirit obtained from sugar cane) or wine, and drunk daily before meals. Coherently with traditional claims, antidepressant (Piato et al., submitted), neuroprotective (Siqueira et al., 2004), antioxidant (Siqueira et al., 2007), and memory facilitating effects (da Silva et al., 2004) were identified with a standardized PO ethanol extract (POEE). We have established that POEE (intraperitoneally or orally administered to mice) significantly and dose-dependently decreases immobility in the tail suspension and forced swimming tests, apparently through effects on D 1 dopamine and -noradrenergic receptors (Piato et al., submitted). ...
... Ptychopetalum olacoides ethanol extract (POEE) was obtained as follows: dried ground roots (2.5 kg) were extracted with ethanol (12 l) in Soxhlet (40 h), and evaporated under reduced pressure (6% yield). POEE contains saponin, phenolic and terpenic compounds; HPLC fingerprinting was carried out on an HP 1100 system equipped with a photodiode array detector (Siqueira et al., 2003Siqueira et al., , 2007). ...
ETHNOPHARMACOLOGY RELEVANCE: Ptychopetalum olacoides Bentham (PO) (Olacaceae), known as Marapuama, is regarded as a "nerve tonic" in the Amazon. Traditional uses include states of lassitude with noticeable lack of desire/motivation, and to manage particularly stressful (physical and/or psychological) circumstances. Suggestive of antidepressant activity, we have established that a specific PO ethanol extract (POEE) significantly decreases immobility in the tail suspension and forced swimming tests.
The aim of this study was to verify the effects of POEE in the unpredictable chronic mild stress (UCMS) depression model in mice, given the construct and face values of the UCMS as an experimental model of depression and the traditional use of this species.
Over 6 weeks BALB/c mice were subjected to the UCMS protocol. The effects of POEE (50, 100, 300mg/kg, p.o.) and imipramine (20mg/kg, i.p.) were evaluated in relation to coat state, splash-test grooming, and corticosterone levels.
The coat state degradation, decreased grooming and increased serum corticosterone induced by UCMS were prevented by POEE and imipramine treatments.
In addition to supporting traditional claims and previously reported antidepressant properties for POEE, this study shows that POEE prevents stress-induced HPA hyperactivity.
... Marapuama-based remedies are employed for various conditions, including facilitating recovery from stroke, and keeping up with highly stressful psychological and/or physical circumstances [28]. A standardized ethanol extract obtained from P. olacoides (POEE) roots possesses nootropic [29], antioxidant [30,31], and neuroprotective [32] properties. The pharmacodynamic basis of POEE facilitatory effects on diverse memory types [29] includes anticholinesterase actions [33], and the involvement of dopamine D 1 and β adrenergic receptors unpublished data. ...
... The extract was evaporated under reduced pressure resulting in the POEE (150 g, 6% yield). Preliminary phytochemical screening gave positive results for saponin, phenolic and terpenic compounds [31,33]. ...
Nootropic, antioxidant, and neuroprotective properties have been shown in a standardized ethanol extract of Ptychopetalum olacoides (POEE), a medicinal plant traditionally used by the Amazonian elderly population. It has been revealed that POEE mechanisms of action include anticholinesterase effects, and involve beta-adrenergic and dopamine D(1) receptors. The purpose of this study was to verify the role of serotonin receptors in the promnesic effects of this standardized extract. The step-down task in mice and selective serotonin antagonists were used. The study reveals that POEE promnesic effects on short-term (acquisition, consolidation and retrieval) and long-term (retrieval) declarative aversive memories are increased by 5HT(2A) (but not 5HT(1A)) serotonin antagonists (spiperone and pindolol, respectively). The observed synergism between POEE and spiperone can be interpreted as the combined effects of two subeffective doses of two 5HT antagonists, or the known synergism between an acetylcholinesterase inhibitor (POEE) and a 5HT antagonist. In conclusion it is suggested that 5HT(2A) serotonin receptors are relevant for the promnesic effects of this extract, adding to its multiple mechanisms of action.
... Ayres et al., 2015, p. 201; Deng et al., 2010; Figueiredo et al., 2016; Klein et al.Ayres et al., 2015; Barbosa et al., 2008; Coleta et al., 2006; Deng et al., 2010; Li et al., 2011; Otify et al., 2015; Petry et al., 2001) Increased time spent in elevated plus maze open arms (Coleta et al., 2006; Figueiredo et al., 2016; Otify et al., 2015; Petry et al.Coleta et al., 2006) Increased number of transitions and time in the light compartment on light-dark procedure (Sena et al., 2009) Antidepressant Reduced immobility time on forced (Jovelina Samara Ferreira Alves et al.Bittencourt et al., 2014; Boasquívis et al., 2018; Da Silva Bittencourt et al., 2020; Dabulici et al., 2020; Ferrari, 2002; K. N. Machado et al., 2021; Majhenič et al., 2007; Martins, 2010; Mattei et al., 1998; Mingori et al., 2017; Peixoto et al., 2017; Portella et al., 2013; Roggia et al., 2020; Sereia et al., 2019; Veloso et al., 2017; Yamaguti-Sasaki et al., 2007; Zamberlan et al.et al., 2017; Sereia et al., 2019; Zamberlan et al., 2020) Anxiolytic effect dependent on 5-HT/DA/Glu systems (Rangel et al., 2013) Antidepressant effect partially (Campos et al.time in water on forced swim test (Espinola et al., 1997) Reduced methylmercury-induced (Algarve et al.. L. Da Silva et al., 2007; Da Silva et al., 2009, 2008, 2004; Figueiró et al., 2011) Reverted amnesia induced by scopolamine and MK-801 on stepdown inhibitory avoidance (Da Silva et al., 2009) Increased time spent in the light compartment on light-dark procedure in unpredictable chronic mild stress model (Piato et al., 2010)Increased latency on step-down inhibitory avoidance and recognition index after 24h on object recognition test (A. L.Da Silva et al., 2007) ...
Given the difficulty of treating some psychiatric and neurodegenerative conditions, Brazil's rich biodiversity provides an optimal scenario for exploring therapeutic alternatives based on bioactive compounds from natural products. 30 non-scientific books were consulted to identify plants popularly used for any purposes related to feelings of well-being. Native species with four or more citations in different books were considered as most popular species, and underwent a systematic review in PubMed, LILACS and Periódicos CAPES databases, including articles in the area of psychopharmacology published up to December 2023, written in Portuguese, English or Spanish. The major use of the 27 native species identified in the survey was as stimulant or calmative, mainly through aqueous extractions of leaves and barks. The systematic review includes 568 articles on the psychopharmacological effects of 23 of the original 27 native species identified, from which 28 articles corresponds to clinical stage. Although most of the studies reviewed support the popular use attributed to some species, it is noteworthy that the vast majority relate to preliminary studies, especially on in vitro antioxidant activity, and that clinical studies are still very scarce. In this sense, this review may serve not only as a guide to the effects popularly and scientifically attributed to Brazilian flora products, but also to encourage future research and the development of alternative therapies against psychological stress.
... Outros estudos tratam do efeito terapêutico das plantas medicinais a partir do etnoconhecimento para as mais diversas condições de saúde, como ansiedade e depressão, edema, dor de cabeça, Parkinson, tratamento de distúrbios alimentares, miíase, déficits cognitivos em epilepsia e demência, doenças do sistema nervoso e disfunção sexual(Berlowitz et al., 2023; Almeida Junior et al., 2021; Albino et al., 2021;Katchborian-Neto et al., 2020; Renelli et al., 2020;Luzuriaga-Quichimbo, 2020;McKenna et al., 2011;Siqueira et al., 2007; Batista et al., 2023).Os estudos em etnobotânica visam a catalogação das espécies mais utilizadas em tratamentos de saúde em comunidades tradicionais e aldeias indígenas(Sousa et al., 2019; Vasquez-Ocmín et al., 2018; Alves et al., 2017; ...
O objetivo inicial das buscas era identificar estudos que tratassem da medicina tradicional amazônica (MTA) na formação inicial de enfermeiros. A partir disso, foram sendo elaboradas as combinações de palavras-chave que poderiam atender às demandas dessa revisão sistemática de literatura (RSL); no entanto, as diferentes estratégias de buscas utilizadas não resultaram em nenhum achado. Com isso, optou-se por realizar esta RSL com os dados identificados através da palavra-chave “Traditional Amazonian Medicine”. Desse modo, este artigo tem por objetivo conhecer o que está sendo produzido na literatura científica acerca da Medicina Tradicional Amazônica. O tipo de estudo adotado é uma revisão sistemática da literatura, a partir do método Systematic Search Flow (SSF). O portfólio é composto por 77 artigos, que abordam uma variedade de temas, incluindo o uso de plantas medicinais, práticas terapêuticas indígenas, efeitos terapêuticos de substâncias psicoativas como a ayahuasca e tratamentos para diferentes condições de saúde. Além disso, há um interesse crescente na integração da MTA com a medicina alopática, como é o caso de estudos que investigam o uso de substâncias psicoativas em terapias para toxicodependentes e para transtornos mentais. O estado do conhecimento da medicina tradicional amazônica revela que as pesquisas desenvolvidas nessa temática se centram na etnofarmacologia e etnobotânia, fator que pode ser atribuído pelo interesse da indústria farmacêutica em descobrir novos ativos e desenvolver novos medicamentos. Estas pesquisas são importantes também para validar o conhecimento tradicional sobre plantas medicinais. Contudo, esse tipo de estudo não é suficiente para abarcar a complexidade de tratamentos, rituais e técnicas empregadas na MTA, tampouco aos diferentes aspectos culturais que constituem esse saber.
... The antioxidant potential of the ethanolic extract from root barks of P. olacoides was demonstrated against different challenges such as nitric oxide, superoxide, and peroxyl radicals (Siqueira et al. 2002). Siqueira et al. (2007) have shown that acute administration of the extract (100 mg/kg, ip) to 14-month-old mice decreased free radical production and lead to a decrease in lipid peroxidation in important cerebral areas. Moreover, the extract increased the activity of glutathione peroxidase and catalase in the hippocampus, while the catalase activity was also increased in the cortex, striatum, and cerebellum. ...
The Ptychopetalum olacoides Benth. (Olacaceae) is an Amazonian tree popularly known as muirapuama or marapuama, among other names, which is used for several central nervous system related problems. The roots and occasionally the bark roots are the main medicinal parts employed and are prepared as an alcoholic infusion, tinctures, and tea. Phytochemical studies revealed that the roots contain tannins, flavonoids, and several terpenoids, while the presence of alkaloids is not clear. Most studies used ethanolic or hydroalcoholic extracts prepared with the roots of the plant. These studies indicate that the species has promising potential for treating central nervous system disorders, acting as an antidepressant, an anti-stress, a neuroprotective agent, and improving cognition. Although some herbal products contain P. olacoides in their composition, clinical studies are still needed to confirm the effects observed in pre-clinical studies.
... Ptychopetalum uncinatum is native to the Amazon rainforest, regionally known as "marapuama, muiratã, muirapuama, pau-homem and liriosma"" There are hypotheses about its secondary plant metabolism, and data on its phytochemical composition and pharmacological functions are not yet fully elucidated. However, few studies report its use for sexual dysfunction and stimulating action on the central nervous system, attributed to the presence of alkaloids, tannins and essential oils [20,21,22]. ...
In this study, an HPLC-DAD method was developed and validated for the quantification of bioactive phenolics in herbal medicines containing Cynara scolymus (Globe artichoke), Maytenus ilicifolia Mart ex Reiss “Espinheira santa” and Ptychopetalum uncinatum “Marapuama”. The samples were lyophilized and 5.0 g of solid were extracted with 30 mL of methanol acidified with 100 μL of concentrated HCl, under magnetic stirring at 40 °C for 30 min. Separation was carried out on a C18 column with analytical solvents constituting a binary elution mixture, consisting of (A) ultrapure water (Millipore, USA), containing 1.0% acetic acid (v v− 1) and (B) methanol (HPLC grade). Spectrophotometric detection was performed at a wavelength of 260 nm for vanillic acid; 280 nm for (+) – catechin and 330 nm for chlorogenic acid. The method to determine bioactive phenolics in herbal medicines showed adequate linearity, repeatability and accuracy. The limits of detection (LOD) and quantification (LOQ) were 0.025 μg g− 1 and 0.031 μg g− 1, respectively. The concentrations (minimum–maximum in mg g− 1) of chlorogenic acid (in samples containing C. scolymus) and vanillic acid (in herbal medicines containing P. uncinatum “Marapuama”) ranged from 71.28 to 925.99 and 17.35 to 19.21, respectively. The catechin content was 0.69 mg g− 1 in Maytenus ilicifolia Mart ex Reiss “Espinheira santa”. Therefore, the results showed that the developed method is simple, less toxic, fast and reliable for the determination of bioactive phenolics in herbal medicines.
... Several studies have provided evidence of the effects of this plant drug on the central nervous system. Initially it was found to show effects as an anxiogenic (Silva et al., 2002), anticholinesterase (Siqueira et al., 2003), neuroprotective (Siqueira et al., 2004), facilitation of memory recovery in young and old mice (Silva et al., 2004), antioxidant (Siqueira et al., 2007), antidepressant (Paiva et al., 1998;Piato et al., 2009), and anti-stress (Piato et al., 2010). Its aphrodisiac potential has not been confi rmed experimentally or clinically, although some of its effects, such as antidepressant and antioxidant activity, may improve sexual performance (Feldman et al., 1994). ...
Croton echioides Baill., Euphorbiaceae, is a small tree found in Bahia, Northeastern Brazil. Its stem bark has been widely sold as an aphrodisiac and tonic, as a substitute for the roots of Ptychopetalum olacoides Benth. Olacaceae, the Amazon Muira Puama or Marapuama, and C. echioides is characterized as the "Northeastern Marapuama". This contribution describes a morphoanatomical analysis and pharmacognostic study of stem bark of this species. The stem has a thick cortex with compound starch grains and laticifers; a sclerenchymatic sheath which consists of brachysclereids with large crystals externally to the phloem, and abundant fiber in the secondary xylem, as the main features of the species. The data obtained for water content (9.26±0.07%), water-soluble extractives (3.92±0.19%), total ash (1.24±0.06%) and acid-insoluble ash (0.16±0.01%), together with the chromatographic profile obtained by TLC, contribute to the quality control and standardization for the plant drug. The pharmacological screening indicated LD50 values above 500 mg/kg orally and equal to 500 mg/kg by the i.p. route, as well as some stimulant potential, depending on the dose.
... The roots and bark of the species are known as Marapuama or Muirapuama and have been used as a neuromuscular tonic for the treatment of chronic rheumatism, sexual impotency, and central nervous system disorders [2]. Previous pharmacological studies have indicated that the EtOH extract of P. olacoides produces a series of beneficial effects on the central nervous system in mice [3][4][5][6][7]. As part of our ongoing project to search for natural products with neurotrophic properties [8][9][10][11][12], we have continued to explore bioactive diterpenoids from the MeOH extract of the bark of P. olacoides, which exhibited neurite outgrowth-promoting activity in NGF-mediated PC12 cells [13,14]. ...
Eight new clerodane type diterpenoids, named 7-oxo-kolavelool (1), 7alpha-hydroxykolavelool (2), 6alpha,7alpha-dihydroxykolavenol (3), 12-oxo-hardwickiic acid (4), ptycholide I (5), ptycholide II (6), ptycholide III (7), and ptycholide IV (8) were isolated from the MeOH extract of the bark of a Brazilian medicinal plant, Ptychopetalum olacoides. The structures of 1-8 were elucidated by analyzing spectroscopic data and by comparing their NMR data with those of the previously reported compounds kolavelool (la), kolavenol (3a), hardwickiic acid (4a), and ptychonolide (5a). Compounds 5 and 6 existed as a 1:1 mixture of inseparable epimers at C-15.
... Special metabolites constitute important active molecules, which differ widely in terms of structure and biological properties (Argolo et al. 2004). However, a profound scientific study and standardization of these plants, as phytotherapeutics, is necessary to guarantee safety in their utilization by the public health system (Siqueira et al. 2007). In fact, Brazilian biodiversity has significantly contributed to the discovery of new pharmacologically active molecules from medicinal plants, ranging from antioxidants to anti-tumor activities (Silva et al. 2005). ...
Reactive oxygen species (ROS) are thought to underline the process of ageing and the pathogenicity of various diseases, such as neurodegenerative disorders and cancer. The use of traditional medicine is widespread and plants still present a large source of natural antioxidants that might serve as leads for the development of novel drugs. In this paper, the alcoholic extract from leaves of Hyptis fasciculata, a Brazilian medicinal plant, and isoquercitrin, a flavonoid identified in this species, showed to be active as 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavengers. The extract of Hyptis fasciculata and isoquercitrin were also able to increase tolerance of the eukaryotic microorganism Saccharomyces cerevisiae to both hydrogen peroxide and menadione, a source of superoxide. Cellular protection was correlated with a decrease in oxidative stress markers, such as levels of ROS, protein carbonylation and lipid peroxidation, confirming the antioxidant potential of Hyptis fasciculata and isoquercitrin.
... P. olacoides ethanol extract (POEE) was prepared as detailed elsewhere : briefly, dried ground roots (2.5 kg) were extracted with ethanol (12 L) in Soxhlet (40 h), and this was evaporated under reduced pressure (6% yield). Analytical HPLC was carried out on an HP 1100 system equipped with a photodiode array detector (Agilent Technologies, Santa Clora, CA, United States) and HPLC fingerprinting was performed as detailed elsewhere (Siqueira et al., 2007). ...
Depression has become of universal major importance, and it is therefore vital to expand the armamentarium for treating the condition. Lack of motivation and lassitude are major symptoms treated with the use of Marapuama (Ptychopetalum olacoides, PO) remedies by communities in the Brazilian Amazon. Considering the prominence of such symptoms in depression, the present study was designed to verify the effects of a standardized PO ethanol extract (POEE) on the forced swimming (FST) and tail suspension tests (TST). POEE i.p. (15-100 mg/kg) and oral (300 mg/kg) resulted in a significant and dose-related anti-immobility effect. We further examined the involvement of dopamine, noradrenaline and serotonin in these antidepressant-like effects. POEE effects were prevented when catecholamine synthesis was inhibited by -alpha-methyl-rho-tyrosine (AMPT) (100 mg/kg, i.p.), while inhibition of serotonin synthesis with rho-chlorophenylalanine methyl ester hydrochloride (PCPA) (100 mg/kg, i.p.) was devoid of effect. The blockade of beta-adrenergic (propranolol 2 mg/kg, i.p.) and D(1) dopamine (SCH 23390 0.5 mg/kg, i.p.) receptors prevented POEE anti-immobility effects; by contrast, blockade of D(2) dopamine (sulpiride 2 and 50 mg/kg, i.p.) receptors was ineffective. Consistent with traditional use, the results indicate that POEE possesses antidepressant-like effects, possibly mediated by beta-adrenergic and D(1) dopamine receptors.
Common name: Silver wattle, French mimosa.
Ptychopetalum olacoides is a folk medicinal plant for health care in market, especially in Brazil. Fourteen known compounds were isolated from P. olacoides and their chemical structures were elucidated by extensive spectroscopic data, including 1D NMR, 2D NMR, UV, IR and HR-ESI-MS. The 14 known compounds were identified as N-trans-feruloyl-3,5-dihydroxyindolin-2-one (1), magnoflorine (2), menisperine (3), 4-coumaroylserotonin (4), moschamine (5), luteolin (6), 4′-methoxyluteolin (7), 3-methoxyluteolin (8), 3, 7-dimethoxyluteolin (9), caffeic acid (10), ferulic acid (11), vanillic acid (12), syringic acid (13) and ginsenoside Re (14). To our knowledge, compounds (1–6, 13–14) were isolated from the plant for the first time. Additionally, quantitative analysis results indicated that calibration equations of compounds (1–3, 6, 9, 11–13) exhibited good linear regressions within the test ranges (R² ≥ 0.9990) and magnoflorine and menisperine were the major constituents in the barks of P. olacoides. The contents of magnoflorine and menisperine accounted for 75.96% of all analytes. However, the content of phenolic components was smaller and the highest content was no more than 1.04 mg/g. Collectively, these results suggested that alkaloids are the dominant substances in P. olacoides, which can make a difference for the quality control and further use of P. olacoides.
In Brazil, many plants are used as tonic, fortifier, aphrodisiac, anti-stress, among other uses that are similar to the indications of an adaptogen. In general, such plants are used unspecifically, in situations of stress and fatigue, in the recovery after a previous pathological or debilitating state, or simply aiming at the maintenance of a healthy state. This article discusses the popular terms employed in the Brazilian folk medicine for the plants with this profile, their particularities and limitations. The article also discusses the possible mechanisms of action of an adaptogen and compares the main Brazilian plants used for that purpose: guarana (Paullinia cupana Kunth, family Sapindaceae), muirapuama (Ptychopetalum olacoides Benth., Olacaceae), catuaba (Anemopaegma arvense (Vell.) Stellfeld & J.F. Souza, Bignoniaceae, and Trichilia catigua A. Juss., Meliaceae), nó-decachorro (Heteropterys aphrodisiaca O. Mach, Malpighiaceae), damiana (Turnera diffusa Willd. ex Schult., Turneraceae) and pfaffia or Brazilian ginseng (Pfaffia sp, Amaranthaceae).
Objective
To further characterize the acetylcholinesterase inhibitors (AChE-Is) pattern of Ptychopetalum olacoides ethanol extract (POEE) on the cytosolic globular monomer (G1) and membrane bound globular tetramer (G4) AChE isoforms in brain areas relevant for cognition.Methods
The G1 and G4 AChE isoforms were prepared according to the reported methods and the determination of AChE activity used was adapted from colorimetric method.ResultsPOEE mostly inhibited G1 in hippocampus (75%), and G4 in frontal cortex (58%) and striatum (75%) (P < 0.05). Kinetic analysis indicated that POEE-induced AChE inhibition in hippocampus was of a competitive nature for G1 but uncompetitive for G4.Conclusion
Considering the high density of cholinergic projection to the cortex and striatum, and the usefulness of conserving cytosolic acetylcholine to replenish synaptic vesicles in a highly active cognition site such as hippocampus, we argue that this could be a desirable profile for a clinically relevant AChE-I.
Evidence suggests that periorbital hyperchromia (dark circles) occurs mainly as a consequence of postinflammatory hemodynamic congestion producing a typical bruising aspect on the lower eyelids.
To evaluate the clinical effects of Pfaffia paniculata/Ptychopetalum olacoides B./Lilium candidum L.-associated compound (PPLAC) on periorbital hyperchromia and to study in vitro its underlying anti-inflammatory and antioxidant mechanisms.
Twenty-one volunteers presenting with periorbital hyperchromia received a serum sample containing 5.0% PPLAC, which was applied topically in the periorbital area twice a day for 28 days. Skin color was measured using variations in the individual typological angle (DeltaITA(0)) and skin luminance (DeltaL*) calculated in the area around the eyes and in the adjacent area. Colorimetric readings were taken at the onset and end of the 28-day treatment. Volunteers were also asked to fill out a questionnaire concerning the improvement in "dark circles." The anti-inflammatory and antioxidant effects of PPLAC were measured by quantification of prostaglandin E(2), leukotriene B(4), histamine, and superoxide dismutase levels using an in vitro model of human skin culture.
Topical application of PPLAC led to a significant improvement in skin luminance and tone in the periorbital area, which was demonstrated by increased values of ITA(0) and L* in about 90% of volunteers. In addition, subjects reported reduced intensity and improved appearance of "dark circles." A dose-dependent decreased production of inflammatory mediators, concomitant to increased antioxidant enzyme levels, was observed in our in vitro studies, under basal and lipopolysaccharide-stimulated conditions.
Although the precise mechanisms related to PPLAC remain to be clarified, our results indicate that the reduction in the inflammatory process as well as the antioxidant protection against deleterious elements may be considered as an integral approach to preserve the integrity of vascular endothelium, preventing the hemodynamic congestion that culminates in the formation of "dark circles" around the eyes.
From the MeOH extract of Ptychopetalum olacoides, which is used in Brazilian folk medicine for the treatment of chronic degenerative conditions of the nervous system, four novel clerodane-type diterpenoids named 6alpha,7alpha-dihydroxyannonene (1), 7alpha,20-dihydroxyannonene (2), 7alpha-hydroxysolidagolactone I (3), and ptycho-6alpha,7alpha-diol (4) were isolated by bioassay-directed fractionation using NGF-differentiated PC12 cells. The structures of 1-4 were established by extensive NMR spectroscopic analyses and chemical conversion. Compounds 1 and 2 significantly enhanced NGF-mediated neurite outgrowth in PC12 cells at concentrations ranging from 0.1 to 50.0 microM for 1 and 0.1 to 30.0 microM for 2, whereas 3 and 4 had no morphological effect on NGF-mediated PC12 cells in the same concentration range. The structure-activity relationship of these compounds is also discussed.
Four new clerodane-type diterpenoids, ptychonolide (1), 20-O-methylptychonal acetal (2), and an equilibrium mixture of ptychonal hemiacetal (3) and ptychonal (4), were isolated from the MeOH extract of the bark of a Brazilian plant, Ptychopetalum olacoides. The structure of 1 was elucidated as a clerodane-type diterpenoid on the basis of spectroscopic data, whereas 2 was assigned to an acetal derivative of 1. Compounds 3 and 4 existed as an equilibrium mixture. A mixture of compounds 3 and 4 was found to exhibit neurite outgrowth-promoting activities on NGF-mediated PC12 cells at concentrations ranging from 0.1 to 10.0 microM.
Homemade remedies with Ptychopetalum olacoides (PO) roots are used by Amazonian peoples for treating various age-related conditions. We previously reported that Ptychopetalum olacoides ethanol extract significantly improved step-down inhibitory avoidance long-term memory in adult and reversed memory deficits in aging mice. Adding to previous data, this study shows that a single i.p. administration of Ptychopetalum olacoides ethanol extract (POEE 50 and 100 mg/kg) improved step-down inhibitory avoidance short-term memory (STM) 3 h after training in adult (2.5 month) mice; comparable results were obtained with POEE given p.o. at 800 mg/kg. Moreover, memory improvement was also observed in aging (14 months) mice presenting memory deficit as compared to adult mice. Furthermore, POEE (100 mg/kg) improved non-aversive memory systems in adult mice in an object recognition paradigm. Consistently with its traditional use this study add to previously reported data and reinforces that POEE facilitates memory processes. Although the acetylcholinesterase inhibitory properties described for this extract may be of relevance for improving memory processes, the molecular mechanism(s) underlying the memory improvement here reported needs further scrutiny.
Roots of Ptychopetalum olacoides Bentham (Olacaceae), known as Marapuama, are prepared in alcoholic infusion for treating “nervous weakness” by Amazonian Caboclos. “Nervous weakness” can be described as a syndrome having several symptoms, among which the following are emphasized: lassitude, depression, sexual impotence and tremors. Based on ethnopharmacological data, we have considered the hypothesis that PO may have psychopharmacological effects, by interacting with different neurotransmitter systems: (i) the dopaminergic system, considering its use as an appetite modulator and to counteract tremors, as well as for its alleged sexual arousing properties; (ii) the noradrenergic system, again for its use against tremors and/or depression; and/or (iii) the serotonergic system, also related to depression and sexual arousal. This paper reports that P. olacoides hydroalcoholic extract potentiated yohimbine-induced lethality, rever sed reserpine-induced ptosis and prevented apomorphine-induced stereotypy. The data indicates that P. olacoides has central nervous system effects and supports the hypothesis of its interaction with dopaminergic and/or noradrenergic systems.
Alcohol infusions of roots from Ptychopetalum olacoides Bentham (PO; Olacaceae) have been used for treating many diseases in which free radicals are likely to be implicated. Of particular interest are the uses amongst the elderly (to ameliorate cognitive functions), and by patients recovering from pathologies associated with damage to the central nervous system (such as stroke). The aim of this study was to evaluate the antioxidant properties of a PO ethanol extract (POEE) by using various in vitro systems. POEE acted as a scavenger of nitrogen oxides as well as superoxide generated by the xanthine-xanthine oxidase system. The extract also showed a high antioxidant capacity using a luminol chemiluminescence derived from a thermolabile diazocompound. We suggest that the therapeutic effects attributed to P. olacoides could be in part associated to its oxygen free radical scavenging capacity.
Oxygen free radicals and oxidative events have been implicated as playing a role in bringing about the changes in cellular function that occur during aging. Brain readily undergoes oxidative damage, so it is important to determine if aging-induced changes in brain may be associated with oxidative events. Previously we demonstrated that brain damage caused by an ischemia/reperfusion insult involved oxidative events. In addition, pretreatment with the spin-trapping compound N-tert-butyl-alpha-phenylnitrone (PBN) diminished the increase in oxidized protein and the loss of glutamine synthetase (GS) activity that accompanied ischemia/reperfusion injury in brain. We report here that aged gerbils had a significantly higher level of oxidized protein as assessed by carbonyl residues and decreased GS and neutral protease activities as compared to young adult gerbils. We also found that chronic treatment with the spin-trapping compound PBN caused a decrease in the level of oxidized protein and an increase in both GS and neutral protease activity in aged Mongolian gerbil brain. In contrast to aged gerbils, PBN treatment of young adult gerbils had no significant effect on brain oxidized protein content or GS activity. Male gerbils, young adults (3 months of age) and retired breeders (15-18 months of age), were treated with PBN for 14 days with twice daily dosages of 32 mg/kg. If PBN administration was ceased after 2 weeks, the significantly decreased level of oxidized protein and increased GS and neutral protease activities in old gerbils changed in a monotonic fashion back to the levels observed in aged gerbils prior to PBN administration. We also report that old gerbils make more errors than young animals and that older gerbils treated with PBN made fewer errors in a radial arm maze test for temporal and spatial memory than the untreated aged controls. These data can be interpreted to indicate that oxidation of cellular proteins may be a critical determinant of brain function. Moreover, it also implies that there is an age-related increase in vulnerability of tissue to oxidation that can be modified by free radical trapping compounds.
The various cell groups in the human hypothalamus show different patterns of aging, which are the basis for changes in biological rhythms, hormone production, autonomic functions, and behavior. The suprachiasmatic nucleus (SCN), the clock of the brain, exhibits circadian and seasonal rhythms in vasopressin synthesis that are disrupted later in life. Furthermore, the age-related sexual differences in the number of vasoactive intestinal polypeptide neurons in this nucleus reinforces the idea that the SCN is not only involved in the timing of circadian rhythms but also in the temporal organization of reproductive functions. The sexually dimorphic nucleus of the preoptic are (SDN-POA), or intermediate nucleus, is twice as large in men as in women, a difference that arises between the ages of two to four years and puberty. During aging a dramatic, sex-dependent decrease in cell number occurs, leading to values which are only 10-15% of the cell number found in early childhood. The vasopressin and oxytocin producing cells in the supraoptic nucleus (SON) and paraventricular nucleus (PVN) are examples of neuron populations that seem to stay perfectly intact in old age. Parvocellular corticotropin-releasing hormone-containing neurons are found throughout the PVN and are even activated in the course of aging, as indicated by their increase in number and by their coexpression with vasopressin. Part of the arcuate nucleus of the hypothalamus (ARH), or tubero-infundibular nucleus, contains hypertrophic neurons in postmenopausal women. These hypertrophied neurons contain neurokinin-B, substance P, and estrogen receptors and probably act on LHRH neurons as interneurons. The tuberal lateral nucleus (NTL), involved in feeding behavior and energy metabolism, does not show any neuronal loss in senescence. These findings indicate that each cell group of the human hypothalamus has its own sex-specific pattern of aging. In fact, some hypothalamic nuclei show a dramatic functional decline with aging, whereas others seem to become more active later in life.
Alcohol infusions of roots of Ptychopetalum olacoides Benth. (PO), known as Marapuama or Muirapuama, are used in the Brazilian Amazon as a 'nerve tonic'. Over the years PO has been found increasingly in phytoformulations and regarded as a stimulant, claimed to enhance physical and mental performances. This study determined that a P. olacoides ethanol extract (30, 100 and 300 mg/kg) decreased exploratory behaviour in the hole-board test, without interfering with locomotion or motor coordination (rota-rod test). The data are comparable to that obtained with pentylenetetrazol (40 mg/kg), suggesting an anxiogenic effect of P. olacoides.
The time course of oxidative damage in different brain regions was investigated in the gerbil model of transient cerebral ischemia. Animals were subjected to both common carotid arteries occlusion for 5 min. After the end of ischemia and at different reperfusion times (2, 6, 12, 24, 48, 72, 96 h and 7 days), markers of lipid peroxidation, reduced and oxidized glutathione levels, glutathione peroxidase, glutathione reductase, manganese-dependent superoxide dismutase (MnSOD) and copper/zinc containing SOD (Cu/ZnSOD) activities were measured in hippocampus, cortex and striatum. Oxidative damage in hippocampus was maximal at late stages after ischemia (48-96 h) coincident with a significant impairment in glutathione homeostasis. MnSOD increased in hippocampus at 24, 48 and 72 h after ischemia, coincident with the marked reduction in the activity of glutathione-related enzymes. The late disturbance in oxidant-antioxidant balance corresponds with the time course of delayed neuronal loss in the hippocampal CA1 sector. Cerebral cortex showed early changes in oxidative damage with no significant impairment in antioxidant capacity. Striatal lipid peroxidation significantly increased as early as 2 h after ischemia and persisted until 48 h with respect to the sham-operated group. These results contribute significant information on the timing and factors that influence free radical formation following ischemic brain injury, an essential step in determining effective antioxidant intervention.
A protein determination method which involves the binding of Coomassie Brilliant Blue G-250 to protein is described. The binding of the dye to protein causes a shift in the absorption maximum of the dye from 465 to 595 nm, and it is the increase in absorption at 595 nm which is monitored. This assay is very reproducible and rapid with the dye binding process virtually complete in approximately 2 min with good color stability for 1 hr. There is little or no interference from cations such as sodium or potassium nor from carbohydrates such as sucrose. A small amount of color is developed in the presence of strongly alkaline buffering agents, but the assay may be run accurately by the use of proper buffer controls. The only components found to give excessive interfering color in the assay are relatively large amounts of detergents such as sodium dodecyl sulfate, Triton X-100, and commercial glassware detergents. Interference by small amounts of detergent may be eliminated by the use of proper controls.
Constituents extracted from the leaves of the Ginkgo biloba tree possess beneficial properties that may buffer the aging nervous system from deterioration due to oxidative stress. In the present investigation, a standardized extract of G. biloba (EGb 761) or an equal volume of the vehicle was administered (100 mg/kg/day) to senescent (20-month) C57BL/6 male mice for up to 82 consecutive days. Animals were tested twice in the Morris water maze (MWM) after 28 and 70 days of treatment. No differences were observed in acquisition or retention of performance on the water maze. Elevated-plus maze (EPM) trials were conducted prior to and subsequent to the chronic treatment regimen. Marked baseline differences in plus-maze performance were present in the first experiment. A second experiment used a matched-pairs design to minimize preexisting differences. Results supported the hypothesis that EGb 761 may serve as an antistress buffer, attenuating the increase in anxiety typically observed in animals after cold water exposure. Tissue samples from the hippocampus and cortex were analyzed by Western blot for the transcription factor cyclic-AMP response element binding (CREB) protein. EGb 761 had no significant effect on immunoreactivity to CREB from either the hippocampus or the cerebral cortex.
The charge (relative concentration) of antioxidants in different extracts of medicinal plants used in southwest Amazonia regions of Beni (Bolivia) and Madre de Dios (Peru) was determined employing a procedure based on the quenching of luminol-enhanced chemiluminescence derived from the thermolysis of 2,2'-azo- bis (2-amidinopropane) (ABAP) as the free radical source. Total reactive antioxidant potential (TRAP) and total antioxidant reactivity (TAR) values were determined (in Trolox equivalents) using this method, and 65 extracts showed antioxidant activity. Of these, 46 demonstrated TRAP values > 100 µM. The highest activity was noted in a methanol bark extract of Copaifera reticulata (TRAP = 7500 µM). On the other hand, the highest TAR value was obtained in the methanol root extract of Dracontium loretense. IC 50 and eficiency (compared to Trolox) values were also calculated. The results obtained indicate that the large antioxidant capacity of some medicinal plants may be due to the presence of both very reactive and low reactive antioxidants, the later in rather high concentrations, and that the therapeutic action claimed for some of these plants could be due, in part, to their capacity for scavenging oxygen free radicals which are involved in many diseases. The use of luminol-enhanced chemiluminescence proved to be a simple, sensitive and reproducible method that can be used to determine the antioxidant capacity in complex mixtures such as plant extracts.
RÉSUMÉ
Selon la théorie du radical libre, le vieillissement est produit par l'effet endommageant des radicaux d'oxygène. Ces radicaux sont produits de façon constante et deviennent toxiques quand le niveau de la concentration intracellulaire devient élevé. Malgré cet effet d'infamie, des études récentes indiquent qu'ils peuvent avoir un côté bénéfique en régulant certains mécanismes dans une cellule. Mais, avec le vieillissement, il semble que la capacité de réguler la concentration intracellulaire s'abaisse, et l'élévation du niveau devient toxique avec le temps. Il faut aussi examiner les effets endommageant des radicaux libres sur un niveau supérieur. À cause de la prééminence de l'hypothalamus comme centre régulateur homéostatique, il est possible que l'hypothalamus puisse jouer un rôle critique dans le processus de vieillissement. La perte progressive des fonctions dans ce centre peut mener à des changements systémiques qui peuvent provoquer des perturbations dans tout l'organisme.
The effect of antioxidants and biological fluids on the intensity of luminol induced chemiluminescence by radicals derived from the thermolysis of 2,2′-azo-bis(2-amidinopropane) has been employed to monitor TRAP and TOTAL ANTIOXIDANT REACTIVITY (TAR) levels. The latter parameter, which considers not only the quantity of oxidants but also their reactivity, is considered a potentially more useful index of the antioxidant status of a biological fluid. The results obtained employing human blood plasma and human urine show that the detected antioxidant capacity of both fluids is mainly related to the uric acid concentration of the samples.
Some markers of oxidative injury were measured in different rat brain areas (hippocampus, cerebral cortex, striatum, hypothalamus, amygdala/piriform cortex and cerebellum) after the systemic administration of an excitotoxic dose of kainic acid (KA, 9 mg kg(-1) i.p.) at two different sampling times (24 and 48 h). Kainic acid was able to lower markedly (P < 0.05) the glutathione (GSH) levels in hippocampus, cerebellum and amygdala/piriform cortex (maximal reduction at 24 h). In a similar way, lipid peroxidation, as assessed by malonaldehyde and 4-hydroxyalkenal levels, significantly increased (P < 0.05) in hippocampus, cerebellum and amygdala/piriform cortex mainly at 24 h after KA. In addition, hippocampal superoxide dismutase (SOD) activity decreased significantly (P < 0.05) with respect to basal levels by 24 h after KA application. On the other hand, brain areas such as hypothalamus, striatum and cerebral cortex seem to be less susceptible to KA excitotoxicity. According to these findings, the pattern of oxidative injury induced by systemically administered KA seems to be highly region-specific. Further, our results have shown that a lower antioxidant status (GSH and SOD) seems not to play an important role in the selective vulnerability of certain brain regions because it correlates poorly with increases in markers of oxidative damage.
A protein determination method which involves the binding of Coomassie Brilliant Blue G-250 to protein is described. The binding of the dye to protein causes a shift in the absorption maximum of the dye from 465 to 595 nm, and it is the increase in absorption at 595 nm which is monitored. This assay is very reproducible and rapid with the dye binding process virtually complete in approximately 2 min with good color stability for 1 hr. There is little or no interference from cations such as sodium or potassium nor from carbohydrates such as sucrose. A small amount of color is developed in the presence of strongly alkaline buffering agents, but the assay may be run accurately by the use of proper buffer controls. The only components found to give excessive interfering color in the assay are relatively large amounts of detergents such as sodium dodecyl sulfate, Triton X-100, and commercial glassware detergents. Interference by small amounts of detergent may be eliminated by the use of proper controls.
Performance of rats on a motor learning paradigm that has been demonstrated to be dependent upon cerebellar norepinephrine (NE) was studied in 20-month-old F344 rats. The behavioral task is identical to that described by Watson and McElligott: Rats are trained on a runway consisting of aluminum pegs arranged in a regular pattern. Rats receive a water reward at either end of the runway. Subsequent to training, rats are tested for running times on a runway with irregularly spaced rods. The ability of rats to improve their performance (decrease their running times) on this novel motor task is diminished in young rats that have received 6-hydroxydopamine lesions. Rats at 20 months of age are known to have deficits in cerebellar noradrenergic transmission; thus, the hypothesis to be tested was to determine if aged rats demonstrated performance deficits similar to young rats depleted of central stores of NE. The rate of acquisition of the task was determined by the decrease in running times with successive days of training. The ability of 20-month-old F344 rats to acquire proficiency on the novel motor task was impaired and the rate of acquisition of the novel motor task was not different from the young 6-hydroxydopamine-lesioned rats. In an attempt to distinguish between alterations in motor coordination and motor learning, additional tests of psychomotor performance were assessed for all groups of rats. These tests included a walking on 2.5- and 5-cm rods, speed of running on the motor task, and number and types of mistakes made on the motor learning task.(ABSTRACT TRUNCATED AT 250 WORDS)
A free radical is any species capable of independent existence that contains one or more unpaired electrons. Free radicals and other reactive oxygen species are frequently proposed to be involved in the pathology of several neurological disorders. Criteria for establishing such involvement are presented. Development of new methods for measuring oxidative damage should enable elucidation of the precise role of reactive oxygen species in neurological disorders.
Publisher Summary This chapter discusses methods to determine carbonyl content in oxidatively modified proteins. The methods described are (1) reduction of the carbonyl group to an alcohol with tritiated borohydride; (2) reaction of the carbonyl group with 2,4-dinitrophenylhydrazine to form the 2,4-dinitrophenylhydrazone; (3) reaction of the carbonyl with fluorescein thiosemicarbazide to form the thiosemicarbazone; and (4) reaction of the carbonyl group with fluorescein amine to form a Schiff base followed by reduction to the secondary amine with cyanoborohydride. Van Poelje and Snell have also quantitated protein-bound pyruvoyl groups through formation of a Schiff base with p-aminobenzoic acid followed by reduction with cyanoborohydride. Although a systematic investigation has not appeared, this method should also be useful in detecting other protein-bound carbonyl groups. Carbonyl content of proteins is expressed as moles carbonyl/mole subunit for purified proteins of known molecular weight. For extracts, the results may be given as nanomoles carbonyl/milligram protein. For a protein having a molecular weight of 50,000, a carbonyl content of 1 mol carbonyl/mol protein corresponds to 20 nmol carbonyl/mg proteins.
We examined cerebral lipid peroxidation, estimated by a thiobarbituric acid test, in rat brain regions after 30 minutes of severe forebrain ischemia and at recirculation periods of up to 72 hours. The lipid peroxide levels remained unaltered in all brain regions during ischemia and during the first hour of recirculation but were selectively increased between 8 and 72 hours of recirculation in the ischemia-sensitive regions of the hippocampus, striatum, and cortex. The most pronounced increases (30-37%) were seen at 48 hours of recirculation. In contrast, lipid peroxide levels were unchanged in infarcted brain regions 24 hours after intracarotid injection of microspheres, indicating that reoxygenation of the ischemic brain is a prerequisite for lipid peroxidation. We assessed the lipid peroxidation capacity of cerebral homogenates obtained from rats subjected to ischemia and recirculation by measuring the production of lipid peroxides after aerobic incubation. The homogenates from rats exposed to 30 minutes of ischemia or to 1 hour of recirculation were not more susceptible to peroxidation. However, the production of lipid peroxides was selectively increased in the hippocampus, striatum, and cortex at 8-48 hours of recirculation, suggesting a loss of efficacy of the antioxidant systems. These results, showing a delayed and long-lasting increase in lipid peroxidation that occurs in ischemia-sensitive brain regions and parallels the development of neuronal necrosis, support the hypothesis that free radical processes participate in postischemic neuronal damage.
The capillary loops of the subcommissural organ of the rabbit consist of nonfenestrated endothelial cells interconnected by circular tight junctions. The endothelial wall is surrounded by a perivascular connective tissue sheath. Horseradish peroxidase (HRP), after intravenous injection, cannot leave the capillary lumen because of the interendothelial tight junctions. After intraventricular injection, HRP enters the intercellular clefts from the ventricles through apical gap junctions and travels toward the perivascular sheath. After crossing the extracellular compartment of the perivascular space and the subendothelial basal lamina, HRP reaches the distal part of the interendothelial cleft but never the luminal side of the tight junction. The lack of transport of this protein molecule does not favor the concept of basal secretion.
Publisher Summary Catalase exerts a dual function: (1) decomposition of H 2 O 2 to give H 2 O and O 2 (catalytic activity) and (2) oxidation of H donors, for example, methanol, ethanol, formic acid, phenols, with the consumption of 1 mol of peroxide (peroxide activity). The kinetics of catalase does not obey the normal pattern. Measurements of enzyme activity at substrate saturation or determination of the K s is therefore impossible. In contrast to reactions proceeding at substrate saturation, the enzymic decomposition of H 2 O 2 is a first-order reaction, the rate of which is always proportional to the peroxide concentration present. Consequently, to avoid a rapid decrease in the initial rate of the reaction, the assay must be carried out with relatively low concentrations of H 2 O 2 (about 0.01 M). This chapter discusses the catalytic activity of catalase. The method of choice for biological material, however, is ultraviolet (UV) spectrophotometry. Titrimetric methods are suitable for comparative studies. For large series of measurements, there are either simple screening tests, which give a quick indication of the approximative catalase activity, or automated methods.
Publisher Summary This chapter presents a procedure for the preparation of glutathione peroxidase, which is regarded as a major protective system against endogenously and exogenously induced lipid peroxidation. Two types of methods are used for determining the activity of glutathione peroxidase. One involves a direct measurement of unconsumed glutathione (GSH) at fixed time periods by polarographic GSH analysis' (Method 1), or by the dithionitrobenzoic acid method (Method 2). The second approach takes advantage of the capability of glutathione reductase, with nicotinamide adenine dinucleotide phosphate (NADPH), to regenerate GSH from oxidized GSH. The decrease in NADPH is continuously measured spectrophotometrically, while the GSH concentration in the enzymatic cycle remains essentially constant (Method 3). A convenient source for the preparation of glutathione peroxidase is bovine blood including the following steps: hemolysate; organic solvent precipitation; phosphate precipitation; absorption to phenyl-sepharose; and washing on diethylaminoethyl (DEAE)–sephadex, S-300 sephacryl, and hydroxylapatite column.
To investigate the relationship between oxidative stress and aggrevation of the disease in patients with malaria.
In the present study lipoperoxidation was demonstrated during the acute phase of malaria by a significant decrease in polyunsaturated fatty acids (PUFA). The lowest values of PUFA were obtained for C20:4 and C22:6, which were the main targets of reactive oxygen species (ROS) when parasitemia was higher than 1%. Similarly, plasma vitamins E and A were significantly reduced during the acute phase of malaria owing to their consumption in part as antioxidants. However, evaluation of the antioxidant enzymatic system in red blood cells of malaria patients indicated no significant difference from controls. Only superoxide dismutase activity tended to decrease when parasitemia increased.
The results suggest that superoxide radicals are the main ROS produced during the acute phase of malaria, and that rejuvenation of RBC during hemolysis involving increased enzyme activities interacts to protect RBC from excessive superoxide radical production.
The aging brain undergoes a process of enhanced peroxidative stress, as shown by reports of altered membrane lipids, oxidized proteins, and damaged DNA. The aims of this review are to examine: (1) the possible contribution of mitochondrial processes to the formation and release of reactive oxygen species (ROS) in the aging brain; and (2) the age-related changes of antioxidant defenses, both enzymatic and nonenzymatic. It will focus on studies investigating the role of the electron transfer chain as the site of ROS formation in brain aging and the alterations of the glutathione system, also in relation to the effects of exogenous pro-oxidant agents. The possible role of peroxidative stress in age-related neurodegenerative diseases will also be discussed.
We have demonstrated that aged rats show impairments in learning patterned motor movements. Similar behavioral impairment is observed in rats with noradrenergic lesions. Norepinephrine is known to act as a neuromodulator in the cerebellar cortex because it can augment the action of GABA and other neurotransmitters. This effect of NE to augment the signal to noise ratio of GABAergic inputs to cerebellar Purkinje neurons is a possible substrate for NE's effect on motor learning. Aged rats demonstrate deficits in the modulatory actions of NE to augment GABAergic inhibitions when both substances are locally applied onto cerebellar Purkinje neurons. In this report, we examined how motor learning and cerebellar noradrenergic function varied in individual young and 20-month-old Fischer 344 rats. There was a significant correlation between the loss of the neuromodulatory actions of norepinephrine (NE) in the cerebellar cortex and the rate of learning a novel motor task in individual rats. This report thus demonstrates for the first time a correlation between age-related impairments in motor plasticity and specific neurophysiological deficits in cerebellar Purkinje neurons in individual animals.
Free radicals and oxidative damage have been implicated in brain aging and several neurodegenerative diseases. The purpose of the present study was to determine whether antioxidants could alleviate age-associated cognitive and motor changes. Aged 24-month-old male Sprague-Dawley rats were treated for 4-5 months with daily i.p. injections of spin-trapping compound phenyl-alpha-tert-butylnitrone (PBN; 32 mg/kg) and alpha-tocopherol (200 mg/kg) or with vehicles. Antioxidant-treated animals also received ascorbate in their drinking water. In Morris water maze testing after two months, antioxidant-treated rats exhibited significantly greater memory retention than vehicle-treated rats in water maze testing. Subsequent tests for passive avoidance behavior and motor activity/skill revealed no effect of antioxidant treatment. In a separate group of aged 33-month-old rats that received the same combination of antioxidants for only 14 days, antioxidant treatment did not affect basal levels of brain lipid peroxidation (as indexed by TBAR formation) compared to controls. The results of this study provide initial evidence that chronic antioxidant treatment can improve cognitive function during aging, thus supporting the 'free radical hypothesis of aging' related to brain function.
The role of oxidative stress in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-mediated neurotoxicity is as yet unclear and the evidence for generation of oxygen free radicals as a primary event in the neurotoxicity is yet to be demonstrated. The present study was undertaken to ascertain the potential role of oxidative damage, and the protective role, if any, of the antioxidant, glutathione (GSH), in MPTP-induced neurotoxicity. Exposure of sagittal slices of mouse brain to MPTP resulted in significant increases of reactive oxygen species (ROS) and malondialdehyde (MDA, the product of lipid peroxidation) and decreases in GSH content. Pretreatment of mouse brain slices, in vitro, with GSH or GSH isopropyl ester attenuated MPTP toxicity as assessed by the tissue activity of the mitochondrial enzyme, NADH-dehydrogenase (NADH-DH), and by leakage of the cytosolic enzyme, lactate dehydrogenase (LDH), from the slice into the medium. In vivo administration of MPTP (30 mg/kg body weight, s.c.), to mice resulted in significant lowering of GSH in the striatum and midbrain, 2 h after dosage; ROS levels in the striatum and midbrain increased after 4 and 8 h, respectively. In the striatum significant inhibition of rotenone-sensitive NADH ubiquinone-1 oxido-reductase (Complex 1) was observed transiently 1 h after MPTP administration. The enzyme activity recovered thereafter; significant inhibition of mitochondrial Complex I was observed in the striatum only 18 h after MPTP dose. In the midbrain, mitochondrial Complex I was inhibited only 18 h after MPTP dose; no change was observed at the early time points examined. Thus, the depletion of GSH and increased ROS formation preceded the inhibition of the mitochondrial enzyme in the midbrain. Evidence presented herein from both in vitro and in vivo studies support that MPTP exposure generates ROS resulting in oxidative stress.
The free radical theory of aging predicts that reactive oxygen species are involved in the decline in function associated with aging. The present paper reports that diets supplemented with either spinach, strawberries or blueberries, nutritional sources of antioxidants, reverse age-induced declines in beta-adrenergic receptor function in cerebellar Purkinje neurons measured using electrophysiological techniques. In addition the spinach diet improved learning on a runway motor task, previously shown to be modulated by cerebellar norepinephrine. Motor learning is important for adaptation to changes in the environment and is thus critical for rehabilitation following stroke, spinal cord injury, and the onset of some neurodegenerative diseases. These data are the first to indicate that age-related deficits in motor learning and memory can be reversed with nutritional interventions.
Manganese superoxide dismutase (MnSOD) overexpression has been shown to reverse the malignant phenotype in a variety of tumor cell lines. The inhibition of proliferation and reversal of the malignant phenotype has been attributed to an increase in H(2)O(2) production as a result of the dismutation reaction. However, direct evidence in support of this hypothesis has not been forthcoming. To evaluate the contribution of H(2)O(2) in the regulation of cell growth in response to MnSOD overexpression, control and MnSOD-overexpressing HT-1080 fibrosarcoma cells were transfected with constructs that direct catalase to either the mitochondrial or cytosolic compartments. Overexpression of catalase in either compartment reversed the proliferative and clonogenic inhibition associated with MnSOD overexpression, blocked the increase in the steady state levels of H(2)O(2) as measured by flow cytometric analysis of 2', 7'-dichlorofluorescein diacetate, and increased protection from the cytotoxicity of H(2)O(2). In addition, mitochondrial or cytosolic catalase enhances respiration through complex I and II in both control and MnSOD overexpressing cell lines and reverses a MnSOD-dependent decrease in net ATP production. Thus, catalase reverses the proliferative inhibition associated with MnSOD overexpression and may also play an important role in metabolic regulation.
This study pertains to the role of lupeol, a pentacyclic triterpene, against the toxic manifestations of chronic cadmium exposure. Cadmium is a potent nephrotoxin on prolonged exposure. Cadmium (as cadmium chloride) at a dose of 1 mg kg(-1) body weight was administered subcutaneously for 15 days to rats. This led to an increase in the level of lipid peroxides and a decrease in the level of antioxidants in the kidney. Lupeol, when supplemented at a dosage of 40 mg kg(-1) body weight concurrent with cadmium administration, showed an improvement in the antioxidant status. The level of lipid peroxides also showed a significant decrease, which shows the nephroprotective action of the drug.
The life span of a species is thought to be determined by the rate of mitochondrial damage which in turn is inflicted by free radicals in the mitochondria during the course of normal metabolism. The level of lipid peroxidation and antioxidants were measured in liver and kidney mitochondria of young and aged rats before and after DL-alpha-lipoic acid supplementation. In both liver and kidney, mitochondrial lipid peroxidation increased with age and a decrease in the enzymatic and non-enzymatic antioxidants were observed. DL-alpha-lipoic acid treated aged rats showed a decrease in the level of lipid peroxides and an increase in the antioxidant status. Our results conclude that supplementation of lipoic acid restores the depleted mitochondrial antioxidant status and suggest that it could be an effective therapeutic agent in treatment of age-associated disorders where free radicals are the major causative factor.
The free radical scavenging abilities of the structurally related steroids beta-sitosterol, beta-sitosterol glucoside (plant sterols and sterolins), cholesterol, and dehydroepiandrosterone sulphate (DHEAS) were compared with melatonin (an efficient free radical scavenger) in an in vitro system which measures lipid peroxidation of platelet membranes in the presence of iron (Fe2+). Lipid peroxidation is a process whereby cellular membranes are damaged due to the oxidative deterioration of polyunsaturated lipids, which may lead to cell death and disease in living organisms. Substances such as vitamin E protect cellular membranes against oxidative damage due to their chemical structures. The steroids cholesterol, beta-sitosterol, beta-sitosterol glucoside and dehydroepiandrosterone (DHEA) are structurally related to each other. During aging, serum concentrations of DHEA, DHEAS and melatonin decrease, while the concentration of cholesterol tends to increase. The aim of the present study was to compare the role these substances play in lipid peroxidation over a wide concentration range. At concentrations lower than the free iron in the reaction mixture, all the steroids investigated decreased lipid peroxidation. At higher concentrations, cholesterol and beta-sitosterol increased lipid peroxidation, while DHEAS and melatonin continued to decrease lipid peroxidation.
Constituents extracted from the leaves of the Ginkgo biloba tree possess beneficial properties that may buffer the aging nervous system from deterioration due to oxidative stress. In the present investigation, a standardized extract of G. biloba (EGb 761) or an equal volume of the vehicle was administered (100 mg/kg/day) to senescent (20-month) C57BL/6 male mice for up to 82 consecutive days. Animals were tested twice in the Morris water maze (MWM) after 28 and 70 days of treatment. No differences were observed in acquisition or retention of performance on the water maze. Elevated-plus maze (EPM) trials were conducted prior to and subsequent to the chronic treatment regimen. Marked baseline differences in plus-maze performance were present in the first experiment. A second experiment used a matched-pairs design to minimize preexisting differences. Results supported the hypothesis that EGb 761 may serve as an antistress buffer, attenuating the increase in anxiety typically observed in animals after cold water exposure. Tissue samples from the hippocampus and cortex were analyzed by Western blot for the transcription factor cyclic-AMP response element binding (CREB) protein. EGb 761 had no significant effect on immunoreactivity to CREB from either the hippocampus or the cerebral cortex.
It has been widely recognized that the hippocampus and striatum are clearly more susceptible to oxidative stress than the remaining brain regions. However, the mechanism involved is not known. The activities of the antioxidant enzymes CuZn-superoxide dismutase (SOD), Mn-SOD and catalase were measured in the hippocampus and striatum and the results were compared to cortex and cerebellum (less susceptible to oxidative stress) after 3 h of a global transient ischemia/reperfusion. CuZn-SOD activities were reduced in all brain regions, but mainly in the hippocampus and striatum. Mn-SOD activity was lowered in the striatum, whereas catalase activity was reduced in the hippocampus and striatum. Our findings indicate that in the earlier phase of ischemia/reperfusion the decay in activities of catalase and SOD may be related with the high susceptibility of the hippocampus and striatum to oxidative damage.
The cholinergic hypothesis of Alzheimer disease (AD) has provided the rationale for the current pharmacotherapy of this disease, in an attempt to downgrade the cognitive decline caused by cholinergic deficits. Nevertheless, the search for potent and long-acting acetylcholinesterase (AChE) inhibitors that exert minimal side effects to AD patients is still an ongoing effort. Amazonian communities use traditional remedies prepared with Ptychopetalum olacoides (PO, Olacaceae) roots for treating various central nervous system conditions, including those associated with aging. The fact that PO ethanol extract (POEE) has been found to facilitate memory retrieval in the step down procedure in young and aged mice prompt us to evaluate its effects on AChE activity in memory relevant brain areas. POEE significantly inhibited AChE activity in vitro in a dose- and time-dependent manner in rat frontal cortex, hippocampus and striatum; a significant inhibition was also found in these same brain areas of aged (14 months) mice after acute administration of POEE (100 mg/kg ip). We propose that such AChE inhibitory activity is a neurochemical correlate of a number of therapeutic properties traditionally claimed for P. olacoides, particularly those associated with cognition.
Although the specific causes of Alzheimer's disease have not yet been determined, considerable circumstantial evidence implicates beta-amyloid, an insoluble polypeptide made up of 39 to 42 amino acids, in the continuing destruction of brain cells that results in the progressive deterioration of the patient's mental ability. The toxic actions of beta-amyloid appear to be due to free radicals generated by a portion of the beta-amyloid molecule. These free radicals damage various parts of the neuron and lead to increased intracellular calcium which is also toxic. beta-Amyloid is formed by the aberrant processing of a much larger precursor protein that is made when cells are damaged. The normal processing of this precursor protein not only prevents the formation of beta-amyloid, but produces a soluble protein that regulates the entry of calcium into neurons and has cytoprotective actions. Interventions to prevent the destruction of neurons and the disruption of brain function by beta-amyloid include the administration of antioxidants and free radical scavengers to reduce further neural damage from deposits of beta-amyloid, the activation of various growth factors to repair damaged cells and restore their functions, and the stimulation of the normal processing of the precursor protein not only to aid in neural repair but more importantly to prevent the formation of additional beta-amyloid.
Amazonian peoples use traditional remedies prepared with Ptychopetalum olacoides (PO) roots for treating various age-related conditions. This study shows that a single intraperitoneally (i.p.) administration of Ptychopetalum olacoides ethanol extract (POEE, 50 and 100mg/kg) improved memory retrieval in step-down inhibitory avoidance (P <or= 0.05 and P <or= 0.01, test session latency 102 [19.38-300] and 192 [91.3-300]s, respectively versus control 24.7 [12.9-89.6]), without interfering with acquisition or consolidation in adult (2.5-month-old) mice. Comparable results were obtained with POEE given p.o. at 800 and 1000mg/kg (P <or= 0.05 and P <or= 0.01, 52.7 [19.5-297.2] and 85.7 [44.4-260.4] versus control 20.5 [8-92.6]). Moreover, memory amelioration was also observed (P <or= 0.01) in aging (14 months) mice presenting memory deficit (14.95 [10.8-41]) as compared to adult (2.5 months) mice (57 [15.7-141.2]), with the extract given acutely i.p. 100 mg/kg (300 [133.1-300] versus control 14.95 [10.8-41]) or p.o. 800 mg/kg (28.4 [15.1-84.6] versus control 11.5 [7.8-23.3]). Indeed, aging mice treated with POEE (800 mg/kg, p.o.) performed as well as adult mice. Consistently with its traditional use, the data suggest that POEE facilitates memory retrieval. Although the antioxidant and acetylcholinesterase inhibitory properties previously described for this extract may be of relevance, the molecular mechanism(s) underlying the improvement in memory retrieval here reported merit further scrutiny.
From indigenous disease concepts to laboratory work hypothesis: the case of “nerve tonics” from the Brazilian Amazon
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