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Journal of Parasitic Diseases: Official Organ of the Indian Society for Parasitology logoLink to Journal of Parasitic Diseases: Official Organ of the Indian Society for Parasitology
. 2025 Mar 6;49(3):735–746. doi: 10.1007/s12639-025-01795-2

Bioactive profiling, larvicidal and pupicidal activities of Crateva adansonii against two developmental stages of malaria vector, Anopheles gambiae

Kayode David Ileke 1, Naheemot Atinuke Babatunde 1, Adebayo Victor Akeju 2,✉
PMCID: PMC12399485  PMID: 40901424

Abstract

Mosquito-borne diseases continue to pose significant public health challenges globally. Synthetic insecticides, raise concerns regarding their environmental impact and human health risks. This study evaluates the effectiveness of Crateva adansonii crude extracts against malaria vector Anopheles gambiae. The leaf, stem and root were extracted with ethanol, GC-MS analysis was done and a concentration of 1%, 5%, 10%, 15% and 20% was used for the bioassay. Mortality data was recoded, and the lethal concentrations were estimated. Bioactive profiling of the crude extracts revealed the presence of hexadecanoic acid, 1-(2-cyclohexyliminocyclopentyl) ethanone, heptadecanoic acid, N-(2-amino-2-oxoethyl) tetradec-2-ynamide and octadecanoic acid for leaf extract. Presence of tetradecane, hexadecane, octadecane, 5-methoxyquinoline-6-carbonitrile, 1-methoxy-3-methylsulfanylbenzene, and Octadecanoic acid was revealed for bark extracts. Additionally, presence of (E)-octadec-6-enoic acid, diethyl bis(trimethylsilyl) silicate, 1-methyl-4-phenyl-5-sulfanylidene-1,2,4-triazolidin-3-one and 3-(furan-2-yl)-4-prop-2-enyl-1 H-1,2,4-triazole-5-thione was revealed for bark extracts. Larvicidal activity recorded 100% larval mortality at 15% and 20% concentrations within 12, 18, and 24 h for the root extracts. 20% concentration of the leaf extract recorded 100% mortality in 6 h. The root extract recorded the lowest lethal concentration against the larvae at 0.04% and 8.07% for LC50 and LC90 respectively at 24 h. Similarly, complete mortality was observed in the leaf and bark extract at 20% concentration in 6 h with the lowest LC50 and LC90 being 1.78 and 14.75 respectively. Ethanolic extract of C. adansonii plant exhibited strong insecticidal activities against malaria vector. Use of plant product as plant-based mosquito control agent will help in reducing the eco-toxic effects of synthetic chemicals.

Keywords: Crateva adansonii, Bioactive profiling, Malaria vector, Larvicidal, Pupicidal

Introduction

Anopheles mosquitoes are unquestionably one of the most medically important vectors of disease pathogens. They transmit pathogens and parasites that are of significant medical importance such as bacteria, nematodes, protozoans, and viruses that are responsible for deadly human diseases such as Lymphatic Filariasis and Malaria (Caraballo 2014). They endanger about four billion people, or over half of the global population, in tropical and subtropical regions, and have also had a significant impact on mankind’s development, not only socioeconomically but also politically (WHO 2017). According to WHO (2023), in 2022, 249 million cases of malaria with 608,000 deaths were reported while 244 million cases with 610,000 deaths were reported in 2021. Plasmodium falciparium, which is the causal agent of malaria in Africa is vectored by A. gambiae, therefore, An. gambiae is mostly referred to the African malaria vector. Anopheles mosquitos require a blood meal preferably, human blood meal for reproduction, their eggs develop into larvae in stagnant water bodies from anthropogenic activities such as irrigation for agriculture, flooded human settlements among others while their adults are found mostly in human settlements. Complex of An. gambiae comprises of six named species which include: An. arabiensis, An. melas, An. merus, An. bwanbae, An. gambiae sensu stricto, An. quadriannulatus, one unnamed species and several ‘incipient’ species (Coetzee et al. 2013).

The management of mosquito populations is very critical in reducing disease transmission and these control method ranges from biological control to chemical control and mechanical control. Chemical control is the most widely used because of their rapid effectiveness and simple application that have led to successful outcomes (Meier et al. 2022). The use of chemical insecticides has been shown to be degrading to the environment and harm non-target species and human health (Abdel-Tawab 2016). Each year, over 200,000 individuals die due to exposure to and misuse of synthetic chemicals (Tudi et al. 2022). Besides the ecological concerns associated with synthetic insecticide use, there is also the challenge of insect vector resurgence and resistance to insecticides (Ojo 2016). To address these challenges, it is imperative to explore alternative methods for managing insect vectors. Studies have consistently demonstrated that plant derivatives are effective in controlling mosquitoes while posing minimal risk to the environment (Demirak and Canpolat 2022; Muhammed et al. 2022; Ahmad et al. 2023; Botina and Martins 2024).

Crateva adansonii belongs to the family Capparidaceae and it is a medicinal plant commonly known as sacred garlic pear tree, temple plant, Caper tree, three leaf caper. It is locally called “Eegun orun” or “ajanaka” among the Yorubas in western Nigeria, or “Ungududu” among the Hausa in northern Nigeria and “Amakarode” among the Igbos in Eastern Nigeria). It is a small tree that can grow between 3 and 10 m tall, with three-lobed leaves. It has white flowers which appear when all the leaveshas fallen off the tree. Its fruit is edible, has a spherical shape and it is about 3.5–5 mm in size (Akhtar and Isman 2000). Different parts of Crateva adansonii plant are used as medicine and food traditionally. The leaves are used as vegetable in soups or sauces and can be used to relieve joint pains when applied externally; infection of the eye, earache and toothache are also treated with the fresh juice from the leaves (Abudullahi et al. 2012). Urinary tract infections, renal and vesical calculi and burning micturition are also treated with the bark (Arbonnier 2004). Grinding the bark into powder is also useful for treating rheumatism, asthma, itch, and epilepsy (Sivarajan and Balachandran 1994). Betulinic acid, diosgenin, lupeol, friedelin, and ceryl alcohol are all produced from the bark. Bark paste is used topically to swollen areas (Burkill 1990).

Crateva adansonii plant contain a range of chemicals that have demonstrated effectiveness against insects. Its essential oil has been employed in various methods to manage storage pests of maize and cowpea (Sanyaolu et al. 2020). No study has been published on the chemical composition of C. adansonii leaves, stem bark and root and their activities of against An. gambiae developmental stages. Hence, it becomes imperative to evaluate the phytochemical content and larvicidal activities of C. adansonii extracts against Anopheles gambiae.

Methodology

Collection and identification of plant materials

Fresh Crateva adansonii leaves, stem and root without defects was sourced from Iseyin, Oyo State, Nigeria and they were identified by Botanist at the Biology Department, Federal University of Technology, Akure, Ondo State, Nigeria. The plants materials were properly cleaned with distilled water, let to air dry for a month in the lab (Fig. 1a–d), and then ground with a pestle and mortar into a fine powder. The finished powder was kept for preservation purposes in dark bottles that are refrigerated until the extraction process was completed.

Fig. 1.

Fig. 1

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aCrateva adansonii plant. bCrateva adansonii Leave. cCrateva adansonii Stem Bark and Leave. dCrateva adansonii Root

Plant extraction and concentration preparation

This study employed a modified version of Udo (2011) cold maceration extraction method, in which 500 g of Crateva Adansonii powder was diluted with 1.5 L of absolute ethanol in an extraction bottle which was gently stirred with a glass rod for a period of three days. A rotary evaporator operating at a speed of 3 to 6 rpm, under controlled temperatures of 30 to 40 °C for a period of 8 h was used to concentrate the liquid filtrate. The concentrated extract was refrigerated at 4 °C until needed. The plant extracts were prepared at various concentrations following the method outlined by Simon-Oke and Akeju (2019). Different concentrations of 1%, 5%, 10%, 15% and 20% was made by diluting 0.1 ml, 0.5 ml, 1 ml, 1.5 ml, 2 ml of the crude extracts in 9.9 ml, 9.5 ml, 9 ml, 8.5 ml, 8.0 ml of sterile distilled water respectively. These concentrations were used for the larvicidal bioassay. The larvicidal bioassay were performed at an optimal room temperature of 26 ± 2 °C with a relative humidity of 80 ± 2% and a photoperiod consisting of 12 h of light followed by 12 h of darkness (12 L:12D).

Profiling of the compounds present in the leaves, stem and root of Crateva adansonii

The chemical compounds present in Crateva adansonii was identified through Gas Chromatography-Mass Spectrometry analysis (GC-MS). This analysis was performed with a gilent 7890 A gas chromatograph system (Agilent 19091-433HP, USA) coupled with a mass spectrometer. The system was equipped with an HP-5 MS fused silica column (5% phenyl methyl siloxane, 30.0 m × 250 μm, film thickness 0.25 μm) and interfaced with a 5675 C Inert MSD fitted with a Triple-Axis detector. Helium gas was employed as the carrier gas at a column flow rate of 1.0 ml/min. The GC-MS conditions were optimized with an ion-source temperature of 250 °C, interface temperature of 300 °C, a pressure of 16.2 psi, and a 1.8 mm out time. The sample was injected in split mode with a split ratio of 1:50 using a 1 µl injector, with an injection temperature set at 300 °C. The column temperature was programmed to start at 36 °C for 5 min, then ramped up to 150 °C at a rate of 4 °C/min, followed by a final increase to 250 °C at a rate of 20 °C/min, which was held for 5 min. The total elution time was 47.5 min. The relative percentage of each chemical compound was determined by comparing its average peak area to total areas. Compounds were identified with their retention time and the interpretation of mass spectrum with the National Centre for Biotechnology Information (NCBI) database (Gomathi et al. 2015).

Mosquito larva collection

Larvae of An. gambiae mosquito were collected from stagnant water bodies and identified using the morphological keys established by Gillies and De Meillon (1968). The mosquito larvae were separated from other small aquatic insects and organisms and then transferred into a sterile container filled with distilled water. The containers were sufficiently large to prevent overpopulation until third/fourth larvae stage development, the stage required for the Larvicidal bioassay. The larvae were fed with baker’s yeast once and twice daily at the start of the experiment and during the later stages of growth respectively. The water from the containers was changed daily to prevent the formation of scum on the water surface.

Larvicidal bioassay

The study followed the procedure outlined by the WHO (2005, 2019) was followed. The optimal humidity of larvicidal bioassay was set at 70–75% RH followed the standard given by WHO (2019) and CDC (2020). Twenty third/fourth larval instars from the culture were added into the different concentrations of the plant crude extracts (1%, 5%, 10%, 15% and 20%), along with a set of control group of larvae introduced into sterile distilled water while Temephos (0.0125%) was used as the positive control. All concentrations were replicated thrice. Larval mortality was recorded after 6, 12, 18 and 24 h. Larval mortality data was collected by recording larvae that did not respond when their siphon was pricked with a needle recorded as dead. Using the formula below, the larval mortality percentage was calculated.

graphic file with name 12639_2025_1795_Article_Equa.gif

Pupicidal bioassay

The Pupicidal bioassay used is a similar approach to the larvicidal bioassay. The optimal humidity of Pupicidal bioassay was set at 80–90% RH followed the standard given by WHO (2019) and CDC (2020). Twenty (20) two-day-old pupae was individually introduced into the various concentrations of plant extracts (1%, 5%, 10%, 15% and 20%), along with a control set containing sterile distilled water while Temephos (0.0125%) was used as the positive control. All concentrations tested was replicated three times. Pupae that did not respond to any stimulus (needle pricks) was marked as dead. Using the formula below, the number of dead pupae was counted and recorded after 6, 12, 18 and 24 h of exposure. The pupal mortality percentage was calculated as:

graphic file with name 12639_2025_1795_Article_Equb.gif

Data analysis

Larvicidal and Pupicidal bioassay results was analyzed with one-way ANOVA, where separation of means of the three replicate was done with Duncan’s Multiple Range Test. Lethal concentrations (LC) required to cause 50% and 90% (LC50 and LC90) mortalities were analyzed with Probit analysis in the bioassay because percentage mortalities was recorded (Finney 1971).

Results

Phytochemical analysis of C. adansonii bark extract

The bark extract analysis with GC-MS revealed a mixture of chemical compounds presented in Table 1 which were identified and characterized by their elution time, chemical formula, peak intensity, and molar weight. The compound with the highest concentration was bis(6-methylheptyl) benzene-1,2-dicarboxylate with a 19.70% peak area. The second most abundant compound is the Octadecanoic acid, or stearic acid, accounted with a 19.64% peak area. Another significant compound is hexadecanoic acid (palmitic acid) with a 17.22% peak area. Other compounds include 2-ethylbenzotriazole with a 4.03% peak area. Naphtalene with a peak area of 0.83%. Cyclopentadecane (2.57%), 5-methoxyquinoline-6-carbonitrile (2.56%) Octadecane (2.07%) and 2-(4-methylcyclohex-3-en-1-yl) propan-2-ol (2.18%) are also present.

Table 1.

Chemical composition of C. adansonii bark extract

PK Compound Retention time Area % Molecular formular Molar mass (g/mol)
1 1-ethyl-2-methylbenzene 3.387 0.22 C9H12 120.19
2 Naphtalene 3.765 0.22 C10H8 128.17
3 Undecane 4.442 0.38 C11H24 156.31
4 2-ethyl-1,4-dimethylbenzene 4.687 0.28 C10H14 134.22
5 trimethyl-[methyl-bis(trimethylsilyloxy) silyl]oxysilane 5.542 0.55 C10H30O3Si4 310.68
6 1,3,5,2,4,6-trioxatrisilinane 5.698 0.18 H6O3Si3 138.30
7 2-bromo-4-chloroaniline 5.820 0.14 C6H5BrClN 206.47
8 2,2,4,4,6,6,8,8-octamethyl-1,3,5,7,2,4,6,8-tetraoxatetrasilocane 7.453 0.82 C8H24O4Si4 296.61
9 2,6-ditert-butyl-4-methylphenol 9.953 0.11 C15H24O 220.35
10 1,2,3,4,6a,7,8,9,10,10a,11,11a-dodecahydrobenzo[b]quinolizin-6-one 12.230 0.15 C13H21NO 207.31
11 Tetradecanoic acid 12.841 0.33 C14H28O2 228.37
12 1-chlorooctadecane 13.130 0.12 C18H37Cl 288.9
13 N-(2-amino-2-oxoethyl) tetradec-2-ynamide 13.674 0.16 C16H28N2O2 280.41
14 Pentadecanoic acid 13.863 0.75 C15H30O2 242.40
15 (1S,15S)-bicyclo[13.1.0]hexadecan-2-one 14.674 0.72 C16H28O 236.39
16 Hexadecanoic acid 14.996 29.45 C16H32O2 256.42
17 1-(2 cyclohexyliminocyclopentyl) ethanone 15.308 0.09 C13H21NO 207.31
18 (9Z)-cycloheptadec-9-en-1-one 15.608 0.50 C17H30O 250.4
19 Heptadecanoic acid 15.807 0.49 C17H34O2 270.5
20 4a-methyl-7-propan-2-yl-1,3,4,5,6,7,8,8a-octahydronaphthalen-2-one 16.007 0.11 C14H24O 208.34
21 Ethyl 7-amino-[1,2,4] triazolo[1,5-a]pyrimidine-6-carboxylate 16.063 0.10 C8H9N5O2 207.19
22 11-octadecenoic acid, methyl esther 16.618 44.93 C19H36O2 296.5
23 Octadecanoic acid 16.763 1.72 C18H36O2 284.5
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Phytochemical analysis of C. adansonii leaf extract

The leaf extract analysis revealed a mixture of chemical compositions presented in Table 2 that are characterized with peak intensity, elution time, chemical formula, and molar weight. The most abundant compound present is 11-octadecanoic acid methyl esther, with a 44.93% peak area. The Second most abundant compound is hexadecanoic acid (palmitic acid) with a 29.45% peak area percentage. Other compounds present are pentadecanoic acid (0.75%), (9Z)-cycloheptadec-9-en-1-one (0.50%) and heptadecanoic acid (0.49%) were also present. Aromatic hydrocarbons and siloxane-based compounds, such as undecane (0.38%), trimethyl-[methyl-bis(trimethylsilyloxy)silyl] oxysilane (0.55%), and 22,446,688-octamethyl-13,572,468-tetraoxatetrasilocane (0.82%), were also identified.

Table 2.

Chemical composition of C. adansonii leaf extract

PK Compound Retention time Area % Molecular formular Molar mass (g/mol)
1 2,2,4,4,6,6,8,8-octamethyl-1,3,5,7,2,4,6,8-tetraoxatetrasilocane 3.176 1.60 C8H24O4Si4 296.61
2 1,3,5-trimethylbenzene 3.387 1.20 C9H12 120.19
3 3,3-dimethylhexane 3.865 1.49 C8H18 114.23
4 Undecane 4.442 1.21 C11H24 156.31
5 2-ethylbenzotriazole 4.587 4.03 C8H9N3 147.18
6 Heptanoic acid 5.031 0.69 C7H14O2 130.18
7 Naphthalene 5.498 0.83 C10H8 5.498
8 2-(4-methylcyclohex-3-en-1-yl)propan-2-ol 5.676 2.18 C10H18O 154.25
9 Dodecane 5.820 1.11 C12H26 170.33
10 2,4-dichloro-3,5-dimethylphenol 6.675 1.11 C8H8Cl2O 191.05
11 Tetradecane 8.475 1.69 C14H30 198.39
12 Hexadecane 10.908 1.43 C16H34 226.44
13 Octadecane 13.097 2.07 C18H38 254.5
14 5-methoxyquinoline-6-carbonitrile 13.486 2.56 C11H8N2O 184.19
15 1-methoxy-3-methylsulfanylbenzene 14.486 1.25 C8H10OS 154.23
16 Hexadecanoic acid 14.874 17.22 C16H32O2 256.42
17 Cyclopentadecane 16.408 2.57 C15H30 210.4
18 Octadecanoic acid 16.719 19.64 C18H36O2 284.5
19 bis(6-methylheptyl) benzene-1,2-dicarboxylate 19.796 19.70 C24H38O4 390.6
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Phytochemical analysis of C. adansonii root extract

The root extract analysis with GC-MS disclosed a mixture of chemical elements which are presented in Table 3. Among which are (E)-octadec-6-enoic acid which was the most abundant compound with a peak area percentage of 40.58%. Another major compound present is hexadecanoic acid (palmitic acid), with a 20.64% peak area. Siloxane compound, 22,446,688-octamethyl-13,572,468-tetraoxatetrasilocane (1.78%) was identified. Other compounds include Octadecane with the peak area of 2.07%, 3-(furan-2-yl)-4-prop-2-enyl-1 H-124-triazole-5-thione with the peak area of 1.07%, Oct-2-yn-1-ol with the peak area of 0.35%, and N, N-Bis(2,4-Dimethylphenyl) acetamide with the peak area of 0.51%.

Table 3.

Chemical composition of C. adansonii root extract

PK Compound Retention time Area % Molecular formular Molar mass (g/mol)
1 2,2,4,4,6,6,8,8-octamethyl-1,3,5,7,2,4,6,8-tetraoxatetrasilocane 3.231 1.78 C8H24O4Si4 296.61
2 methyl 3-methyl-2,4-dihydro-1,3-benzoxazine-6-carboxylate 3.609 0.13 C11H13NO3 207.23
3 ethyl 3-methyl-4,5,6,7-tetrahydro-1 H-indole-2-carboxylate 4.253 0.13 C12H17NO2 207.27
4 6,10,14-trimethylpentadecan-2-one 4.453 0.53 C18H36O 268.5
5 4-phenylpyrido[2,3-d]pyrimidine 4.720 0.19 C13H9N3 207.23
6 N, N-bis(2,4-dimethylphenyl)acetamide 4.820 0.51 C18H21NO 267.4
7 2,2,4,4,6,6,8,8,10,10-decamethyl-1,3,5,7,9,2,4,6,8,10-pentaoxapentasilecane 5.342 0.41 C10H30O5Si5 370.77
8 methyl 3,3-dimethyldiaziridine-1-carboxylate 5.842 0.23 C5H10N2O2 130.15
9 1-methylpyrrolidin-2-one 6.109 0.13 C5H9NO 99.13
10 1,3,5,7,9-Pentaethylcyclopentasiloxane 7.664 0.39 C10H25O5Si5 365.73
11 4-fluoro-3-(trifluoromethyl)benzamide 9.975 0.20 C8H5F4NO 207.12
12 4-phenylpyrido[2,3-d]pyrimidine 10.541 0.13 C13H9N3 207.23
13 8-[(2 S,3R)-3-octyloxiran-2-yl]octan-1-ol 12.275 0.37 C18H36O2 284.5
14 17-hydroxy-10,13-dimethyl-3-oxo-2,6,7,8,9,11,12,14,15,16-decahydro-1 H-cyclopenta[a]phenanthrene-17-carbonitrile 12.997 0.13 C20H27NO2 313.4
15 1-ethoxy-4-[(Z)-2-nitroprop-1-enyl]benzene 13.741 0.21 C11H13NO3 207.23
16 (E)-octadec-5-ene 13.908 0.39 C18H36 252.5
17 5-N-(4-fluorophenyl)-3-N-methyl-1 H-1,2,4-triazole-3,5-diamine 14.074 0.15 C9H10FN5 207.21
18 oct-2-yn-1-ol 14.685 0.35 C8H14O 126.2
19 Hexadecanoic acid 14.874 20.64 C16H32O2 256.42
20 3-hydroxy-5,5-dimethyl-2-(prop-2-enyliminomethyl)cyclohex-2-en-1-one 15.407 0.16 C12H17NO2 207.27
21 N-(2-amino-2-oxoethyl)tetradec-2-ynamide 15.496 0.16 C16H28N2O2 280.41
22 4-(3-methoxyphenoxy)-1,2,5-oxadiazol-3-amine 15.685 0.63 C9H9N3O3 207.19
23 (8 S,9 S,10R,13R,14 S,17R)-17-[(E,2R,5R)-5-ethyl-6-methylhept-3-en-2-yl]-10,13-dimethyl-2,3,6,7,8,9,11,12,14,15,16,17-dodecahydro-1 H-cyclopenta[a]phenanthrene 15.841 1.87 C29H48 396.7
24 (3R,8 S,9 S,10R,13R,14 S,17R)-17-[(E,2R,5R)-5,6-dimethylhept-3-en-2-yl]-10,13-dimethyl-2,3,8,9,11,12,14,15,16,17-decahydro-1 H-cyclopenta[a]phenanthren-3-ol 15.996 1.43 C28H44O 396.6
25 (E)-octadec-6-enoic acid 16.485 40.58 C18H34O2 282.5
26 Diethyl bis(trimethylsilyl) silicate 18.918 0.17 C10H28O4Si3 296.58
27 1-methyl-4-phenyl-5-sulfanylidene-1,2,4-triazolidin-3-one 20.040 0.14 C9H9N3OS 207.25
28 [(E)-(3,5-dimethylphenyl)methylideneamino]thiourea 20.218 0.16 C10H13N3S 207.3
29 3-(furan-2-yl)-4-prop-2-enyl-1 H-1,2,4-triazole-5-thione 20.596 1.07 C9H9N3OS 207.25
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Larvicidal activities of C. adansonii crude extracts against Anopheles gambiae

Larvae mortality of Anopheles mosquito was observed for all concentrations of the three plant parts used. The mortality percentage is recorded in Table 4. Larvae exposure to leaf extracts 1%, 5%, 10%, 15% and 20% for 6 h yielded a mean percentage of 0, 46.7, 76.7, 96.7, and 100%, for the bark extracts, 3.33, 15.00, 25.00, 50.00, 28.33 and 55 0.00% mean mortality percentage were recorded while 13.33, 20.00, 31.67, 40.00 and 56.67% mortality percentage was recorded after 6 h exposure to the root extracts. Twelve hours of exposure to the leaf, bark and root extracts recorded a 25.00, 63.33, 85.00, 96.67 and 100.00% mortality percentage, a 3.33, 18.33, 38.33, 50.00 and 68.33% mortality percentage and a 31.67, 41.67, 51.67, 100.00 and 100.00% mortality percentage respectively. 100% mortality percentage was recorded in 20% concentration of the leaf extracts at six, twelve, eighteen and twenty-four hours of exposure, for the root extracts, 100% mortality was recorded at 15% and 20% after twelve, eighteen and twenty-four hours of exposure while the highest mortality percentage recorded at 20% after twenty-four hours is 76.67%.

Table 4.

Larvicidal effect of C. adansonii crude extract on Anopheles gambiae mosquito

Plant extract Conc. (v/v %) Period of exposure
6 h 12 h 18 h 24 h
Leaf 1 0.00 ± 0.00a 25.00 ± 0.00c 25.00 ± 0.00bc 25.00 ± 0.00b
Bark 3.33 ± 3.33a 3.33 ± 3.33a 21.67 ± 1.67b 26.67 ± 1.67b
Root 13.33 ± 1.67b 31.67 ± 1.67d 58.33 ± 4.41e 70.00 ± 0.00de
Leaf 5 46.67 ± 4.41f 63.33 ± 1.67g 66.67 ± 3.33f 66.67 ± 3.33d
Bark 15.00 ± 2.89b 18.33 ± 3.33b 31.67 ± 4.41c 40.00 ± 5.00c
Root 20.00 ± 0.00bc 41.67 ± 1.67e 66.67 ± 1.67f 76.67 ± 3.33ef
Leaf 10 76.67 ± 3.33i 85.00 ± 2.89h 85.00 ± 2.89h 85.00 ± 2.89g
Bark 25.00 ± 0.00cd 38.33 ± 1.67e 46.67 ± 1.67d 63.33 ± 3.33d
Root 31.67 ± 1.67de 51.67 ± 1.67f 76.67 ± 3.33g 81.67 ± 1.67g
Leaf 15 96.67 ± 3.33j 96.67 ± 3.33i 98.33 ± 1.67i 98.33 ± 1.67h
Bark 28.33 ± 1.67cd 50.00 ± 2.89f 63.33 ± 1.67e 70.00 ± 0.00de
Root 40.00 ± 0.00ef 100.00 ± 0.00i 100.00 ± 0.00i 100.00 ± 0.00h
Leaf 20 100.00 ± 0.00j 100.00 ± 0.00i 100.00 ± 0.00i 100.00 ± 0.00h
Bark 55.00 ± 7.64gh 68.33 ± 4.41g 75.00 ± 2.89g 76.67 ± 1.67ef
Root 56.67 ± 6.01h 100.00 ± 0.00i 100.00 ± 0.00i 100.00 ± 0.00h
Positive control 0.0125 100.00 ± 0.0j 100.00 ± 0.00i 100.00 ± 0.00 i 100.00 ± 0.00 i
Negative control 0 0.00 ± 0.00a 0.00 ± 0.00a 0.00 ± 0.00a 0.00 ± 0.00a
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Mortality mean ± Standard error followed by the same alphabet along the column are not significantly different (p > 0.05) from each other using New Duncan Multiple Range Test (NDMRT). Positive control: Temephos

No mortality was recorded at 6 h from exposure to leaf extracts at 1% concentration while 25% mortality percentage was recorded at twelve, eighteen and twenty-four hours of exposure for the other concentrations. Lowest mortality percentage of bark and root extracts at 1% concentration was 3.33% and 13.33% respectively.

The highest concentration (20%) yielded a percentage mortality of 100% after 6 h for the leaf extracts. However, the same concentration yielded 100% mortality after twelve hours for the root extracts while for the bark extracts, the highest concentration did not yield a 100% mortality, instead yielded the highest mortality percentage of 76.67% mortality at twenty-four hours indicating a 23.33% difference in comparison with the mortality percentage of the leaf and root extracts. The overall results indicate that the larvicidal activities of the plant crude extracts had a positive correlation with the concentrations and exposure time which means that increased mortality was observed as concentration and exposure time increased. This results also indicates that C. adansonii leaf extracts are the most active as it was the only extract that recorded 100% mortality faster than the rest after 6 h of larval exposure.

Lethal concentrations (LC) of C. adansonii crude extracts against An. gambiae larvae

The concentrations of C. adansonii required to cause mortality of 50% and 90% in Anopheles mosquito larvae are shown in Table 5. The results demonstrated a clear time-dependent effect, with an in increase in exposure time leading to increased larvicidal activity across all plant parts. The leaf extract demonstrated highest larvicidal activity across all exposure times. After 6 h of exposure, the LC50 was 5.58%, and the LC90 was 12.08%, with a high correlation (R2 = 0.94). At longer exposure period, the LC50 values decreased significantly, indicating 2.54% after 12 h and 2.48% after 18 h which remained consistent with the LC50 value at 24 h. Similar pattern was noticed with the LC90 values which further indicates the leaf extract’s strong larvicidal activity. Highest LC90 value was recorded at 18 and24 hours at 10.56%. The high value of the correlation coefficients (R2 = 0.94–0.99) further suggest a strong cause-effect relationship between exposure and mortality.

Table 5.

Lethal concentrations of C. adansonii extracts against An. gambiae larvae

C. adansonii crude extract Exposure period (h) Intercept ± S.E. Slope ± S.D. R 2 LC50 (LCL - UCL) LC90 (LCL -UCL) p value
Leaf 6 2.11 ± 0.07 3.88 ± 0.26 0.94 5.58 (4.09–7.62) 12.08 (8.86–16.47) 0.49
12 4.19 ± 0.12 1.99 ± 0.50 0.95 2.54 (1.51–4.29) 11.50 (6.82–19.41) 0.64
18 4.31 ± 0.13 1.69 ± 0.59 0.99 2.48 (1.39–4.41) 10.56 (5.95–18.79) 0.72
24 4.31 ± 0.13 1.69 ± 0.59 0.99 2.48 (1.39–4.41) 10.56 (5.95–18.79) 0.72
Bark 6 3.10 ± 0.15 1.32 ± 0.76 0.92 27.17 (13.55–54.49) 251.78 (125.56–504.87) 0.31
12 3.07 ± 0.12 1.69 ± 0.59 0.97 13.70 (8.09–23.20) 78.77 (46.52–133.37) 0.72
18 4.05 ± 0.18 1.06 ± 0.95 0.86 7.89 (3.58–17.40) 65.64 (29.78–144.67) 0.35
24 4.27 ± 0.18 1.05 ± 0.95 0.93 4.98 (2.24–11.07) 59.78 (26.89–132.88) 0.69
Root 6 3.76 ± 0.21 0.89 ± 1.13 0.85 25.19 (9.60–66.11) 229.19 (87.36–601.324) 0.33
12 4.51 ± 0.36 0.49 ± 2.02 0.96 10.02 (1.96–51.39) 200.35 (39.08–1027.07) 0.00
18 5.18 ± 0.39 0.48 ± 2.07 0.91 0.42 (0.08–2.47) 23.46 (3.34–117.64) 0.10
24 5.51 ± 0.55 0.36 ± 2.75 0.97 0.04 (0.003–0.47) 8.07 (0.66–98.28) 0.28
Open in a new tab

Note: R2 = Statistical measure of mortality proportion in regression model

S. E. = Standard error

S. D. = Standard deviation

LD50 = Lethal dosage at which 50% population response

LD90 = Lethal dosage at which 90% population response

LCL = Lower confidence limit

UCL = Upper confidence limit

p value = Chi -square (χ 2) significant

The bark extract also showed significant time-dependent activity in the larvicidal activity. After 6 h, the LC50 was 27.17%, with an LC90 of 251.78%. As the exposure time increased, the LC50 values increased to 7.89% after 12 h and 4.98%after 24 h, with corresponding LC90 values of 65.64% and 59.78%. The correlation coefficients for the bark extract ranged from 0.86 to 0.97. The root extract initially demonstrated a lower larvicidal activity in comparison to the activity of the leaf and bark extracts, with an LC50 of 25.19% after 6 h. However, the root extract’s mortality activity improved significantly with time. After 18 h of exposure, the LC50 value was recorded to be 0.42%, and after 24 h, the extract showed the highest mortality activity with an LC50 of 0.04%. The corresponding LC90 values of 23.46% and 8.07% after 18 and 24 h indicated a similar trend, demonstrating that the root extract’s activity increased with prolonged exposure. The high correlation coefficients (R2 = 0.85–0.97) indicated a strong relationship between exposure time and larval mortality. The larvicidal activity of C. adansonii crude extracts increased with exposure time, with the leaf extract showing the most consistent effectiveness and the root extract demonstrating a high potency after extended exposure.

Pupicidal activities of C. adansonii crude extracts against Anopheles gambiae

Pupae mortality of An. gambiae were recorded for all concentrations of the extracts for the three plant parts used. The mortality percentage is recorded in Table 6. Pupal exposure to leaf extracts at 1%, 5%, 10%, 15% and 20% concentration for 6 h yielded a mean mortality percentage of 25.00, 31.67, 40.00, 98.33, and 100%. For the bark extracts, 1.67, 10.00, 41.67, 75.00 and 100.00% mean mortality percentage were recorded while 3.33, 11.67, 20.00, 26.67 and 36.67% mortality percentage were recorded after 6 h exposure to the root extracts. Twelve hours of exposure to the leaf, bark and root extracts recorded a 26.67, 38.33, 50.00, 98.33 and 100% mortality percentage, a 10.00, 23.33, 53.33, 91.67 and 100% mortality percentage and a 11.67, 20.00, 21.67, 36.67 and 45.00% mortality percentage respectively. 100% mortality percentage was recorded in 20% concentration of the leaf and bark extracts at six, twelve, eighteen and twenty-four hours of exposure while no concentration of the root extracts yielded a 100% mortality.

Table 6.

Pupicidal effect of C. adansonii crude extract on Anopheles gambiae mosquito

Plant extract Conc. (%) Period of exposure
6 h 12 h 18 h 24 h
Leaf 1 25.00 ± 2.99d 26.67 ± 1.67c 28.33 ± 1.67c 28.33 ± 1.67b
Bark 1.67 ± 1.67a 10.00 ± 0.00b 33.33 ± 1.67d 40.00 ± 0.00c
Root 3.33 ± 3.33a 11.67 ± 1.67b 18.33 ± 1.67b 25.00 ± 0.00b
Leaf 5 31.67 ± 1.67e 38.33 ± 1.67de 38.33 ± 1.67e 43.33 ± 1.67c
Bark 10.00 ± 0.00b 23.33 ± 3.33c 51.67 ± 1.67g 68.33 ± 4.41f
Root 11.67 ± 1.67b 20.00 ± 2.89c 28.33 ± 1.67c 41.67 ± 1.67c
Leaf 10 40.00 ± 0.00f 50.00 ± 0.00fg 71.67 ± 1.67h 90.00 ± 0.00h
Bark 41.67 ± 1.67f 53.33 ± 7.26g 71.67 ± 1.67h 80.00 ± 0.00g
Root 20.00 ± 2.89c 21.67 ± 1.67c 30.00 ± 2.89cd 41.67 ± 1.67c
Leaf 15 98.33 ± 1.67h 98.33 ± 1.67hi 98.33 ± 1.67ij 98.33 ± 1.67ij
Bark 75.00 ± 0.00g 91.67 ± 1.67h 95.00 ± 0.00i 95.00 ± 0.00i
Root 26.67 ± 1.67d 36.67 ± 1.67d 43.33 ± 1.67f 51.67 ± 1.67d
Leaf 20 100.00 ± 0.00h 100.00 ± 0.00i 100.00 ± 0.00j 100.00 ± 0.00j
Bark 100.00 ± 0.00h 100.00 ± 0.00i 100.00 ± 0.00j 100.00 ± 0.00j
Root 36.67 ± 1.67f 45.00 ± 2.89ef 51.67 ± 1.67g 61.67 ± 1.67e
Positive Control 0.0125 100.00 ± 0.00h 100.00 ± 0.00i 100.00 ± 0.00 i 100.00 ± 0.00 i
Negative Control 0 0.00 ± 0.00a 0.00 ± 0.00a 0.00 ± 0.00a 0.00 ± 0.00a
Open in a new tab

Mortality mean ± Standard error followed by the same alphabet along the column are not significantly different (p > 0.05) from each other using New Duncan Multiple Range Test (NDMRT). Positive control Temephos

The highest mortality percentage recorded for roots extracts at 20% concentration after twenty-four hours is 61.67%. Lowest mortality percentage was recorded from exposure to bark extracts for 6 h at 1% concentration while 10%, 33.33% and 40.00% mortality percentage was recorded at twelve, eighteen and twenty-four hours respectively. The highest used concentration (20%) yielded a percentage mortality of 100% after 6 h for the leaf and bark extracts. However, the same concentration yielded a 61.67% mortality after twenty-four hours of exposure for the root extracts indicating a 38.33% difference in comparison with the mortality percentage of the leaf and bark extracts. The overall results indicate that the extracts had increasing mortalities proportionate to increasing concentrations and time of exposure. This results also indicates that C. adansonii leaf and bark extracts are the most active as they are the only extracts that recorded 100% mortality faster than the root extracts after 6 h of pupal exposure.

Lethal concentrations (LC) of C. adansonii crude extracts against An. gambiae pupae

The concentrations of C. adansonii required to cause mortality of 50% and 90% in Anopheles mosquito larvae are shown in Table 7. A clear temporal effect was observed as longer exposure times increased the Pupicidal activity of the crude extracts. The leaf extract showed high variability in LC50 and LC90​ values across exposure times. The LC50 value dropped drastically from 55.55 at 6 h to 2.95 at 24 h, indicating increased in effectiveness over time. The decrease in the confidence intervals from 6.97 to 442.95 at 6 h to 1.66–5.27 at 24 h further suggests reliability in the result of increased effectiveness with extended exposure.

Table 7.

Lethal concentrations (LC) of C. adansonii crude extracts against An. gambiae pupae

C. adansonii Crude Extract Exposure period (h) Intercept ± S.E. Slope ± S.D. R 2 LC50 (LCL - UCL) LC90 (LCL -UCL) p value
Leaf 6 4.31 ± 0.46 0.39 ± 2.52 0.93 55.55 (6.97–442.95) 985.36 (123.58–7856.48) 0.00
12 4.36 ± 0.30 0.59 ± 1.68 0.96 12.05 (3.07–47.26) 939.64 (239.49–3686.57) 0.00
18 4.33 ± 0.18 1.01 ± 0.99 0.76 4.64 (2.03–10.58) 88.04 (38.57–200.98) 0.00
24 4.26 ± 0.13 1.63 ± 0.61 0.74 2.95 (1.66–5.27) 19.75 (11.06–35.24) 0.05
Bark 6 2.65 ± 0.09 2.24 ± 0.45 0.89 11.05 (7.32–16.67) 43.21 (28.62–65.22) 0.00
12 3.44 ± 0.10 1.98 ± 0.51 0.78 6.32 (3.97–10.05) 31.77 (19.98–50.52) 0.01
18 4.37 ± 0.14 1.52 ± 0.66 0.78 2.69 (1.45–4.97) 21.45 (11.59–39.71) 0.24
24 4.64 ± 0.15 1.44 ± 0.69 0.89 1.78 (0.91–3.50) 14.75 (7.51–28.96) 0.77
Root 6 3.12 ± 0.19 1.10 ± 0.91 0.98 51.52 (21.56–123.14) 976.44 (408.56–2333.66) 0.88
12 3.79 ± 0.27 0.72 ± 1.39 0.83 46.99 (14.09–156.64) 971.39 (291.42–3237.97) 0.43
18 4.03 ± 0.28 0.66 ± 1.52 0.86 30.09 (8.41–107.63) 846.18 (245.41–2917.65) 0.63
24 4.29 ± 0.27 0.66 ± 1.52 0.90 11.62 (3.37–40.05) 574.04 (160.50–2053.05) 0.86
Open in a new tab

R2 = Statistical measure of mortality proportion in regression model

S. E. = Standard error

S. D. = Standard deviation.

LD50 = Lethal dosage at which 50% population response

LD90 = Lethal dosage at which 90% population response

LCL = Lower confidence limit

UCL = Upper confidence limit

p value = Chi -square (χ 2) Significant

The leaf extract showed R2 values close to 1 which indicate a strong correlation between mortality and concentration. Similarly, as observed in the Pupicidal activity of the leaf extract, the bark extract was also highly effective, as a LC50 value of 11.05 was needed to achieve mortality in 50% of the mosquitoes in 6 h, with a further decrease to 1.78 at 24 h. Similar pattern was also observed in the concentration required to kill 90% of the mosquito population as it decreased from 43.21 at 6 h to 14.75 at 24 h. The bark extract maintained high R2 values throughout the exposure periods, with values of 0.89 at 6 h and 0.89 again at 24 h. This indicates a consistently strong correlation between concentration and mortality. The root extract demonstrated a slight difference to the observations from the leaf and bark extract. For the root extract, the concentration required to kill half of the pupae population in 6 h was 51.52 at 6 h and 11.62 at 24 h. The LC90 values were also consistently higher than those of the other plant parts, indicating that the leaf and bark extracts were more effective in controlling An. gambiae pupae than the root extracts.

Discussion

The analysis of the phytochemical composition of C. adansonii bark extract revealed a cocktail of chemical composition that are responsible for its mosquitocidal activity. bis(6-methylheptyl) benzene-1,2-dicarboxylate is present in the highest percentage in the bark part of the plant and it has been found to be useful used as plasticizers, to soften polyvinyl chloride primarily. They are also mainly used in the production of a products, such as gum, building materials, cosmetics, detergents, surfactants, packaging materials, toys, paints, pharmaceuticals, food products, and textiles. They are also contained in personal care items such as body and hair sprays, skin moisturizer, nail paints and liquid soap. Ghafari et al. (2022) reported significant repellence and larvicidal activity of bis(6-methylheptyl) benzene-1,2-dicarboxylate against Anopheles mosquito. Similarly, Ramamurthy and Krishnaveni (2014) observed that the ethanolic leaf extract of Mukia maderaspatana containing bis(6-methylheptyl) benzene-1,2-dicarboxylate had moderate larvicidal and Pupicidal activity against Ae. aegypti. One of the identified compounds Cyclopentadecane, has been reported to be used in the fragrance industry for its musky odor. It is a monounsaturated fatty acid commonly found in various plant and animal fats and is known for its anti-inflammatory and antioxidant properties (Arctander 1969; Aitzetmüller et al. 2006; Lee et al. 2011).

Octadecanoic acid also known as stearic acid is also a significant component of C. adansonii bark. Stearic acid is a saturated fatty acid with a moisturizing and emulsifying properties which is used in cosmetics, soaps, insecticides, and detergents (Barel and Maibach 2009; Spitz 2013). Dey et al. (2023) observed mortality activity of Annona squamosal extract which has a significant amount of stearic acid against Anopheles mosquito larvae. De-melo et al. (2018) reported significant activity of this compound against Culex larvae and it was found to affect the metabolism and the morphology of their midgut along with their fat body. Stearic acid was also reported to possess mosquito biting deterrent effect (Cantrell et al. 2011). Hexadecanoic acid, another significant chemical compound of the bark extract is found as glycerol ester in oils and fats. Dey et al. (2023) reported mortality activity of Annona squamosal extract with a solid amount of hexadecanoic acid against Anopheles mosquito larvae. Hassan and Jebanesen (2022) observed a significant mosquitocidal activity of hexadecenoic acid against Anopheles mosquito.

Chemical composition of the leaf extracts includes 11-octadecanoic acid methyl ester, which is the most abundant compound has been reported for its Larvicidal activity against Culex mosquito (Baz et al. 2024). Palmitic acid also known as hexadecanoic acid is present in large amount in the bark extract. This compound is also present in the root and leaf extract, additionally confirming the mosquitocidal properties of this plant. Like stearic acid, palmitic acid is a saturated fatty acid and is widely used cosmetics and skincare products because of its moisturizing properties. It helps to create a smooth, protective barrier on the skin through its ability as a thickening agent and emollient (Lodén and Torma 2001; Barel and Maibach 2009). Pentadecanoic acid is also a major compound of the leaf extract, and it has been used for surface active agents, finishing agents, agricultural chemicals (non-pesticidal) and adhesives and sealant chemical (Lide 2008; Elvers et al. 2013). Other compounds present are pentadecanoic acid and (9Z)-cycloheptadec-9-en-1-one and heptadecanoic acid were also present. Aromatic hydrocarbons and siloxane-based compounds present include undecane, trimethyl-[methyl-bis (trimethylsilyloxy) silyl] oxysilane.

(E)-octadec-6-enoic acid was the most abundant compound in the root extract. It is a monounsaturated fatty acid commonly found in plant oil and in several animal and vegetable fats and oils. It is used in cosmetics industry because of its moisturizing properties. Khaldi et al. (2022) reported the presence of (E)-octadec-6-enoic acid in Melia azedarach L which showed significant activity against fourth instar mosquito larvae and the LC50 were calculated to be 35.49 and LC90 to be 79.61 mg/L. It was also observed from the results, that Melia azedarach oil had significant ovicidal activity with cause a noticeable reduction on the development of mosquito eggs into larvae and effective repellency against adult mosquitoes. Hexadecanoic acid which is the second most abundant compound has also been reported for its mortality and repellence activity against Anopheles mosquito. This compound is also present in the bark extract of C. adansonii which is a strong indication of the mosquitocidal activities of the C. adansonii plant. Octadecane, which is different from the octadecanoic acid is also a chemical composition of the root extract and has been reported for its larvicidal and repellent properties in Ageratum houstonianum (Hadidy et al. 2022).

The results from the Pupicidal activity of these plant extracts showed that the plant extracts were effective in causing mortality of An. gambiae pupae. Similar to what was observed in the Larvicidal activity of the leaf extracts, the leaf extracts also demonstrated maximum mortality against the pupae at 6 h exposure time. The bark extract was also highly effective as it caused maximum mortality. These findings agree with the observation of Koomson et al. (2022) who observed a significant Pupicidal activity of Alchornea cordifolia against An. gambiae. Similarly, a dosage dependent response was also observed in the Pupicidal activity of this plant extracts. Furthermore, it was notice that the plant extract was more effective on the larvae compared to the pupa. The physiological differences in cuticle, metabolic rate and hormonal makes the larvae to be more susceptible and sensitivity to insecticides than the pupae. Other factors that make the more susceptible and sensitivity to insecticides include behavioral and biochemical differences. This piece of information makes the larvae of insect vector a major target in the planning disease vector control.

The lethal concentration values (LC50 and LC90) reported in this study was influenced by the period of exposure. It was observed that the LC50 and LC90 values decrease significantly as the exposure period lengthened. This was in conformity with previous insecticidal research that used crude extracts of plants to control various mosquito species of mosquito (Asiry et al. 2017; Abok et al. 2018). Furthermore, C. adansonii leaf extract recorded the lowest LC50 and LC90 for both the larvicidal and Pupicidal activities while the bark extract recorded the highest lethal concentration values for the larvae mortality with the root extracts recording the highest lethal concentration values for the pupa mortality. The result from this study is in conformity with past studies that studied the activity of plants as an environmentally-friendly mosquito-control agents (Ileke et al. 2017; Ileke and Olabimi 2019; Ileke and Adesina 2019).

Conclusion

This study has examined the larvicidal and pupicidal effects of C. adansonii ethanolic extract against Africa malaria vector An. gambiae. Ethanolic extract of C. adansonii plant exhibited strong insecticidal activities against the larvae and pupae stage of the vector. The bark extract exhibited lower larvicidal activity compared to the leaf extract while the root extract demonstrated moderate larvicidal activity. Incorporation of this plant as natural mosquito control agent will help in mitigating the hazardous effects of synthetic chemicals. Furthermore, research on this plant-based insecticides can lead to the discovery of new bioactive compounds with potential applications in the management of vector of public health importance. With our current findings, development of new insecticides in the management of aquatic stages of malaria vector using understudied plant is a promise to mitigate morbidity of malaria infection.

Acknowledgements

The first author, Dr. KD Ileke is grateful for the award of Small Research Grant by Financial Forensic Studies and Diagnostic Limited (FFSD) in May 2024. The authors also acknowledged the technical assistance of the Laboratory staff of the Department of Biology, FUTA.

Author contributions

Kayode David Ileke and Naheemot Atinuke Babatunde designed the study. The laboratory investigation was carried out by Kayode David Ileke, Naheemot Atinuke Babatunde and Adebayo Victor Akeju. Adebayo Victor Akeju and Naheemot Atinuke Babatunde carried out the data analysis. Naheemot Atinuke Babatunde wrote the manuscript. Kayode David Ileke and Adebayo Victor Akeju reviewed the manuscript. All authors read and approved the final manuscript.

Funding

Financial Forensic Studies and Diagnostic Limited (FFSD) Small Research Grant funded the research work.

Declarations

Conflict of interest

The authors declare no competing interests.

Ethical approval

This study was conducted with the approval of School of Postgraduate Studies Research Committee. All procedures and techniques used in these studies were in accordance with the National Institute of Health Guidelines for the Care and Use of Laboratory Animals.

Consent to participate

Not Applicable.

Consent to publish

All authors grant permission for the publication.

Footnotes

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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