Acute toxicity assessment of camphor in biopesticides by using Daphnia magna and Danio rerio

Abstract

29:e2014008 quickly, thereby minimizing the risk of environmental pollution or residual toxicity [1]. Plant-derived insecticidal substances were first commercialized about 160 years ago. Currently, more than 150 related products are produced and sold in 39 countries, and the manufacture of such products is expected to increase [2]. The research institute involved in the present joint research, Environment and Future Ltd., has developed and been selling a bio-pesticide (SSAGRI; Nature & Future, Gokseong, Korea) with camphor extracted from camphor trees as the major ingredient. Eco-friendly bio-pesticides supposedly differ in their degree of influence according to the chemical structure of the extract and toxicity expression mechanism and in their toxicity and susceptibility according to the type of pesticide and the type of organism that is influenced [3,4]. In the "Rural Development Administration's Bio-pesticide Registration Standard and Assessment Method, " the regulatory authorities for the registration and management of pesticides demand that additional toxicity data for aquatic organisms be included, along with Daphnia magna (D. magna) toxicity assessment results, for pesticides that are considered the most hazardous, to reduce the uncertainty of predictions of ecosystem effects because of differences in susceptibility based on the type of pesticide and organism. Thus, in the present research, the toxicities to D. magna and Danio rerio (D. rerio) were assessed. D. rerio is relatively easy to rear, affordable to maintain, and produces 200-300 embryos in a single external fertilization, and therefore, are appropriate for toxicity assessment [5]. In addition, the assessment period was set to 96 hours, allowing quick derivation of toxicity results. Moreover, they are commonly used in medical fields as the DNA concordance rate with humans is nearly 90%. In the present research, the acute toxicity of camphor, which is one of the major ingredients of bio-pesticides, was assessed as the median lethal concentration (LC50) by using the median effective concentration (EC50) for killing and immobilization of water fleas and juvenile D. rerio and its embryos. Additionally, simple lethality offers a limit to assessing the effect of toxicity by determining the degree of toxicity. To assess the influence of the major ingredients of pesticides on the morphologies of different organisms, a fish embryo toxicity test [5] was conducted in addition to the existing toxicity assessment method. For measuring the toxicity in the most sensitive species, the no observed effect concentration (NOEC) and predicted no effect concentration (PNEC) [6] were derived from the acute toxicity assessment results for the two study species, which would provide the stable concentration range for the ecosystem. The results will allow the ecosystem toxicity effect of the plant extracts to be assessed ahead of the development of eco-friendly pesticides with plant-derived extracts. Further, this would be helpful for setting a usage standard within the stable range. Materials and Methods Structure and Physiochemical Characteristics of Camphor Camphor (Sigma-Aldrich Co., Louis, MO, USA; GC level), extracted from Cinnamomum camphora, has a molecular weight of 152.23, octanol-water partition coefficient of 2.38, and a low water solubility but high solubility in ethanol. The structure is shown in Figure 1. The concentration of camphor reagent used in the present research is ≥ 95.0%. was distributed from a chemical toxicity research institute and was cultured in accordance with OECD test guideline 202 [7]. The rearing condition was set to pH 7.5 ± 0.2, temperature of 20 ± 1°C and photoperiod of 16:8-hour light/dark (L/D). For food, 7-10 × 10 7 cells/L of Chlorella vulgaris was provided every day. Rearing water was changed 3 times a week, and the number of female parents was set to 10/L. D. rerio The D. rerio used in acute fish toxicity assessment was distributed from Kyungpook National University. Rearing conditions were set to a temperature of 28 ± 1°C and photoperiod of 16:8-hour L/D; rearing water was dissolved with 0.065 g/L of sodium bicarbonate (Daejung Chemicals & Metals Co., Ltd., Siheung, Korea; extra pure level), and the pH was set to pH 7.5 ± 0.2 with 12% sodium phosphate monobasic anhydrous (Daejung Chemicals & Metals) solution, while the conductivity was set to 0.39-Figure 1. Chemical structure of camphor.