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Chemical composition and insecticidal activity of Mentharotundifolia L. on Rhyzoperthadominica (F.) (Coleoptera: Bostrichidae) from Algeria.

ABSTRACT

Natural products are good alternative to synthetic pesticides due to their reduced harmful impacts on human health and environment pollution. Pesticides based on plant extracts have been showed their efficacy against a range of pests. This study was conducted to determine the insecticidal activity of essential oils from Mentharotundifolia L. (Lamiaceae) against pest stored-product insect Rhyzoperthadominica (F.) (Coleoptera; Bostrichidae).The obtained oils were analyzed by gas chromatography-mass spectrometry (GC-MS) and 49 compounds were identified. The major compounds identified in the oil were Piperitenone oxide (72.87%) followed by Limonene (2.64%), (E-)Caryophyllene (2.58%) and Germacrene D (2.06 %). Results revealed that at the end of 30 minutes of exposure, the highest concentrations of essential oil applied on grains and on filter paper discs (0.4[micro]l) generates 25% and 50.65% mortality respectively. These rates increased up to 53.3% and 100% respectively after 60 minutes of exposure whereas no mortality was recorded in the acetone treated controls. Median lethal dose ([LD.sub.50]) values of the essential oil from M. rotundifolia against adults of R.dominica on filter paper discs and on grains were [0,25.10.sup.-2][micro]l/[cm.sup.2] and [0,85.10.sup.-2][micro]l/g respectively. Median lethal time ([LT.sub.50])values are reported with the essential oil tested on filter paper and on grains at 0.4[micro]l/ml of concentrations 21.23 minutes and 41.66 minutes respectively. The results suggest that the essential oil of M .rotendifolia might be used as a readily accessible source of natural insecticide agent against R.dominica(F.)adults.

KEYWORDS: Rhyzoperthadominica(F.), Mentharotundifolia L., Insecticidal activity, Essential oil, Chemical composition.

INTRODUCTION

Rhyzoperthadominica(F.)belongs to the family of Bostrichidae (Coleoptera) is one of the most destructive and major insect pest of various stored grain products in tropical and subtropical region. This pest-stored can attacked different variety of cereal grain stored, but prefer wheat[14]. The control of this pest stored insects in storage systems mainly depends on fumigants. There is an urgent need to develop safe alternatives that have the potential to replace the toxic fumigants, yet are effective, economical and convenient to use [5].

The alternatives to conventional insecticides were much recommended. For this plant oil extracts from aromatic plants have been widely investigated. Due to their toxicities against many insects, many researches were established during the last decade [31]. Most of the essential oil constituents are monoterpenoids, which are secondary plant chemicals and considered to be of little metabolic importance. The toxicity activity of many plant extracts against different stored-product pests has been study in different part of world [4, 6, 11]. Essential oils from different plant species possess ovicidal, larvicidal, and repellent properties against various insect species and are regarded as environmentally compatible pesticides [12]. The genus of Mentha includes 18 species and 11 hybrids, among which several species are cultivated for essential oil production. Indeed, mint oils are among the most important essential oils produced in the world. In Algeria the genus of Mentha is represented by six species: M. rotundifolia, M. longifolia, M. spicata, M. aquatica, M. pulegium and M. piperita[26]. Mentharotundifolia L.grow spontaneously in Algeria [10] which has the current Algerian name is "Timarssat", belongs to Lamiacea family and is generally found in humid regions of Algeria. It is an aromatic plant, widely used in traditional medicine [16]. In the present study, the chemical compounds of essential oil of M. rotendifolia from Setif region were determined and the insecticidal activity of the essential oil was tested against the adult stages of the stored-products pest R. dominicaand lethal dose and lethal time were calculated. The essential oil with different concentrations was applied respectively on grain and on paper filter with adults in Petri dish.

MATERIALS AND METHODS

Plant material:

The aerial parts of Mentharotundifolia L. were collectedin Amoucha (North of Setifregion, Algeria) in July 2013 during flowering stage. The fresh aerial parts are dried in the shade away from light at room temperature for used in the extraction of essential oil.

Essential oil distillation and analysis:

100 g of M. rotundifolia aerial part were separately hydro-distilled during approximately for 3 hours using a Clevenger-type apparatus. The essential oils were dried over anhydrous sodium sulfate and stored in sterile tubes at 4[degrees]C until analysis and the treatments.

Chromatographic analysis of essential oils:

The essential oils were analysed on a Hewlett-Packard gas chromatograph Model 5890, coupled to a Hewlett-Packard model 5971, equipped with a DB5 MS column (30 m X 0.25 mm; 0.25 [micro]m), programming from 50[degrees]C (5 min) to 300[degrees]C at 5[degrees]C/min. with a 5 minutes hold. Helium was used as the carrier gas (1.0 ml/min); injection in split mode (1:30); injector and detector temperatures, 250 and 280[degrees]C, respectively. The mass spectrometer worked in EI mode at 70 eV; electron multiplier, 2500 V; ion source temperature, 180[degrees]C; MS data were acquired in the scan mode in the m/z range 33450[19] The identification of the components was based on comparison of their mass spectra with those of NIST mass spectral library[19, 21]and those described by Adams, as well as on comparison of their retention indices either with those of authentic compounds or with literature values[2].

Culture of insect pest:

Adults of R. dominicawas reared in a plastic bottle of 1 l containing wheat which were covered by a fine mesh cloth for ventilation at 25 [+ or -]1[degrees]C, 60[+ or -]5% relative humidity.

Bioassay:

Contact toxicity of the essential oil on filter paper:

Four doses were prepared by diluting each time in 1ml acetone the respective volumes of 0.1, 0.2, 0.3 and 0.4 [micro]l of essential oil of M.rotundifolia. Each of the solutions thus prepared has been answered uniformly on a washerfilter paper (Whatman No. 1) 9 cm in diameter (63.62c[m.sup.2]) placed in a glass Petri dish of the same diameter. This washer was left at room temperature for 15 minutes to allow complete evaporation diluting solvent. For the fifth dose or witness, the puck was treated only with acetone. 20 adults non-sexed taken from their environment and breeding aged seven days has been introduced into each Petri dish containing a washer treated; then the plates were immediately closed. Three replicates were performed for each dose and dead insects were counted every 15 minutes[22].

Contact toxicity of the essential oilon seeds:

The four concentration levels: 0.1, 0.2, 0.3 and 0.4 [micro]l/mlof essential oilacetone solution have been prepared. A sore each dose was added to 20g of seeds contained in a Petri dishand then the whole was homogenized so that all the seeds are uniformly coated. Control plates, the seeds were treated only with acetone. Each dish infested with of 20 insect adults non-sexed and aged to seven days after emergence seeds were treated with each dose prepared. Three replicates were performed for each dose and dead insects were counted every 30 minutes of exposure.

Statistical Analysis:

The probit analysis model was used to estimating lethal dose ([LD.sub.50]) and lethal time ([LT.sub.50]) for insect dose response(insects adults killed) of the M. rotundifolia oil extracts. Lethal Time or lethal dose is the period of time or dose (concentration) required for a proportion of a large group of organisms to respond after being exposed to a specific dose of an injurious agent, such as a drug or radiation or pathogen at a given concentration under a defined set of conditions[23].

The model can be used to estimate [LT.sub.50] in serial-time-dose-mortality data where the effect of time on percentage of kill at one concentration or at several concentrations. [LT.sub.50] and [LD.sub.50] values are obtained by linear interpolation.

In order to determine statistically significant differences in toxicity among the insecticidal activities and exposure times, analysis of variance (ANOVA) was carried out using software package Costat. The results showed significant differences at P <0.0001 levels.

RESULTS AND DISCUSSION

Chemical composition of the essential oil of MentharotundifoliaL:

The plant extractions provide essential oils of yellowish color with a very strong and persistent odor of mint. The yield (0.95%), calculated from dry material. Our results are comparable to results reported for the same species in Algeria (Rouina, Miliana, Chlef)by Bradaet al. [10]but lower than those recorded from Beja, (1.26%) and Bizerte(1.04%) in Tunisia [27], and in Morocco (1.54%) [15].In this study, a total of 49 compounds were identified, which accounted for 97.76% of the essential oil (Table 1).The major component of the essential oils is Piperitenone oxide (72.87%) followed by Limonene (2.64%), (E-) Caryophyllene (2.58%) and Germacrene D (2.06 %).

The essential oil of M. rotundifolia from Algeria is characterized by its high rate of Piperitenone oxide (72.87%). Thisresult is similar to that reported by several studies[10, 18, 31]. Results obtained by Ansari1 et al.,[3] cited Pulegone (69.10%) and Menthone (18.5%) as the major components in M. rotundifolia essential oils from Morocco. Two chemotypes have been found in the essential oils of M. rotundifolia L. growing in Tunisia, which are characterized by the main components Caryophyllene (26.67%), Germacrene D (12.31%), Pulegone (32.09%) and Piperitenone oxide (17.28%) respectively[28]. In others reports, the major compound of Egyptian M. rotundifolia essential oils is Linalool (35.32%) and P-minth-1en-8-ol (11.08%) [7].

The variation of chemical composition of essential oils depends on several factors: botanical origin, genetic factors, weather conditions, phytosanitary problems, type of soil, the method used, the used plant parts, the harvest period of the plant, the degree of drying, the drying conditions, temperature and drying time [8].

Contact toxicity of the essential oil on filter paper:

The percentage of cumulative mortalities of the adults according the time and the dose of the essential oil of M. rotundifoliaused on filter paper is given in Figure 1. We observe a variation in the rate of mortality with the essential oil, the dose tested and the time. The highest dose 0.4 [micro]l/ml causes complete mortality (100%) of the lesser grain borer after 60 minutes exposure. The low value of the [LD.sub.50] (0.16[micro]l) calculated after 30 minutes of exposure confirms the high toxicity of this essential oil on filter paper for these insects. The low value of the [LT.sub.50] is 21.23minutes. The results of the analysis of variance demonstrated a significant (P< 0,0001) difference for the duration of exposure to essential oils and the doses of essential oil tested by contacts on filter paper (Table 2).

Contact toxicity of the essential oil on seeds:

Figure 2 illustrates evolution the percentage of cumulative mortalities relative to control of the adults in function of time and the dose of the essential oil of M. rotundifolia used on seeds. We observe a variation in the rate of mortality with the essential oil, the dose tested and the time. It appears as in the previous case an increase in the percentage mortality adults of R. dominica according time and dose of essential oil. No mortality was observed in the control group during the exposure period. At the higher dose (0.4 [micro]l) we noted 100% mortality of insects after 150 minutes of exposure. The low value of the [DL.sub.50] (0.17[micro]1) after 60 minutes of exposure confirmed the high toxicity of this essential oil, on seeds for these type of insects. The low value of the [TL.sub.50] is 41.66 minutes. The results of the analysis of variance demonstrated a significant (P< 0,0001) difference for the duration of exposure to essential oils and the doses of essential oil tested by contacts on seed(Table 3)

Menthahas historical significance as a medicinal and insecticidal plant in the traditional knowledge system. In the last few decades, many studies have been reported on the insecticidal activity of several Menthaspecies. In this study, essential oil of M. rotundifolia proved to be toxic on seeds and on filter papers against adult R. dominica which leads us to think that the principle assets would probably be one or constituents volatile contenting the essential oil. The toxic effects of this essential oil could depend on its chemical composition and level insect sensitivity. Many studies were carried out on the toxicity of M. rotundifolia against pests of stored product. Clemente et al, [13]tested Dichloromethane extracts from M. rotundifolia on larva of Triboliumcastaneum, significant lethal effects producing higher mortality than the control. The essential oils from M. rotundifolia were tested for their insecticidal activity against Callosobruchusmaculates [7].

Essential oil investigated in our study may be attributed to their having major monoterpenoid components, because some major compounds of the test oils, such as Piperitenone oxide, Limonene, Caryophyllene, [alpha]-pinene, [beta]- pinene possessed insecticidal effects against the test insects[15, 29, 30].

Use of essential oil with insecticidal properties have been reported from all over the world as they are convenient, less expensive, highly effective and safer for the humans and environment. Plants such asLaurusnobilis L, Teucrieumpolium, Menthalongifolia, Thymus daenensis, Achilleawilhelmisii, Artemisahaussknechtii, Lantana camara., Citrus autantium, Cinnamomumzeylanicum, lavandulaofficinalis and Ocimum sanctum have shown insecticidal, antifeedant, repellant and growth regulating properties against various stored grain pests like Triboliumcastaneum (Herbst), Triboliumconfusum (DuVal), Rhyzoperthadominica (Fabr.), Callosobruchuschinensis(L.)[9,17, 20,25,30]

The analysis showed that different types of exposure of essential oil of M. rotendifolia were significantly different from each other and different concentration levels were also significantly different from each other. The [LT.sub.50] and [LD.sub.50] estimated correspond to specific extracts and their different concentration levels.

Conclusion:

In the present research, we performed a phytochemical study of Mentharotundifolia, determined the chemical composition of essential oils and assessed its insecticidal activity. Essential oil from Algeria is characterized by diverse chemical profiles. M. rotundifolia dominated mainly by Piperitenone oxide (72.87%), Limonene (2.64%), (E-)Caryophyllene (2.58%) and Germacrene D (2.06 %)

M.rotundifoliahas showed an important toxic effect against R.dominicaadults. This effect could be attributed to the chemical composition and particularly to the abundance of Piperitenone oxide, Limonene and (E-)Caryophyllenewithout ignoring the synergistic role of minor compounds.

The results and the discussions of this study leads to one concluding that the type of exposure and the concentration of botanical extract used in the study were significantly different from each others. Across all the concentrations 0.4[micro]l/ml is the most potent chemical respectively.

This study is important for R. dominicaresponse (mortality)studies as it has implications regarding the method for analyzing correlated time- dose-response data specifically when the speed of kill is of interest.

M. rotundifoliaessentialoil has a high potential as natural insecticide in stored cereals protection. Further studies should be carried out to increase the number of plants used for pest control in order to obtain cheaper pesticides and environmental pollution will gradually decrease.

ACKNOWLEDGMENTS

The authors would like to appreciate all the team ofLEXVA Laboratory Analytique, Clermont-Limagne, France, also our sincere thanks to Mr.LaourHocine and Mr. MiloudHafsi, Professors at the Ferhat Abbas University ofSetif, Algeriafor the identification of the studied plant species and statistical analysis respectively.

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(1) Radhia Arab, (1) Mustapha Bounechada and (2) Messaoud Ramdani

(1) Laboratory of Amelioration and Development of Vegetable and Animal, Faculty of Nature Life Sciences, Ferhat Abbas University Setif-1, 19000 Setif, Algeria

(2) Laboratory of Natural Ressource Valorisation, Faculty of Nature Life Sciences, Ferhat Abbas University Setif-1, 19000 Setif, Algeria

Address For Correspondence:

Radhia Arab, Laboratory of Amelioration and Development of Vegetable and Animal, Faculty of Nature Life Sciences, Ferhat Abbas University Setif-1, 19000 Setif, Algeria

E-mail: arab.radhia@yahoo.fr

This work is licensed under the Creative Commons Attribution International License (CC BY).

http://creativecommons.org/licenses/by/4.0/

Received 28 August 2016; Accepted 18 October 2016; Available online 22 October 2016

Table 1: Chemical composition of essential oil of MentharotundifoliaL.
from Algeria

N[degrees]  compound                     RT       %

1           [alpha]-Thujene               6.0236   0.0745
2           [alpha]-pinene                6.1927   1.486
3           Camphene                      6.5712   0.5325
4           Sabinene                      7.1054   0.7703
5           [beta]-pinene                 7.2123   1.5513
6           1-Octen-3-ol                  7.2969   0.9129
7           Myrcene                       7.4883   1.1911
8           Octanone 2                    8.0047   0.0661
9           Limonene                      8.3743   2.6464
10          1,8-Cineole                   8.4366   0.2031
11          (Z)-Ocimene                   8.5123   0.291
12          [beta]-(E)-Ocimene            8.7349   0.0954
13          [gamma]-Terpinene             8.9886   0.0612
14          Cis-Sabinene hydrate          9.2647   0.6307
15          Terpinolene                   9.5585   0.1078
16          Cis-thujone                   9.9058   0.1228
17          1-Octen-3-yl acetate         10.0126   0.7296
18          Allo-Ocimene                 10.4267   0.1848
19          Lavandulol                   11.1524   0.0953
20          Borneol                      11.3483   1.945
21          Terpinen-4-ol                11.4907   0.2645
22          [alpha]-Terpineol            11.7712   0.2837
23          Shisifuran                   12.1496   0.193
24          Hexyl isovalerate            12.5993   0.0573
25          Trans- Piperitone epoxide    12.8531   0.2931
26          cis-Carvone oxide            13        0.3815
27          Isobornyl acetate            13.3918   0.1504
28          Sesamol                      13.4319   0.3388
29          Piperitenone oxide           14.999   72.8737
30          [beta]-Bourbonene            15.1637   0.238
31          [beta]-Elemene               15.2394   0.3137
32          Z-Jasmone                    15.3017   1.1362
33          [alpha]-Gurjunene            15.5244   0.0977
34          (E-) Caryophyllene           15.7514   2.58
35          [beta]-Copaene               15.8939   0.2748
36          Muurola 3,5 dieneCis         16.1254   0.28
37          [beta]-Sesquiphellandrene    16.1877   0.6619
38          [alpha]-Humulene             16.3079   0.2832
39          Trans-Muurola-4(14),5-diene  16.4014   0.3234
40          Germacrene D                 16.7086   2.0645
41          Bicyclogermacrene            16.9223   0.0702
42          [gamma]-Amorphene            16.9891   0.0781
43          [delta]-Amorphene            17.2429   0.0663
44          cis-Calamenene               17.2918   0.1539
45          Germacrene D-4-ol            18.1555   0.1319
46          Caryophyllene oxide          18.2446   0.1437
47          Viridiflorol                 18.4316   0.1805
48          1,10-di-epi-Cubenol          18.7165   0.0848
49          [alpha]-Cadinol              19.2864   0.0726

Table 2: Results of the analysis of variance for the factor duration of
exposure to essential oil and the factor doses of essential oil tested
byesults contacts on filter paper

Times of treatment  Means   Doses        Means

90 minutes          16.0a   0.4[micro]l  14.26a
60minutes           14.4b   0.3[micro]l  13.13b   0000***
45minutes           12.06c  0.2[micro]l  12.53bc
30 minutes           7.13d  0.1[micro]l  12.13d
15minutes            2.46e  0[micro]l    0e

#Means within columns with the same letter are significantly different
using LSD at level (P< 0,0001)

Table 3: Results of the analysis of variance for the factor duration of
exposure to essential oils and the factor doses of essential oil tested
by contacts on seed

Time of exposition  Means   Doses        Means   P

180 minutes         16a     0.4[micro]l  15.33a  0000***
150 minutes         15.53a  0.3[micro]l  14.27b
120 minutes         13.8b   0.2[micro]l  13.38c
90 minutes          11.6c   0.1[micro]l  13.05c
60 minutes           7.53d  0[micro]l    0d
30 minutes           2.8e

#Means within columns with the same letter are significantly different
using LSD at level (P< 0,0001)
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Author:Arab, Radhia; Bounechada, Mustapha; Ramdani, Messaoud
Publication:Advances in Environmental Biology
Article Type:Report
Geographic Code:6ALGE
Date:Oct 1, 2016
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