Enitome E. Bafor

Acute toxicity studies of the leaf extract of Ficus exasperata on haematological parameters, body weight and body temperature.

INTRODUCTION The plant, Ficus exasperata is popularly used in Nigeria and in several parts of Africa for a variety of ailments. AIM OF STUDY This study was thus mapped out to investigate the toxicity profile of the aqueous leaf extract (AET) on haematological parameters, body weight and body temperature in mice. METHODOLOGY In the present study, AET was evaluated for acute toxicity over 24 h and 14-day periods. The LD(50) was assessed via oral and intraperitoneal administration. RESULTS The LD(50) was indeterminable via the oral route but was determined to be 0.54 g/kg i.p. In the 24h and 14 days single dose study, oral administration of 2.5, 5, 10 and 20 g/kg of AET produced neither mortality nor changes in behavior or any other physiological activity in mice. Body weights and body temperatures were not significantly altered. Haematological analysis showed no marked differences in any of the parameters examined (WBC count, platelet and haemoglobin estimation) in either the control or treated groups. However, the 14 days daily dose study showed significant increase in body temperature (p<0.05) and a significant decrease in the red blood cell count, haemoglobin count and haematocrit values (p<0.05), while other parameters remained unchanged. CONCLUSION In summary, AET was found to be relatively safe on short-term oral administration. However, chronic toxicity studies are required for the support of the safe use of this plant.

The leaves of Ficus exasperata Vahl (Moraceae) generates uterine active chemical constituents

ETHNOPHARMACOLOGICAL RELEVANCE In the search for new, safe and efficacious uterine active agents, the plant Ficus exasperata was subjected to phytochemical screening and pharmacological analysis. MATERIALS AND METHODS Ethyl acetate and methanolic leaf extracts of Ficus exasperata were fractionated and purified by a series of chromatographic techniques. The isolation process was guided by in vitro functional uterine assays involving the use of C57Bl/6 female mice. Identification of the active chemical constituents was performed by several spectroscopic techniques which included 1D and 2D nuclear magnetic resonance (NMR) and high resolution mass spectrometry (HRMS). The uterine effects of these compounds were investigated on spontaneous, oxytocin-induced and high KCl-induced contractions using isolated uterine segments of non-pregnant female mice. The activity of different compounds on the amplitude (maximum tension above basal force) and frequency of uterine contractions were simultaneously measured and then statistically analysed. The structure-activity relationships were also examined where possible. RESULTS These studies led to the identification of some new phytochemical derivatives. Pharmacological assay revealed the presence of both uterine stimulatory and inhibitory constituents. The new pheophytin/pheophorbide derivatives, flavonoids, fatty acids and glycerol derivatives significantly reduced the frequency and amplitude of uterine contraction, while KCl salt, pyrimidine and pheophorbide-b derivatives significantly augmented both spontaneous and agonist-induced contractions. CONCLUSION This study has demonstrated that Ficus exasperata generates secondary metabolites which have proven effective in the significant inhibition of uterine contractions and thus a potential source of new tocolytic agents. Additionally, uterine stimulatory constituents were also generated some of which may be potential drugs for contraception and/or labour facilitation. Lead compounds generated from this study are the pheophytin/pheophorbide derivatives, pyrimidine derivatives and flavonoid derivatives.

Characterisation of the antiproliferative constituents and activity of Ficus exasperata (Vahl) on ovarian cancer cells – a preliminary investigation

Abstract Ovarian cancer is one of the most common gynaecological cancers today. This study therefore investigates the anticancer effects of Ficus exasperata extracts and fractions on ovarian cancer cells. The antiproliferative activity of the crude extracts (1 mg/mL) was assessed using the MTT assay on A2780 (ovarian cancer) cell line. Bio-activity guided fractionation was performed and preliminary identification was further achieved using high resolution mass spectrometry and nuclear magnetic resonance spectroscopy. All crude extracts tested exhibited antiproliferative activity except for the methanol extract which interestingly showed proliferative effects. Five fatty acids were identified from the active fractions (FB1-10 and FB1-12). FB1-12 exhibited an IC50 value of 15.20 μg/mL. The least potent fraction (FB1-4 + 5) had an IC50 value of 34.51 μg/mL. H1-HEX and H1-MET exhibited 97.2 and 97.9%, respectively, compared to control. This study therefore provides proof-of-principle that fatty acids of Ficus exasperata exhibit significant antiproliferative effects on ovarian cancer cells.

In vitro determination of the mechanism of the uterine stimulatory effect of Newbouldia laevis

The uterine stimulatory effect of the ethanol leaf extract of Newbouldia laevis (Beauv.) Seemann ex Bureau (Bignoniaceae) was evaluated in the presence of some antagonists in vitro in an attempt to elucidate the mechanism of action of the extract. The extract was tested in the presence and absence of phentolamine (4.09 and 40.91 nM), diphenhydramine (4.45 and 44.47 nM), atropine (1.18 and 11.91 nM), and verapamil (2.03 and 20.35 nM). The effect of the antagonists on the extract and on oxytocin used as a reference drug in this study was evaluated. The EC50 and Emax were determined and statistically analyzed using one way ANOVA and Dunnett’s post hoc test. There was no significant difference in the EC50 and Emax of the extract and oxytocin in the presence of phentolamine. Diphenhydramine and atropine significantly inhibited (p <0.01) the extract but both drugs had no effect on oxytocin. However, significant differences (p <0.01) were observed in the EC50 and Emax of the extract and oxytocin in the presence of verapamil. These results suggest that the leaf extract of N. laevis contracts the uterus by opening voltage-operated calcium channels and/or by activation of muscarinic receptors.

A role of alpha-tocopherol and phylloquinone in the modulation of uterine contractility and reproductive function in mouse models

BACKGROUND AND AIM Alpha-tocopherol has been implicated in reproduction processes, and deficiency of phylloquinone has been associated with serious complications in pregnancy. This study was therefore aimed at investigating the effects of phylloquinone and alpha-tocopherol on uterine contractility and female reproductive function using mouse models. MATERIALS AND METHODS Both in vivo and ex vivo animal models were employed and designed to assess changes on uterine contractility and reproductive functions in the non-pregnant uterus. The effect of alpha-tocopherol and phylloquinone on spontaneous uterine contractions, oxytocin-induced uterine contractions (11.82nM) and high KCl-induced tonic uterine contractions (80mM) were assessed. The effect of subcutaneous administration of alpha-tocopherol (10mg/kg) on reproductive hormone levels and reproductive tissues were also determined. RESULTS Alpha-tocopherol increased the force of contractions while phylloquinone decreased the force of uterine contractions. Plasma levels of luteinizing hormone (P<0.01), estrogen (P<0.01) and progesterone (P<0.001) were elevated in the presence of alpha-tocopherol after 6 days subcutaneous administration. CONCLUSIONS Alpha-tocopherol and phylloquinone have been shown to directly modulate uterine contractility and reproductive function and may contribute to the management and treatment of reproductive disorders.

Toward understanding myometrial regulation: metabolomic investigation reveals new pathways of oxytocin and ritodrine activity on the myometrium

In recent times, additional pathways involved in the regulation of the myometrium have been suggested. This also holds true for the effect of drugs such as oxytocin (OT) and β-adrenergic agonists on the myometrium. Knowledge of these additional pathways will certainly prove useful in designing better therapies for pathologies of the myometrium. This study was therefore aimed at investigating the possibility of other pathways involved in the activities of both OT and ritodrine (RIT; a β-adrenergic agonist) in the myometrium by utilizing metabolomics and bioinformatics. High-resolution Fourier transform mass spectrometry (HRFTMS) and nuclear magnetic resonance (NMR) spectroscopy coupled with functional uterine assays were used for an innovative assessment. In vitro pharmacological assay of OT (1 nmol/L) and RIT (0.1 nmol/L) on isolated mice uteri mounted in 3 mL organ baths was performed. Mice uteri, treated with OT or RIT, as well as the physiological buffer in which the uterine tissues were immersed, were rapidly collected and analyzed using HRFTMS, proton (1H)-NMR, and bioinformatics. Resulting data were analyzed via pairwise chemometric comparison models, with P ≤ .05 considered statistically significant. In addition to previously known metabolites, nicotinamide adenine dinucleotide, γ-aminobutyric acid, and sphingosine were significantly associated with the activity of OT, whereas the activity of RIT was associated with a downstream involvement of prostaglandin F1 and phosphatidylinositol signaling. These findings add evidence to the reports on additional regulation of myometrial activity by these drugs and suggest newer pathways for therapeutic manipulation.In recent times, additional pathways involved in the regulation of the myometrium have been suggested. This also holds true for the effect of drugs such as oxytocin (OT) and β-adrenergic agonists on the myometrium. Knowledge of these additional pathways will certainly prove useful in designing better therapies for pathologies of the myometrium. This study was therefore aimed at investigating the possibility of other pathways involved in the activities of both OT and ritodrine (RIT; a β-adrenergic agonist) in the myometrium by utilizing metabolomics and bioinformatics. High-resolution Fourier transform mass spectrometry (HRFTMS) and nuclear magnetic resonance (NMR) spectroscopy coupled with functional uterine assays were used for an innovative assessment. In vitro pharmacological assay of OT (1 nmol/L) and RIT (0.1 nmol/L) on isolated mice uteri mounted in 3 mL organ baths was performed. Mice uteri, treated with OT or RIT, as well as the physiological buffer in which the uterine tissues were immersed, were rapidly collected and analyzed using HRFTMS, proton (1H)-NMR, and bioinformatics. Resulting data were analyzed via pairwise chemometric comparison models, with P ≤ .05 considered statistically significant. In addition to previously known metabolites, nicotinamide adenine dinucleotide, γ-aminobutyric acid, and sphingosine were significantly associated with the activity of OT, whereas the activity of RIT was associated with a downstream involvement of prostaglandin F1 and phosphatidylinositol signaling. These findings add evidence to the reports on additional regulation of myometrial activity by these drugs and suggest newer pathways for therapeutic manipulation.

Amelioration of Escherichia coli-induced endometritis with ascorbic acid in non-pregnant mouse models

Infection‐induced endometritis is associated with infertility. The outcome with oral antibiotics remains poor. This study therefore investigates the role of ascorbic acid in resolving endometritis.

In vitro inhibitory effect of methanol leaf extract of Stachytarpheta jamaicensis (Verbenaceae) on nonpregnant rat uterus

Purpose: To investigate the effect of the methanol extract of the plant, Stachytarpheta jamaicensis (Verbenaceae) (SJ) on uterine smooth muscles of non-pregnant female rats, with the aim of examining the oxytocic or otherwise effect of the extract. Methods: In the first phase of experiments, the effects of SJ (0.41 and 4.01 mg/ml) on oxytocin (OT) induced uterine contractions were determined and repeated after addition of salbutamol (SBL) (41.7 nM). In the second phase, verapamil (VER) (2.03 μM), SBL (41.7 nM), and SJ (0.41 and 4.01 mg/ml) were applied to the tissues after pre-contraction with K+ (80 mM). In the third phase, the effects of VER, SBL, and SJ on CaCl 2 -induced contractions (0.03 – 10.83 mM) were examined. In the fourth and final phases, the second phase experiments were repeated in a calcium-free medium and in the absence and presence of propranolol (1.54 mM) respectively. Results: SJ exhibited significant inhibitory effects on OT and CaCl 2 induced uterine contractions ( p < 0.05). The EC 50 for OT increased from 1.92 ± 0.12 to 7.16 ± 0.16 nM and that for CaCl 2 increased from 0.19 ± 0.09 to 0.76 ± 0.11 mM in the presence of the extract. SJ also significantly inhibited KCl- induced contraction by 39.15 ± 2.13 and 53.23 ± 1.58 % for 0.41 and 4.01 mg/ml of the extract, respectively ( p < 0.01); this inhibition was unaffected by propranolol. On the other hand, SBL and VER showed inhibition of 77.25 ± 1.85 and 79.44 ± 2.27 %, respectively. Conclusion: SJ exerts uterine inhibitory effects in rats which appear unrelated to β 2 -adrenergic receptor stimulation but possibly through inhibition of calcium entry into the cytoplasm. Keywords: Oxytocin, Calcium-free, Uterine contraction, β 2 -Adrenergic receptor stimulation, verapamil, salbutamol, Propranolol, S. jamaicensis

Metabolomics-coupled functional pharmacology of chlorophyll compounds isolated from the leaves of Ficus Exasperata Vahl (Moraceae) provides novel pathways on myometrial activity

New chlorophyll derivatives (pheophytins along with pheophorbide derivatives) were isolated from the leaves of Ficus exasperata and were found to have varying effects on uterine contractility. The current study was therefore aimed at the utilization of mass spectrometry and nuclear magnetic resonance spectroscopy coupled with isolated uterine tissue assay as a platform to assist in the determination of the mechanism of activity of the isolated chlorophyll compounds from the plant F exasperata. The pheophytin and pheophorbide compounds (200 µg/mL) were added to the isolated uterine tissues. Mice uteri, treated with the pheophytin compounds, and the physiological buffer in which the uterine tissues were immersed, were rapidly collected and analyzed using high-resolution Fourier transform mass spectrometry and proton (1H) nuclear magnetic resonance for bioinformatics study. Resulting data were analyzed via pairwise chemometric comparison models, with P < .05 considered statistically significant. Primary signaling pathways found to be correlated with the pheophytins in this study included cyclic adenosine monophosphate, dopamine, extracellular signal-regulated kinases 1/2, and glutamate pathways.

EDITORIAL: Potentials for Use of Medicinal Plants in Female Reproductive Disorders – The Way Forward

In the past years, the use of traditional medicine has gained much recognition worldwide, although more engrained in some cultures than others. This form of medicine relies on the use of certain herbal plants (medicinal plants) and other remedies for beneficial biological effects. In Africa and Asia there appears to be a high reliance on medicinal plants particularly by people of lower income and this has been attributed partly to the often unreachable cost of allopathic drugs, unavailability of modern health care facilities, and also the cultural acceptability of the traditional system. At the same time, there are some fields of thought that appear to disregard or discountenance the importance and use of medicinal plants. Such thoughts may have arisen for several reasons including, the idea that medicinal plant use is mainly for low income earners, that the use of medicinal plants have little to offer, and that medicinal plants have no activity and act more like a placebo. Evidence from scientific research worldwide has however proven that medicinal plants have immense biological and health applicability. Some of the earliest drugs were first discovered from traditionally used plants prior to their availability as synthesized drugs. Medicinal plants should therefore be accorded due consideration and support across different health and scientific disciplines. The use of medicinal plants has found application in several diseases and health conditions and reproductive disorders are not left out. In the traditional system of medicine, plant preparations in the forms of macerations, tinctures, concoctions, or infusions are used for a wide range of diseases. The application of medicinal plants to female reproductive health issues is gaining interest, as reproductive disorders are considered an important public health and social problem. In developing countries, particularly in sub–Saharan Africa, reproductive health disorders pose a major burden and are considered the second most prevalent health care problem in Africa. The starting point for use of plants often begins from traditional healers and natives of a particular culture. The plants utilized vary from culture to culture which is not surprising owing to the vast varieties of plants in existence and also to the varying localization of plants from one geographical region to another. Plants collected may include species with known biological activity on reproductive disorders for which active compound(s) have not been isolated. On collection of plants, phytochemists (natural product chemists) prepare the extracts from the plant materials, which are subjected to biological screening using pharmacologically relevant assays, and then proceed to the isolation and characterization of the active compound(s) through bioassay-guided fractionation. Medicinal plants have played an important role of providing new chemical entities through the years even so for female reproductive health issues. Detailed ethnobotanical surveys have been reported for several native communities around the world and many traditionally-used plants or remedies have been identified. The knowledge and the correct use of these natural medicines has been acquired and improved over many generations. Documentation of traditional knowledge of medicinal plants is crucial, since it provides chemists and pharmacologists with starting points for ―targeted‖ analysis, discovery of novel remedies, and natural drugs for the treatment of pregnancy and birth-related problems.