Environ Sci Pollut Res DOI 10.1007/s11356-015-4148-9
Biosynthesized silver nanoparticles using floral extract of Chrysanthemum indicum L.—potential for malaria vector control Selvaraj Arokiyaraj & Vannam Dinesh Kumar & Vijay Elakya & Tamilselvan Kamala & Sung Kwon Park & Muthiah Ragam & Muthupandian Saravanan & Mohomad Bououdina & Mariadhas Valan Arasu & Kalimuthu Kovendan & Savariar Vincent
Received: 3 September 2014 / Accepted: 19 January 2015 # Springer-Verlag Berlin Heidelberg 2015
Abstract Mosquitoes transmit serious human diseases, causing millions of deaths every year. The use of synthetic insecticides to control vector mosquitoes has caused physiological resistance and adverse environmental effects in addition to high operational cost. Insecticides synthesized of natural products for vector control have been a priority in this area. In the present study, silver nanoparticles (Ag NPs) were green-synthesized using a floral extract of Chrysanthemum indicum screened for larvicidal and pupicidal activity against the first to fourth instar larvae and pupae of the malaria vector Anopheles stephensi mosquitoes.
The synthesized Ag NPs were characterized by using UV–vis absorption, X-ray diffraction, transmission electron microscopy, and energy-dispersive X-ray spectroscopy techniques. The textures of the yielded Ag NPs were found to be spherical and polydispersed with a mean size in the range of 25–59 nm. Larvae and pupae were exposed to various concentrations of aqueous extract of C. indicum and synthesized Ag NPs for 24 h, and the maximum mortality was observed from the synthesized Ag NPs against the vector A. stephensi (LC50 =5.07, 10.35, 14.19, 22.81, and 35.05 ppm; LC90 = 29.18, 47.15, 65.53, 87.96, and
Responsible editor: Philippe Garrigues V. Dineshkumar contributed equally to this work. S. Arokiyaraj Institute of Green Bio Science and Technology, Seoul National University, Pyeongchang, Republic of Korea V. Dinesh Kumar : S. Vincent P.G. Research and Department of Advanced Zoology and Biotechnology, Centre for Environmental Research and Development, Loyola College, Nungambakkam, Chennai 600 034, Tamil Nadu, India S. Arokiyaraj : V. Elakya Department of Biotechnology, Vel Tech High Tech Dr. RR Dr. SR Engineering College, Avadi, Chennai 600 062, Tamil Nadu, India T. Kamala Deparment of Biotechnology, Rajalakshmi Engineering College, Chennai 602 105, Tamil Nadu, India S. K. Park Department of Animal Nutrition and Physiology, National Institute of Animal Science, RDA, Suwon 441-706, Republic of Korea M. Ragam Department of Physics, Fatima College, Madurai 625 018, Tamil Nadu, India
M. Saravanan Department of Medical Microbiology and Immunology, Institute of Biomedical Sciences, College of Health Science, Mekelle University, PO Box 231, Mekelle, Ethiopia
M. Bououdina Nanotechnology Centre, University of Bahrain, PO Box 32038, Sakhir, Kingdom of Bahrain
M. V. Arasu Department of Botany and Microbiology, Addiriyah Chair for Environmental Studies, College of Science, King Saud University, P.O. Box 2455, Riyadh 11451, Saudi Arabia
K. Kovendan (*) Division of Entomology, Department of Zoology, School of Life Sciences, Bharathiar University, Coimbatore 641 046, Tamil Nadu, India e-mail: [email protected]
Environ Sci Pollut Res
115.05 ppm). These results suggest that the synthesized Ag NPs have the potential to be used as an ideal eco-friendly approach for the control of A. stephensi. Additionally, this study provides the larvicidal and pupicidal properties of green-synthesized Ag NPs with the floral extract of C. indicum against vector mosquito species from the geographical location of India. Keywords Anopheles stephensi . Chrysanthemum indicum . Biosynthesis . AgNPs . TEM . XRD
Introduction Mosquito-borne diseases are endemic over 100 countries, transmitting the illness to more than 700 million individuals annually inflicting mortality of nearly two million individuals and a minimum of one million children die every year, leaving 2100 million individuals in danger around the world (Jang et al. 2002). Mosquitoes as vectors of serious human diseases like malaria, filariasis, Japanese encephalitis, dengue fever, Chikungunya, and yellow fever constitute a major public health problem all over the globe particularly within the most inhabited tropical countries like India, China, and others. Anopheles stephensi is the primary vector of malaria in India and other West Asian countries, caused by the mosquito parasite Plasmodium falciparum. It is among the leading causes of human morbidity and mortality in tropical and semitropical countries (Snow et al. 2005). One of the approaches for the management of mosquitoborne infections is the interruption of disease transmission by killing or preventing mosquitoes from biting (Mathew et al. 2009). Nowadays, synthetic insecticides/larvicides have created several ecological issues due to their persistent residual accumulation within the surroundings, development of resistance in target vectors, and chronic effects (Mulla et al. 2003). Therefore, researchers necessitate the search for plant products to control the mosquito vectors. Silver nanoparticles have been proven to possess immense importance and thus have been studied extensively (Sharma et al. 2009; Mohanpuria et al. 2008). The green synthesis method has received considerable attention in recent times due to low cost, environmentally friendly, and one-step process for the biosynthesis process (Shankar et al. 2004; Huang et al. 2007; Prakash et al. 2013). In earlier reports, biosynthesized silver nanoparticles (Ag NPs) are reported to have potential antimicrobial, antiplasmodial, and larvicidal properties (Prakash et al. 2013; Suman et al. 2013). Chrysanthemum indicum (Asteraceae) is a native plant of Asia and Northeastern Europe, commonly known as “Mums.” It is a perennial plant, branching herb, grows up to 25–100 cm in height with yellow daisies. It has been traditionally used for the treatment of cancer, pneumonia, colitis, stomatitis, sore, and fever. This plant is known to possess antibacterial, antioxidant, and oxidative DNA damage preventive activity (Jung 2009;
Debnath et al. 2013). In the present study, silver nanoparticles are synthesized using a floral extract of C. indicum and tested for its larvicidal and pupicidal properties on A. stephensi.
Materials and methods Mosquito culture The eggs of A. stephensi were collected from the National Centre for Disease Control field station of Mettupalayam, Tamil Nadu, using an “O”-type brush. These eggs were delivered to the Entomology laboratory and transferred to 18×13×4-cm enamel trays containing 500 ml of water for hatching. The mosquito larvae were cultured by feeding with pedigree dog biscuits and yeast at a ratio of 3:1. The feeding was continued until the larvae transformed into the pupal stage. The pupae were collected from the culture trays using a dipper and transferred to plastic containers (12×12 cm) containing 500 ml of water. The containers were kept in 90×90×90-cm mosquito cage for adult emergence. Mosquito larvae were maintained at a temperature of 27±2 °C, 75–85 % humidity, and kept under a photoperiod of 14:10 (light/ dark). A 10 % sugar solution was provided for a period of 3 days before blood feeding. The adult female mosquitoes were allowed to feed on the blood of a rabbit (one rabbit/day, exposed on the dorsal side) for 2 days, to ensure adequate blood feeding for 5 days. Once blood fed, enamel trays with water from the culture trays were placed within the cage as oviposition substrates. Materials Flowers of C. indicum (Fig. 1a) were purchased from the flower market of Chennai, Tamil Nadu, India, and taxonomically identified. The voucher specimen was numbered and deposited in our research laboratory (CERD, Loyola College, Chennai) for further reference. Flowers were washed with distilled water, shade-dried, and powdered in a blender. AgNO3 (99.8 %) was purchased from Sigma-Aldrich (St Louis, MO, USA). Phytochemical analysis The aqueous floral extract obtained from the flower of C. indicum was tested for the presence of the phytochemicals like tannins, saponins, flavonoids, terpenoids, steroids, alkaloids, and glycosides according to the method described by Edeoga et al. (2005). Green synthesis of Ag NPs Aqueous extract was prepared by dissolving 10 g of powder in a 300-ml flask with 100 ml of sterile distilled water and boiled for 5 min and then filtered with Whatman filter paper no. 1 and stored at 4 °C for further experiments. The filtrate (4 ml) was treated with 1-mM AgNO3 solution (300 ml) in an Erlenmeyer
Environ Sci Pollut Res Fig. 1 UV–vis absorption spectra of Ag NPs synthesized using floral extract of C. indicum after survey scan 350 to 800 nm. Flower of C. indicum (a). Color changes before and after the process of reduction of Ag NPs (b)
flask at room temperature, resulting to the conversion of reddish brown color indicating the formation of Ag NPs within 5 min.
analysis. Energy-dispersive X-ray (EDX) spectroscopy was also used for the elemental analysis of the samples. Larval and pupal toxicity test
Characterization methods The synthesized Ag NPs were characterized by UV–vis spectrometer (UV-Vis Cyber lab 4000) in the wavelength of 350– 800 nm. The crystal structure of the synthesized Ag NPs was characterized by a PANalytical X’pert PRO XRD with Cu/Kα radiation (λ=1.5418 Å) in a 2θ range of 20° to 80°. The size and surface morphology of the synthesized Ag NPs was determined by transmission electron microscopy (TEM) (JEM1101, JEOL, Tokyo, Japan) at 200- and 500-nm resolution. The fine powder form of Ag NPs was dispersed in ethanol on a carbon-coated copper TEM grid, and images were obtained by operating at an accelerating voltage of 120 kV for further
One gram of aqueous floral extract was dissolved in 100 ml of acetone (1 % stock solution). From the stock solution, various concentrations of the extract were prepared (31.25, 62.5, 125, 250, and 500 ppm). The larvicidal activity was assessed according to WHO (1981). Twenty-five larvae (instars I–IV) and pupae were placed in 249 ml of dechlorinated water in a 500-ml glass beaker, and 1 ml of desired concentrations of floral extract and Ag NPs were added. Larval food was given to each test concentrations. At each tested concentration, two to five trials were made and each trial consists of five replicates (Kovendan et al. 2012). Control mortality was corrected by using Abbott’s formula (Abbott 1925), and the percentage mortality was calculated.
Observed mortality in treatment −Observed mortality in control 100 100 − Control mortality Number of dead larvae=pupae 100 Percentage mortality ¼ Number of larvae=pupae introduced
Corrected mortality ¼
Results and discussion
All data were subjected to analysis of variance; the means were separated using Duncan’s multiple range tests by Alder and Rossler (1977). The average larval mortality data were subjected to probit analysis, and LC50 and LC90 values were calculated by using the method of Finney (1971). SPSS software package version 16 was used for accurate analysis (SPSS 2007). Results with P