Original Manuscript Journal of Evidence-Based Integrative Medicine Volume: 27: 1-8 Anti-plasmodial, Cytotoxic and © The Author(s) 2022 Article reuse guidelines: Antioxidant Activities of Selected sagepub.com/journals-permissionsDOI: 10.1177/2515690X211073709 Ghanaian Medicinal Plants journals.sagepub.com/home/cam Regina Appiah-Opong, PhD1 , Kojo Agyemang, MPhil1, Eunice Dotse, MPhil1, Philip Atchoglo, MPhil1, Kofi Baffour-Awuah Owusu1, Abigail Aning1, Maxwell Sakyiamah, PhD2, Richard Adegle, BSc2, Frederick Ayertey, MPhil2 , Alfred Ampomah Appiah, PhD2, and Alexander K. Nyarko, PhD3 Abstract Malaria affects about half of the world’s population. The sub-Saharan African region is the most affected. Plant natural products have been a major source of antimalarial drugs; the first (quinine) and present (artemisinin) antimalarials are of natural product origin. Some secondary metabolites demonstrate adjuvant antioxidant effects and selective activity. The focus of this study was to inves- tigate the anti-plasmodial activity, cytotoxicities and antioxidant properties of eight (8) Ghanaian medicinal plants. The anti-plasmo- dial activity was determined using the SYBR green assay and the tetrazolium-based colorimetric assay (MTT) was employed to assess cytotoxicity of extracts to human RBCs and HL-60 cells. Antioxidant potential of plant extracts was evaluated using Folin-Ciocalteu and superoxide dismutase assays. Phytochemical contstituents of the plant extracts were also assessed. All the extracts demon- strated anti-plasmodial activities at concentrations <50 μg/ml. Parkia clappertoniana and Terminalia ivorensis elicited the strongest anti-plasmodial activities with 50% inhibitory concentrations (IC50) of 1.13 μg/ml and 0.95 μg/ml, respectively. This is the first report on anti-plasmodial activities of Baphia nitida, Tabernaemontana crassa and Treculia Africana. T. Africana showed moderate anti-plasmo- dial activity with IC50 value of 6.62 µg/mL. Extracts of P. clappertoniana, T. Africana and T. ivorensis (0.4 mg/mL) showed >50% anti- oxidant effect (SOD). The extracts were not cytotoxicity towards RBCs at the concentration tested (200 μg/ml) but were weakly cytotoxic to HL-60 cell. Selectivity indices of most of the extracts were greater than 10. Our results suggest that most of the plant extracts have strong anti-plasmodial activity and antioxidant activity which warrants further investigations. Keywords antioxidant, anti-plasmodial, malaria, medicinal plants, phenolic content Received February 2, 2021. Received revised October 8, 2021. Accepted for publication December 27, 2021. Introduction 1 Malaria is a blood parasitic disease caused by various plasmo- Noguchi Memorial Institute for Medical Research, College of Health Sciences, dium species; the predominant ones been P. falciparum and P. University of Ghana, Accra, Ghana2 Centre for Plant Medicine Research, Mampong-Akuapim, Ghana malariae. The burden of disease and mortalities are dispropor- 3 University of Ghana School of Pharmacy, College of Health Sciences, tionately high in low-income countries. According to the World University of Ghana, Accra, Ghana Health Organization (WHO), an estimated 228 million cases of malaria occurred worldwide in 2018, and nineteen countries in Corresponding Author: Regina Appiah-Opong, Department of Clinical Pathology, Noguchi Memorial sub-Saharan Africa and India accounted for about 85% of the 1 Institute for Medical Research, College of Health Sciences, University of Ghana,global malaria burden. Chemotherapy remains the main avail- P.O. Box LG 581, Legon, Ghana. able treatment regimen, as studies are yet to produce malaria Emails: rappiah-opong@noguchi.ug.edu.gh; ropong@gmail.com Creative Commons Non Commercial CC BY-NC: This article is distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0 License (https://creativecommons.org/licenses/by-nc/4.0/) which permits non-commercial use, reproduction and distribution of the work without further permission provided the original work is attributed as specified on the SAGE and Open Access page (https://us.sagepub.com/en-us/nam/open-access-at-sage). 2 Journal of Evidence-Based Integrative Medicine vaccine for protection against the parasite. The Plasmodium evidence on their medicinal properties. These medicinal plants are parasites like most microorganisms develop resistance over used by Ghanaian Traditional Medicine Practitioners to treat malaria periodic drug therapy. Parasites’ resistance to chloroquine and and other conditions. The plants were authenticated by Mr Heron recently arteminisins have been reported.2,3 The rst chloro- Blagogee, a Senior Botanist at the Center for Plant Medicinefi quine resistance to P. falciparum was recorded in 1978 in non- Research, Ghana, where Voucher specimens were deposited (Table 1). The plant parts were air-dried, pulverized and extracted immune travelers from Kenya and Tanzania, followed by a with 50% ethanol at room temperature for 24 h. The extraction proce- wide-spread resistance across the coastal inland areas and 3,4 dure was repeated three times and the supernatants were pooled. Themost parts of Africa by 1983. Quite recently, reports from ethanol content of the extracts were removed using a BUCHI® Asian countries including Cambodia, China, Myanmar and rotary evaporator. The remaining aqueous extracts were lyophilized Vietnam showed resistance to artemisinin, the current first-line (LABCONCO®, USA) to obtain dried extracts. drug used as antimalarial treatment regimen3 in several coun- tries including Ghana. Thus, drug resistance is a major chal- lenge to malaria control, hence it is imperative to search for Anti-Plasmodial Assay alternative and more potent antimalarials to replace the The chloroquine-sensitive Plasmodium falciparum strain 3D7 was cul- current drugs whenever drug resistance becomes widespread. tured in human red blood cells (RBCs) and RPMI 1640 culture Plant natural products have been enormous reservoirs for anti- medium. Subsequently, anti-plasmodial activity screening of the extracts malarial drug discovery; earlier successful efforts led to the dis- was performed using the SYBR green I-based fluorescence assay covery of the first antimalarial drug, quinine and, subsequently described by Kwansa-Bentum et al.14 The extract stock (50 mg/mL in chloroquine, mefloquine and artemisinin.5 The tropical flora 50% hydroethanol) solutions were diluted serially in a culture medium has an enormous diversity of plants and yet to be elucidated to obtain working concentrations in the range of 1 to 2000 μg/mL for natural products. Ghana, a tropical West African country is the assay. Aliquots of extracts were added sorbitol synchronized parasit- home to over 1000 medicinal plants, and more than 100 plants ized RBCs (ring stage) and incubated as earlier described, at 2% hemat- ocrit and 1% parasitaemia. Wells containing untreated parasites (with have been reported in ethnobotanical studies as therapies for tra- vehicle) were used as negative controls while the positive control exper- ditional malaria treatment.6,7 Previous studies have reported 8,9 iment consisted of parasite infected RBCs (iRBCs) treated with chloro-good anti-plasmodial activities in several plants. quine. The cultures were incubated for 72 h in a humidified chamber in Oxidative damage is a major pathological consequence of an incubator at 37 °C under low oxygen and carbon dioxide levels. malaria infections. Vital organs of the body are affected by oxi- Afterward lysis buffer containing 20 mM Tris-Cl (pH 7.5), 5 mM dative stress, resulting in changes such as hepatomegaly, sple- EDTA, 0.008% saponin, 0.08% triton-X 100 and 1× SYBR green I nomegaly as well as endothelial and cognitive damages. (10 000× in DMSO) was added to the wells, and the contents were Antimalarials currently available on the market often leave mixed gently and incubated in the dark at room temperature (26 °C) traces of these damages after treatment.10 Several medicinal for 1 h. Fluorescence in each well was read using a multi-well plate plants have been reported to have antioxidant properties.11–13 reader (Tecan Infinite M200, Austria) at excitation and emission wave- Phenolics are plant components that could have good antioxi- lengths of 497 and 530 nm, respectively. The fluorescence readings were used to calculate the percentage inhibition of parasite growth. The 50% dant properties. Therefore, we investigated the in vitro antima- inhibitory concentrations (IC50) of the extracts were extrapolated fromlarial and phenolic content of eight (8) medicinal plants in plots of percentage inhibition against extract/drug concentrations. All Ghana. We also assessed the effects of the extracts on superox- chemicals and reagents used for the study were purchased from ide dismutase enzyme (SOD) activity. The SODs are antioxi- Sigma-Aldrich Chemical Company (St. Louis MO, USA) and Gibco dant enzymes that are necessary for life. They convert BRL Life Technologies (Paisley, Scotland). superoxide radical into hydrogen peroxide and molecular oxygen, thus mitigating oxidative stress. Based on their tradi- tional uses and medicinal properties, the plants Cinnamomum Cytotoxicity Assay zeylanicum, Lippia multiflora Moldenke, Morinda lucida The 3-(4, 5-dimethylthiazol-2-yl)-5-diphenyltetrazolium bromide- Benth., Parkia clappertoniana Keay, Terminalia ivorensis based colorimetric selective assay (MTT-Based CSA) was used to A.Chev, Baphia nitida Lodd, Tabernaemontana crassa determine the cytotoxic effect of the extracts on uninfected red blood Benth. and Treculia Africana Decne. e×Trecul, were selected (RBCs) and human HL-60 cells as described.8,11 Briefly, 90 µL of for the study (Table 1). washed RBCs at 2% hematocrit were added to 10 μL aliquots of seri- ally diluted extract preparations (0-1250 μg/mL) in separate wells and HL-60 cells cultured in RPMI 1640 medium were also seeded at 1× Methods 105 cells per well into 96-well microtiter plates, mixed and incubated Plant Collection and Extract Preparation at 37 °C for 72 h. Positive and negative control experiments compris- ing chloroquine and vehicle-treated RBCs and untreated HL-60 cells The leaves of Morinda lucida and Lippia multiflora, roots of respectively were set up. Wells with extracts and culture medium Tabernaemontana crassa, stem-bark of Baphia nitida, Cinnamomum were also set up as color controls for extract-signal correction. After zeylanicum and Treculia Africana, and stem-bark and leaves of incubation for 72 h, 20 µL of 7.5 mg/mL MTT solution (in phosphate Parkia clappertoniana and Terminalia ivorensis were collected from buffer saline) was added to each well and the plate was re-incubated for Mampong-Akuapim in the Eastern region of Ghana by staff of the 2 h and 4 h, respectively for RBC and HL-60 experiments. Formazan Center for Plant Medicine Research, Ghana, based on anecdotal crystals formed from MTT were dissolved by adding acidified Appiah-Opong et al 3 Table 1. Ghanaian Medicinal Plants, Their Families, Parts Used and Voucher Specimen Numbers. Sample ID Plant name Family Part used Voucher specimen No. J48 Morinda lucida Rubiaceae Leaves JJNC008L J49 Parkia clappertoniana Leguminosae Stem-bark, Leaves JJNC050SBL J50 Tabernaemontana crassa Apocynaceae Root JJNC067R J51 Terminalia ivorensis Combretaceae Stem-bark, Leaves JJNCO48SBL J52 Baphia nitida Fabaceae Stem-bark JJNC040SB J53 Lippia multiflora Verbanaceae Leaves JJNC002L J54 Cinnamomum zeylanicum Lauraceae Stem-bark JJNC066SB J55 Treculia africana Moraceae Stem-bark JJNC033SB isopropanol and incubating the plates in the dark at room temperature Saponins (26 °C) overnight. The optical densities of the wells were measured at the wavelength of 570 nm using the multi-well plate reader. Triplicate Aliquots of 2 mL of distilled water were added to 1 mL of each extract experiments were performed. Curcumin was used as a positive control. and shaken vigorously. Observation of a stable and persistent The percentage RBC survival/cell viability in treated wells was calcu- (≥10 min) froth suggests the presence of saponins. lated and plotted against respective extract concentrations. The 50% cytotoxic concentrations (CC50) were determined by regression analy- sis. The selectivity indices (SI) of the extracts ie ratio CC /IC values Terpenoids50 50 were calculated. To a volume of 500 µL extract, 200 mL of chloroform was added, fol- lowed by the gentle addition of drops of concentrated H2SO4. The for- mation of an interface with a reddish-brown color suggests the Determination of Total Phenolic Content existence of terpenoids. Ursolic acid was used as the positive control. Generally, phenols have good antioxidant properties, therefore, the total phenolic content of the extracts was determined by the Tannins Folin-Ciocalteu’s method with slight modifications.13 A concentration of 1 mg/mL of each extract was prepared in distilled water. Gallic acid A volume of 1 mL extract was aliquoted into test tubes and brought to a standard solutions were prepared by serial dilution in the concentration boil at 100 °C. A few drops of 0.1% FeCl3 were added. A brownish- range 0.0156 - 1 mg/mL. A volume of 10 µL of each sample or gallic green or blue-black coloration indicates the presence of tannins. acid solution was aliquoted into the wells of a 24-well plate, in tripli- Gallic acid was used as the positive control. cates. Subsequently, 790 µL of distilled water and 50 µL of Folin-Ciocalteu reagent were added to each of the wells, mixed and incubated at room temperature for 8 min. One hundred and fifty micro- Flavonoids liters of Na2CO3 (20% w/v) were added and the mixtures were further A few drops of dilute NaOH solution were added to 500 µL of the incubated at room temperature for 2 h. Absorbance was read at a wave- extract. An intense yellow color indicated the presence of flavonoids. length of 750 nm. A gallic acid standard calibration curve was plotted. Quercetin was used as the positive control. The total phenolic content of extracts extrapolated from the plot was expressed in gallic acid equivalents (mg GAE/100 g extract). Alkaloids Superoxide Dismutase Assay To test for the presence of alkaloids, 200 µL of Wagner’s reagent (Iodo-potassium iodide) was added to 500 µL of the extract. The for- Effects of the plant extracts on superoxide dismutase (SOD) activity mation of a reddish-brown precipitate indicated the presence of alka- were evaluated as earlier described.15 Aliquots of 20 µL (80 µg/mL) loids. Quinidine was used as the positive control. of each sample were transferred into a 96-well plate, 200 µL 75 mM Tris-HCl buffer (pH 8.2) and 30 µL pyrogallol were added to each well. Corosolic acid was used as a positive control, and negative Statistical Analysis control was 50% hydroethanolic solution. Absorbance was read at The statistical significance between the means of the data was analyzed intervals of 0 and 5 min at the wavelength of 420 nm. The effect on using One-way analysis of variance (ANOVA) followed by Student’s SOD activity was expressed as percent inhibition of the rate of autoox- t-test. A ρ-value of less than 0.05 was considered statistically idation of pyrogallol as determined by the change in absorbance/min. significant. Phytochemical Tests Results Phytochemical tests for the presence of saponins, terpenoids, tannins, Anti-Plasmodial Activities of Plant Extracts alkaloids and flavonoids were carried out as described16,17 (Languon et al. 2018; Shah and Hussain, 2014). Each test was carried out Figure 1a illustrates the dose-response curve of the inhibitory using 10 mg/mL of extract. action of the medicinal plants on the Plasmodium falciparum 4 Journal of Evidence-Based Integrative Medicine Figure 1. Effects of plant extracts on (a) P. falciparum strain 3D7 infected RBCs, (b) uninfected RBCs and (c) HL-60 leukemia cells. Names of the plants are shown in Table 1. Table 2. Anti-Plasmodial Activity Cytotoxicity and Selectivity Indices The IC50 values were in the following increasing order: of Plant Extracts. J51200 95.650 >48.04 22.98 J49 1.133 >200 >200 >176.52 >176.52 extract (62.23 µg/mL). The IC50 value of positive control, chlo- J50 62.230 >200 110.100 >3.21 1.77 roquine was 5.5 ng/mL. There was a significant difference J51 0.956 >200 >200 >209.31 >209.31 between the IC50 values of the extracts and that of chloroquine J52 44.360 >200 >200 >4.51 >4.51 (ρ < 0.05). J53 3.036 >200 >200 >65.88 >65.88 J54 4.051 >200 nd >49.37 Nd J55 6.616 >200 95.520 >30.23 14.44 Cytotoxicities and Selectivity Indices of Plant Extracts CHQ 0.005 0.051 nd 98.04 Nd Cytotoxic effects of the extracts on RBCs are shown in the RBC SI>2 indicates good selectivity of a therapeutic agent; iRBC, represents RBCs survival curves (Figure 1b). All the extracts demonstrated a infected with Plasmodium falciparum strain 3D7. nd: Not determined. weak cytotoxic effect on the human RBCs at the highest extract concentration of 200 µg/mL, with percent cell survival strain 3D7. All the extracts showed concentration-dependent >50%. Similarly, the tested extracts showed weak or no cyto- inhibition of the parasite growth, with a majority of them toxic effects on the HL-60 leukemia cells (at 200 µg/mL) as showing moderate to strong anti-plasmodial action against the shown in Figure 1c. Curcumin, used at positive control for parasite with IC50 values in the range of 0.96 to 6.6 µg/mL. the cytotoxicity assay gave an IC50 value of 16.76 µg/mL. Appiah-Opong et al 5 Thus, generally, the extracts demonstrated weak cytotoxic Parkia clappertoniana, Tabernaemontana crassa, Terminalia action on both human cells tested (Table 2). ivorensis and Treculia Africana. We also evaluated the selec- Selective toxicities of the extracts on the parasites compared tive toxicity of the plants on human cells (uninfected RBCs to RBCs and HL-60 cells are shown in Table 2. Extracts J49 and and HL-60 cells) and their antioxidant properties. J51 with the strongest anti-plasmodial action demonstrated the Phytochemical constituents of the extracts were also highest parasite SI of 176.52 and 209.31, respectively for both investigated. RBCs and HL-60 cells. Recent studies outline stringent endpoint criteria for biolog- ical activity and selective activity. Philippe et al.18 described the anti-plasmodial effects of plant extracts as highly active (IC50≤ Total Phenolic Content of Extracts 5μg/mL); moderately active (5 < IC50≤15 μg/mL); weakly The total phenolic contents of the plant extracts are shown in active (15 50 μg/mL). Figure 2. Total phenolic content of the range 1287.1 to Anti-plasmodial activities of plant extracts have also been cat- 85 969.09 mg GAE/100 g extract were measured. Extract J51 egorized as follows: very active (<5 μg/mL), active (5-50 μg/ demonstrated the highest phenolic content of 85 969.0 mg mL), weakly active (50-100) and inactive (100). 19 Based on GAE/100 g extract whereas extract J48 demonstrated the both categorizations the plant species Cinnamomum zeylani- lowest total phenolic content of 1287.1 mg GAE/100 g extract. cum, Lippia multiflora, Morinda lucida, Parkia clappertoniana and Terminalia ivorensis showed high activity whereas Treculia Africana showed moderate activity. According to the Effect of Plant Extracts on SOD Activity categorization of Philippe et al.18 Baphia nitida is weakly Figure 3 shows the effect of the plant extracts on the activity of active and Tabernaemontana crassa is inactive. Parkia clap- the antioxidant enzyme SOD. Extract J53 caused the highest pertoniana and Terminalia ivorensis (each extracted from a increase in SOD activity which was comparable to the activity mixture of the stem, bark and leaves) showed the strongest of the positive control (CA). Each of the extracts caused a activities with IC50 values of 1.13 ug/mL and 0.95 μg/mL, ≥40% increase in enzyme activity compared to the control. respectively. Earlier studies have reported the anti-plasmodial activities of their parts on P. falciparum strain 3D7. The etha- nolic extract of the stem bark of T. ivorensis was reported as Results of Phytochemical Tests showing an IC50 value of 6.95 μg/mL and the aqueous extracts of the leaves showed the value of 0.64 μg/mL.9 The aqueous Qualitative tests were performed to determine the presence of leaf extract of T. ivorensis however gave an IC value of the phytochemicals saponins, terpenoids, tannins, flavonoids 50 10.52 μg/mL in chloroquine-resistant W2 strains.9 The and alkaloids in the eight plant extracts. Table 3 shows the phy- aqueous extract of the leaves of Parkia biglobosa a common tochemicals identified in the extracts. Extracts J49, J54 and J55 specie of Parkia genus has also been reported to have an IC contained all the five phytochemicals above. 50 value of 56.23 µg/mL while the phenolic fraction of the meth- anolic extract had the value of 0.51 µg/mL in P. falciparum iso- Discussion lates from malaria patients.20,21 Strong activities of the plant extracts in this study may be resulting from the composite activ- The anti-plasmodial effects of medicinal plants from Ghana 6–8 ities from the various components of the extracts. Althoughhave been reported in several studies. In this study, we inves- there was no correlation between the phenolic content of the tigated the in vitro anti-plasmodial action of hydroethanolic extracts of eight (8) plant species; Baphia nitida, Cinnamomum zeylanicum, Lippia multiflora, Morinda lucida, Figure 3. Effect of plant extracts on SOD activity. Eighty micrograms per milliliter (80 µg/mL) of each plant extract was tested. CA: Figure 2. Total phenolic content of plant extracts. Ten micrograms Corosolic acid; *, represents a significant difference between %SOD per milliliter of each plant extract were tested. of the positive control (CA) and the plant extracts (P≤ 0.001). 6 Journal of Evidence-Based Integrative Medicine Table 3. Phytochemical Constituents of Plant Extracts. comparable to the positive control. The antioxidant activity recorded could partly be attributable to the high total phenolic Phytochemical J48 J49 J50 J51 J52 J53 J54 J55 content of the extracts. Interestingly, T. ivorensis which had the Saponins + + + + + + + + highest phenolic content exhibited the strongest anti-plasmodial Terpenoids − + − − − − + + activity. Their antioxidant property could be beneficial for Tannins − + − + − + + + adjunctive therapeutic application in malaria management. Flavonoids + + − + + + + + The anti-plasmodial, antioxidant and cytotoxic effects of P. Alkaloids − + + + − − + + clappertoniana, T. ivorensis and T. africana extracts could +indicates present. partly be attributable to some of the identified chemical constit- −indicates absent. uents of the extracts. All the five phytochemicals tested, sapo- nins, terpenoids, tannins flavonoids and alkaloids were extracts and the anti-plasmodial activities, T. ivorensis and P. present in the three plant extracts except T. ivorensis in which clappertoniana which showed the strongest anti-plasmodial terpenoids were not detected. These findings are corroborated activities, had the highest phenolic content. Different plant con- by earlier reports that revealed the presence of alkaloids, anthra- stituents may act synergistically to improve their combinatorial quinones, flavonoids, glycosides, saponins, steroids, tannins effect. This effect may be due to certain complex formation and triterpenoids in P. clappertoniana; alkaloids, flavonoids, from the various constituents which elicits potent inhibition glycosides, saponins, tannins, and terpenes in T. ivorensis and than their individual effects.22 anthraquinone, cardiac glycosides, flavonoids, polyphenols This study also reports for the first time, the in vitro and saponins in T. africana.29–32 Further studies have eluci- anti-plasmodial activities of B. nitida, T. crassa and T. dated ivorenosides A, B and C as triterpene saponins in T. ivor- africana. T. africana showed moderate anti-plasmodial activity. ensis and, 6,9-dihydro-megastigmane-3-one, 4-hydroxybenzoic Studies have reported comparable anti-diabetic effects of the acid, α-pinene, myrtenal, limonene, camphene and n-hexanoic hydro-acetone extract of its root bark and hemagglutination acid in T. Africana.33,34 These natural product constituents inhibition of the seeds.23,24 However, our study reports the anti- could be investigated further for their anti-plasmodial effects. plasmodial activity of its stem-bark (IC50 value of 6.62 µg/mL). The selective action of the plants for the malaria parasite is an important indicator for the potency selection of medicinal Conclusion plants. It represents the ratio of cytotoxicity to biological activ- ity. Some other studies have described the selective index of This study has indicated a strong anti-plasmodial action and some medicinal plants as; low (4≤ SI<10), selective high selectivity of Terminalia ivorensis, Parkia clappertoniana (10>SI≤25), and highly selective (≥25).9,25,26 P. clappertoni- and Lippia multiflora extracts. We also report for the first time ana and T. ivorensis showed high selectivity towards the anti-plasmodial action of Baphia nitida, Tabernaemontana malaria parasite. This suggests that the extracts possess compo- crassa and Treculia Africana. The strong anti-plasmodial activ- nents with promising anti-plasmodial activity. The low toxicity ity of Terminalia ivorensis could partly be due to its high phe- of these plant extracts to human cells is consistent with earlier nolic content and the other phytochemical constituents. Further studies that reported that the aqueous seed extract of P. clapper- studies are warranted to isolate and characterize the active prin- tonian showed no observable maternal and developmental tox- ciples in the bioactive plant extracts. icities in Sprague-Dawley rats and ICR mice. On the other hand, the aqueous extracts of T. ivorensis demonstrated an SI≥3.48 against human umbilical vein endothelial cells,9 and Acknowledgments the aqueous root extract of T. africana showed no indicative The authors are grateful to the staff of CPMR, Mampong-Akwapim toxicity to the liver, heart and muscles enzymes in vivo.27 and Departments of Clinical Pathology and Parasitology, Noguchi Fifty percent cytotoxic concentration (CC ) of C. zeylanicum Memorial Institute for Medical Research, University of Ghana,50 Legon, Accra, Ghana, particularly Ms. Abigail Aning for their techni- in Jurkat cells was not determined due to limitation in the cal support. amount of plant extract. Antioxidant therapy in malarial infections remains to be fully rationalized. Studies have nonetheless reported its role as a viable Author Contributions therapeutic strategy for alleviating plasmodium-induced oxidative 10 RAO and AKN conceived of the study and participated in its designstress and its associated complications. The increased metabolic and coordination. KA, PA, KB-AO and ED carried out the experimen- rate of the rapidly growing and multiplying parasites leads to the tal studies and data analysis. MMS, RA, FA and AAA worked on the generation of large quantities of redox-active by-products, and, plant samples. KA drafted the manuscript. All authors reviewed the consequential imbalance in the oxidant-antioxidant system. The manuscript. changes have been observed in children with severe malaria and they have been associated with P. vivax malaria.28 All the plants studied showed various antioxidant potentials. Lippia multiflora Data Availability leaf extract exhibited a stronger SOD inductive effect which was Data on this research will be made available on request. Appiah-Opong et al 7 Declaration of Conflicting Interests 9. Komlaga G, Cojean S, Dickson RA, et al. Antiplasmodial activity The author(s) declared no potential conflicts of interest with respect to of selected medicinal plants used to treat malaria in Ghana. the research, authorship, and/or publication of this article. Parasitol Res. 2016;115(8):3185-3195. http://doi:10.1007/ s00436-016-5080-8 Ethical Approval 10. Isah MB, Ibrahim MA. The role of antioxidants treatment on the pathogenesis of malarial infections: a review. Parasitol Res. Ethical approval for the study was obtained from the Institutional 2014;113(3):801-809. http://doi:10.1007/s00436-014-3804-1 Review Board of Noguchi Memorial Institute for Medical Research, College of Health Sciences, University of Ghana (NMIMR-IRB 11. Appiah-Opong R, Asante IK, Osei Safo D, et al. Cytotoxic effects CPN 001/12-13 Revd 2017). Informed consent was sought from vol- of Albizia zygia (DC) J.F. Macbr, a Ghanaian medicinal plant unteers who donated blood samples that were used to culture the against T-lymphoblast-like leukemia, prostate and breast cancer malaria parasite. cell lines. Int J Pharm Pharm Sci. 2016;8(5):392-396. https:// innovareacademics.in/journals/index.php/ijpps/article/view/10656 Funding 12. Acheampong F, Larbie C, Appiah-Opong R, Arthur F, Tuffour I. In vitro antioxidant and anticancer properties of hydroethanolic The author(s) disclosed receipt of the following financial support for extracts and fractions of Ageratum conyzoides. Eur J Med the research, authorship, and/or publication of this article: This work was supported by the Noguchi Memorial Institute for Medical Plants. 2015;7(4):205-214. http://doi:10.9734/EJMP/2015/17088 Research, University of Ghana and Science and Technology 13. Anim MT, Larbie C, Appiah-Opong C, Tuffour I, Owusu KB-O, Research Partnership for Sustainable Development (SATREPS), Japan Aning A. Extracts of Codiaeum variegatum (L.) A. Juss is cyto- toxic on human leukemic. Breast and prostate cancer cell lines. ORCID iDs J Appl Pharm Sci. 2016;6(11):087-093. 14. Kwansa-Bentum B, Agyeman K, Larbi-Akor J, Anyigba C, Regina Appiah-Opong https://orcid.org/0000-0003-4219-7107 Appiah-Opong R. In vitro assessment of antiplasmodial activity Frederick Ayertey https://orcid.org/0000-0003-1691-5646 and cytotoxicity of Polyalthia longifolia leaf extracts on Plasmodium falciparum strain NF54. Mal Res Treat. 2019. References http://doi.org/10.1155/2019/6976298 1. WHO, The World malaria report 2019. Accessed February 2, 15. Alam MN, Bristi NJ, Rafiquzzaman M. Review on in vivo and in 2021. www.who.int/news-room/feature-stories/detail/world-malaria- vitromethods of evaluation of antioxidant activity. Saudi Pharm J. report-2019; 2019. 2013;21(2):143-152. 2. 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