Geographically, Bengkulu Province is located at 2°16’-3°31’ N and 101°01’-103°41’ E, with a maximum temperature of 32.9-34 °C and a minimum temperature of 22-23 °C (Badan Pusat Statistik 2017). Bengkulu Province is located along the west coast of Sumatra Island for approximately 525 km.

Data on medicinal plants used for antimalarial were obtained from results of RISTOJA conducted in 2012. Source of the data were 36 respondents from 7 ethnic groups in Bengkulu Province, namely Serawai, Lembak, Rejang, Muko-muko, Pasemah, Pekal and Enggano.

Number of sample points were determined based on populations of ethnic groups having at least 1,000 members. Six of the 7 ethnic groups in Bengkulu Province presumably have antimalarial concoctions, i.e., Serawai, Lembak, Rejang, Muko- muko, Pasemah, and Enggano (Figure 1).

Respondents were traditional healers from local community who utilized medicinal plants to treat diseases. Those respondents were selected by means of purposive sampling approach based on information from indigenous community leaders, village heads, staff of government health facilities, and local health office.

Method used for data collection in this study is in accordance with the guidelines from 2012 RISTOJA (Indonesia, 2012); (Syafni & Bakhtiar, 2022). Of the 7 ethnic groups that became respondents, only 6 ethnic groups were selected because these 6 ethnic groups utilize medicinal plants to treat malaria.

In this study, all medicinal plants used as antimalarial concoctions by the respondents were inventoried, recorded, and collected. The collected specimens were identified and stored in the Herbarium Tawangmanguensis of the Ministry of Health of the Republic of Indonesia.

Prior to the interview session, the study objectives and methods were explained to the respondents. Informed consent was also obtained and practices related to the recognition, control, and treatment of malaria.

Data processing was started with initial cleaning of the data from errors, followed by grouping the data based on similar knowledge. After the grouping, data were analyzed to determine UV (use value), FL (fidelity level), FUV (family use value), and RFC (relative frequency of citation).

Use value was calculated to figure out the importance of each medicinal plant for each traditional healer. The formula to calculate use value was the following (Najem et al., 2020); (, 2023):

U = number of use reports cited by each respondent for a medicinal plant species

n = total number of respondents interviewed for a medicinal plant species

Fidelity level was determined to analyze the importance of medicinal plants for a given remedy, which was calculated by using the following formula (Najem et al., 2020); (Asiimwe et al., 2021):

Ip = number of respondents who independently reported the use of a specific medicinal plant for the same illness

Iu = overall number of respondents who cited the medicinal plant for any illnesses in general

Family use value was determined to identify the significance of medicinal plant families and was calculated by the following formula (Najem et al., 2020):

UVs = use value of medicinal plant species belonging to the same family

Ns =   total number of medicinal plant species present in each family

Relative frequency of citation was determined to show the local importance of each medicinal plant species, and it was calculated by the following formula (Najem et al., 2020); (Asiimwe et al., 2021):

where:

FC = number of respondents using a given medicinal plant species

N =  total number of interviewed respondents

Plant part value was determined to show the most used plant parts by the respondents (Najem et al., 2020); (Biara et al., 2021); (, 2023). The highest plant part value indicates the most used medicinal plant parts by the respondents and was calculated by using the following formula (Najem et al., 2020); (Biara et al., 2021); (, 2023):

where:

RU = number of uses reported by the respondents for all medicinal plant parts

RU Plant Part = the sum of uses reported by the respondents for each part of the medicinal plant.

To explain knowledge differences on antimalarial medicinal plants in each ethnic group, a cluster dendrogram was established by using R-4.3.2.

Twenty traditional healers from 6 ethnic groups in Bengkulu Province who prescribed antimalarial concoctions made from medicinal plants were selected as respondents in this study (Table 1 ).

In our study, traditional healers who prescribed antimalarial concoctions in Bengkulu Province were found in 6 ethnic groups, i.e., Enggano, Muko-muko, Rejang, Lembak, Serawai, and Pasemah. Traditional healers who prescribed the highest number of antimalarial concoctions were from Pasemah ethnic group, which is related to a larger coverage area, namely Pagaralam, Empat Lawang, Lahat Regency, Ogan Komering Ulu, the Muara Enim area, and Mount Dempo, an active volcanic area. From their study, (Huda et al., 2019) stated that the long distance from the residential area to health facilities encouraged people to dwell into the knowledge of utilizing medicinal plants in overcoming health problems.

Male gender dominated the respondents in our study (12 of 20 respondents). The dominance of male gender as traditional healer is related to people’s perception in requirements of being a traditional healer, one of which is being a male descent of a traditional healer (Djamaluddin et al., 2020), because the medicinal skill should be passed down from their ancestors. The respondents in our study were mostly in lower educational level.

In regard to age, 16 of 20 traditional healers were at the age of 50 or above and one healer even reached the age of 80. This indicated that medicinal profession have not been the interest of younger generations in concurrent with several obstacles that hinder the regeneration efforts by the older traditional healers. Transfer knowledge on medicine requires special skills and basic medical knowledge, which is not easily transferred to potential successors. There are basic requirements for someone to become a traditional healer. A study conducted by (Efrianto, 2018) showed that not everyone has the ability to become traditional healer. (Efrianto, 2018) also mentioned that there are four ways to pass on knowledge, namely learning, inspiration, dreaming and meditating.

Monthly number of patients received by respondents in our study were mostly in the range of 0-25 (11 of 20 respondents). Some respondents even received more than 100 patients per month (4 of 20 respondents). However, our respondents did not keep notes on which of their patients were prescribed antimalarial concoctions.

Our study also showed that most village communities have a good knowledge on traditional medicine and a lot of interests in using traditional medicine. The knowledge and interests were mostly inherited by their families. Knowledge is a factor influencing the use of traditional medicine. Lack of knowledge on health care and modern medicine has resulted in the use of herbal medicine as one of treatments for various diseases (Adiyasa & Meiyanti, 2021).

Our study showed that not all traditional healers have antimalarial prescriptions. In Bengkulu Province, the 6 ethnic groups claimed to have medicinal plants for treating malaria. Traditional healers diagnose a patient as having malaria based on fever symptoms and testimony of the patient. There were 47 medicinal plant species belonging to 29 plant families used for antimalarial medication (Table 2 ). Fifteen plant specimens could not be identified by the scientific names.

Our study showed that C. papaya, P. canescens, and T. crispa had UV, RFC, and FL values of 0.15, 0.15, and 15.00, respectively. Ethno medicinal data were quantitatively analyzed using RFC, indicating the local importance of medicinal plant species. The species having the highest RFC (C. papaya, P. canescens, and T. crispa) were the species most often used by traditional healers in Bengkulu Province to cure malaria, because these plants were easy to find and easy to cultivate. People believe that the bitter taste of C. papaya, P. canescens, and

T. crispa leaves can be used for treating malaria (Shinta, 2005); (Taek, 2020).

Several research results showed that C. papaya, P. canescens, and T. crispa leaves have pharmacological properties as antimalarial medicine. Papaya extract contains a total phenolics of 27.99±1.46 mg GAE/g and a total flavonoids of 18.24±1.36 mg QE/g, which prevent the immune system from destroying platelets, decrease the virus-induced bone marrow damage, and stabilize infected cell membranes, leading to the acceleration of natural healing process by boosting platelets count and averting problems (Babalola et al., 2024). T. crispa contains an active compound called tinocrisposide which works in the erythrocytic phase by inhibiting the growth of parasites in erythrocytes; therefore T. crispa can be an alternative medicine for treating malaria (Abdillah et al., 2015); (Malik, 2015). The ethanol fraction of P. canescens leaves at a dose of 0.084 g/kgBW is the most effective dose for treating malaria and has potential as an antimalarial medicine (Prasiwi et al., 2018).

The fidelity level (FL) represents the proportion of traditional healers who reported utilizing specific plant species to treat a particular sickness in the research area. The highest fidelity level

value for a particular plant species signifies its critical importance and frequent application as a therapeutic agent ( (Padhan & Panda, 2016); (Chaachouay et al., 2019))informant consensus factors (FIC. In our study, C. papaya, P. canescens, and T. crispa had the highest fidelity level (15%), indicating that traditional healers often use these plants as antimalarial medicine in Bengkulu Province.

C. papaya contains many advantageous chemical components in its fruits, leaves, flowers, stems, bark, and roots, such as papain, chymopapain, cystatin, à-tocopherol, ascorbic acid, flavonoids, cyanogenic glucosides, and glucosinolates. Leaf extract of C. papaya were demonstrated to possess antimalarial properties by effectively eliminating Plasmodium falciparum larvae within a concentration range of 25 ug/mL to 150 ug/mL (, 2012). P. canescens leaves contain phytochemical compounds, including flavonoids, phenolics, terpenoids, and alkaloids, which act as antioxidants, antibacterial, anti-inflammatory, antidiabetic, and antimalarial agents (Maigoda et al., 2022) ethanol extract extracted by maceration method using 96% ethanol as solvent from P. canescens. The Dragendorff’s and Mayer test carried out the qualitative phytochemical analysis, FeCl3 test, Salkowski method, Liebermann– Burchard method, foam test, and NaOH reagent. The total phenolic and flavonoid levels were tested using the Folin–Ciocalteu method. In vitro antioxidant activity was carried out using the DPPH (1,1-diphenyl-2-picrylhydrazyl.

In their study, (Suwandi et al., 2018) showed that the aqueous extract from P. canescens leaves had antispasmodic properties which effectively reduced the development of Plasmodium falciparum with an IC50 value of 12.26±1.05 μg/mL. T. crispa plant has been extensively utilized by traditional healers in Bengkulu Province for its antimalarial properties. Research conducted by Niljan et al. (2014) indicated that extract of T. crispa exhibits antimalarial properties in mice infected with Plasmodium berghei, while combining the extract of T. crispa with pyrimethamine enhances the efficacy of T. crispa as antimalarial medicine.

The family use value of medicinal plants in our study showed that both Lamiaceae and Caricaceae had the highest FUV (0.15), while Menispermaceae had the second highest value of 0.13 (Figure 2 ).

Lamiaceae had long been used as traditional medication for treating malaria. Various studies stated that Lamiaceae family, previously referred to as Labiatae, contains several active compounds that act as antimalarials, such as abietane, diterpenes, diterpenoids, triterpenoids, and flavonoids (Tjitraresmi et al., 2020). Caricaceae family, especially C. papaya, plays a role in producing alkaloids having larvicidal activity against Anopheles stephensi mosquitoes (Singh et al., 2022). Alkaloids produced by various plants are known as antimalarial medication (Omagha et al., 2020). Several species of Menispermaceae family, such as Arcangelisia flava (Nain et al., 2022); (Pratama et al., 2023) and Tinopsora crispa (Fitri et al., 2019); (Merici et al., 2020), contain alkaloids in the form of berberine, which function as antimalarial medication.

Herbal materials used by traditional healers consist of various parts of medicinal plants ranging from leaves, flowers, stems, bark, and roots (Figure 3).

Leaves (43.9%) and barks (13.6%) were the most frequent used parts of medicinal plants for treating malaria. Leaves are mostly used in herbal preparations by traditional healers because leaves are easily recognized, taken, and utilized part of medicinal plants. Leaves can be harvested at any time without depending on weather conditions and are the easiest plant part to process or mix as ingredients for herbal preparations (Widiyastuti et al., 2017) forest areas are still the main habitat of medicinal plants germplasm. Sigogor Nature Reserve in Ponorogo Regency, East Java, is a potential area that has a diversity of medicinal plants and has not been widely studied. The exploration of medicinal plants in the Sigogor Nature Reserve area aims to know the diversity, abundance and knowledge of the surrounding community about the existence of these medicinal plants. The method used is explorative survey with qualitative approach. The data collected in the form of secondary data and primary field observation results and literature studies of previous research results. Data analysis was carried out descriptively for the identification of medicinal plant specimens, qualitative vegetation analysis to determine plant habitus type, than percentage of habitus type and plant part was calculated based on observation result. The results of exploration activities show there were 43 species of medicinal plants from 33 families have been found in the sorrounding area of Sigogor. Habitus of medicinal plants found mostly were herbs (39.5%.

In utilizing plant parts for herbal preparations, it is important to consider the conservation aspect of medicinal plants. The use of bark, stems, and roots in herbal preparations can have a long impact on population decrease of medicinal plants, because the harvesting process of those plant parts will include the process of injuring the plants, leading to the plant death and eventually, the extinction of the plants (Bamigboye & Tshisikhawe, 2020).

Medicinal plants used in preparing antimalarial medication in Bengkulu Province came from five different sources, i.e., from the wild (38%), garden (30%), forest (28%), seashore (3%), and market (1%) (Figure 4). Traditional healers mostly (69%) utilize medicinal plant from nature, either from the wilds or harvested from a forest or from a seashore.

Natural conditions of Bengkulu Province are relatively pristine, not much impacted by industrial and residential areas; therefore, allowing the community to obtain medicinal plants from the wilds of the surrounding environment (, 2022). The ease of obtaining medicinal plants both from the wilds (69%) and from gardens (30%) has resulted in low sales of medicinal plants in markets (1%)Figure 4, showing that sources from ‘Garden’ and ‘Forest’ provide a significant contribution, while sources from ‘Seashore’ and ‘Market’ ‘ are the least utilized, indicating that local traditions and knowledge play as the primary roles in supporting local health care (Balinado & Chan, 2017). Only 30% of the raw materials used by traditional healers came from medicinal plants planted or naturally grown in their gardens, while the remaining medicinal plants came from wild plants found in the forest, primarily trees that are challenging to cultivate (Dajic-Stevanovic & Pljevljakusic, 2015).

The preference of utilizing medicinal plants from the wilds over those cultivated ones is related to the concentration of bioactive chemicals produced by the medicinal plants that may be impacted by environmental factors associating with the therapeutic process (Pandey et al., 2023).

In regard to the use of medicinal plants for treating malaria, each traditional healer has different ingredients and methods compared to other ethnic groups. Peculiarities in the utilization of these medicinal plants are greatly influenced by culture, educational background, availability of public health facilities, experiences of traditional healers and availability of medicinal plants resources in residential region of traditional healers ( (Gaoue et al., 2017); (Cain et al., 2018); (McQuaid & Landier, 2018); (Purwoko et al., 2023)).

Our study established a comprehensive overview of different herbal preparations used for antimalarial medication, ingredients specifications, preparation methods, dosage, usage frequency and treatment duration for antimalarial treatment conducted by traditional healers from 6 ethnic groups in Bengkulu Province (Table 3 ).

Usage frequencies varied from once a day to 10 times daily or frequently, and the treatment durations ranged from one day to until recovered. The most common usage frequencies for antimalarial medication were 3 times a day and once daily (Table 3 ), indicating how often traditional healers administered specific antimalarial medication in Bengkulu Province. There were also variations in the usage frequencies of antimalarial therapies based on the type of treatment, which were primarily prescribed to be taken once, twice, or three times per day for a fixed number of days, such as 3 days, 7 days, or a month. Less expected frequencies included half a glass a day or 10 times a day. Some treatments were less frequent, like once a day, once a night, or a few times per month for an extended period, such as 3 months, and there were a few treatments specifying

to take the medication frequently or as needed until symptoms resolve. The usage frequency of prescription made from medicinal plants for antimalarial medication can vary depending on the type of medicinal plants and how it is processed ((Caunca & Balinado, 2021); (Hastiana et al., 2023)). The misuse of medicinal plants, whether excessive consumption of concoction or in combination with other medicinal plants that are not compatible can cause side effects ((Alkhamaiseh & Aljofan, 2020); (Başaran et al., 2022)).

Preparation of traditional antimalarial medications by traditional healers in Bengkulu Province can be categorized as internal and external usages ( Figure 5). For internal usage, the herbal preparations for antimalarial medication were made in the form of decoction, infusion, dry powder in capsules, and raw consumption.

The differences in treatments and medicinal recipes were based on the experience and knowledge of traditional healers learned from their ancestors. This study showed that the most widely used methods in traditional medication practiced by traditional healers in Bengkulu Province were decoction and infusion ( Figure 5).

Our study also showed that the use of medicinal plants in the form of a decoction was most preferred considering that this method provided greater extraction yields and higher concentration of active phytochemicals; thus, potentially increased the efficacy of herbal preparations. In processing medicinal plants, it is important to apply safe heating limits when boiling the medicinal plants, so as not to destroy the active ingredients contained in the plants. Destruction of active ingredients during boiling using high temperature occurs because some secondary metabolite compounds contained in the boiled medicinal plants have thermolabile properties. In addition, decoction is the oldest and simplest way of preparing traditional medicine, whether using fresh or dried ingredients ((Balinado & Chan, 2017); (Caunca & Balinado, 2021); Mir et al. 2021; Sujana et al. 2021). Infusion is preferred for preparing antimalarial medication, along with other traditional medications which use ingredients having bioactive contents that are easily degraded and easily soluble (Petrakou et al. 2020) . Traditional medications used by ethnic groups are generally in the form of decoctions, infusions, raw foods, poultices, powders, and others (Boyzel et al. 2019) .

Plants selected for treating malaria traditionally were those having bitter taste. Several research revealed that some plants have been proven to have antimalarial activity; for instant quinine from Cinchona spp. and artemisinin from Artemisia annua ((Nonaka et al., 2018); Tajuddeen & Van Heerden 2019). Our study investigated almost all of 11 medicinal plants to determine their antimalarial activity and the content of major phytochemical compound, except for Boehmeria grandis, in the Pasemah ethnic group (Table 3 ; (Indonesia, 2013)).

Empirical facts about B. grandis can be used as an initial information for conducting research on the potential B. grandis in controlling malaria. Further research on P. canescens, J. gendarussa, and M. umbellata are still required because their extracts are still in crude form, and the main bioactive compounds that act as antimalarial medication are not yet known.

Several studies had determined bioactive compounds of 7 medicinal plants, namely C. papaya (carpaine) (Arifuddin et al., 2019), T. crispa (13 hydroperoxyoctadeca 9, 11 dienoic acid (13[S] HPODE), Columbine, and berberine),

A. paniculata (andrographolide), C. longa (curcumin), P. angulata (physalin) (, 2021), Z. officinale (gingerenone), and F. tinctoria (berberine).

Each of these bioactive compounds has its mechanism and role as an antimalarial medication. Although the antimalarial mechanism is unknown, it is estimated that carpaine directly inhibits the parasite (Arifuddin et al., 2019); (, 2019); (Dwivedi et al., 2020); (Haldar et al., 2020). At the tests of 13 hydroperoxyoctadeca 9, 11 dienoic acid (13[S] HPODE), the reduction in parasitemia occurs suppressive, prophylactically, and shows a schizonticidal effect. Columbine and berberine have similar mechanisms, working as an antimalarial medication using a dihydrofolate reductase-thymidylate synthase (DHFR- TS) inhibitory mechanism ( (Chester et al., 2017); (Bare et al., 2020); (Purwaningsih et al., 2023)) hypertension, atherosclerosis, and cancer. For decades, anti-adipogenic potential of many herbal extracts has been investigated. One example is Garcinia cambogia extract (GE. The mechanism of andrographolide is to inhibit the growth of parasitemia (Sari et al., 2018). Curcumin has several mechanisms

of action, including anti-inflammatory, antioxidant, antinociceptive, antiparasitic, and wound-healing effects ((Urošević et al., 2022); (Jamil et al., 2023)). Physalin works by inhibiting the function of the P2X7 receptor, which regulates the production of pro-inflammatory cytokines (, 2021) Gingerenone works as an antimalarial medication by binding to plasmepsin II and Plasmodium falciparum dihydrofolate reductase-thymidylate synthase (, 2023).

In Bengkulu Province the 20 traditional healers, who were selected as respondents in this study, had diverse traditional knowledge on malaria treatment. This study developed a dendrogram to represent four main clusters, where one cluster (Pasemah7) has no similarity to the other clusters ( Figure 6).

Traditional healers in Pasemah ethnic group (Pasemah7) had a formula containing 14 medicinal plants having antimalarial activity that differed from the other healers, i.e., A. lavenia, A. ghaesembilla, M. tanarius, P. acerifolium, P. javanicum, S. cordifolia, S. rhombifolia, T. populnea, A. elasticus, M. paradisiaca, G. verticillata, G. apus, O. sativa, and C. halicacabum. One traditional healer from Enggano ethnic group (Enggano-1) had a formula using 4 different medicinal plants having antimalarial activity, namely A. calamus, S. taccada, F. tinctoria, and C. zanthorrhiza.

Our study observed that Pasemah was the ethnic group with the highest usage of medicinal plants for treating malaria. Knowledge on medicinal plants acquired by the Pasemah ethnic group may have been caused by local wisdom, culture, beliefs, and accessibility inherited within the ethnic group (Adi et al., 2020); (, 2023).

Pasemah ethnic group is widely spread in Pagar Alam City, Empat Lawang Regency, Lahat Regency, and Muara Enim Regency (Ernatip & Ajisman, 2007), where malaria cases are high, especially in Muara Enim Regency in which malaria cases are the highest in South Sumatra, with incidence rates of 3,428 (in 2018), 3,215 (in 2019), and 6,266

(in 2020) ((B.P.S., 2023); (, 2023)). In Bengkulu Province, there are open-pit mineral mining areas causing the occurrence of lots of standing water, which supports the development of malaria vector mosquitoes.