Chemical Composition,Antifeedant, Repellent,and Toxicity Activities of the Rhizomes of Galangal,Alpinia galanga Against Asian Subterranean Termites,Coptotermes gestroi and Coptotermes curvignathus (Isoptera: Rhinotermitidae)

Dual choice bioassays were used to evaluate the antifeedant property of essential oil and methanolic extract of Alpinia galanga (L.) (locally known as lengkuas) against two species of termites, Coptotermes gestroi (Wasmann) and Coptotermes curvignathus (Holmgren) (Isoptera: Rhinotermitidae). A 4-cm-diameter paper disc treated with A. galanga essential oil and another treated with either methanol or hexane as control were placed in a petri dish with 10 termites. Mean consumption of paper discs (miligram) treated with 2,000 ppm of essential oil by C. gestroi was 3.30 ± 0.24 mg and by C. curvignathus was 3.32 ± 0.24 mg. A. galanga essential oil showed significant difference in antifeedant effect, 2,000 ppm of A. galanga essential oil was considered to be the optimum concentration that gave maximum antifeedant effect. The essential oil composition was determined using gas chromatography-mass spectrometry. The major component of the essential oil was 1,8-cineol (61.9%). Antifeedant bioassay using 500 ppm of 1,8-cineol showed significant reduction in paper consumption by both termite species. Thus, the bioactive agent in A. galangal essential oil causing antifeeding activity was identified as 1,8-cineol. Repellent activity shows that 250 ppm of 1,8-cineol caused 50.00 ± 4.47% repellency for C. gestroi, whereas for C. curvignathus 750 ppm of 1,8-cineol was needed to cause similar repellent activity (56.67 ± 3.33%). C. curvignathus is more susceptible compare to C. gestroi in Contact Toxicity study, the lethal dose (LD₅₀) of C. curvignathus was 945 mg/kg, whereas LD₅₀ value for C. gestroi was 1,102 mg/kg. Hence 1,8-cineol may be developed as an alternative control against termite in sustainable agriculture practices.     

Insecticidal effects of Ungernia severtzovii bulb extracts against the grain aphid Schizaphis graminum (Rondani)

Insecticidal effect of Ungernia severtzovii bulbs extract has been investigated against the grain aphid Schizaphis graminum. LC₅₀ value of the ethanolic extract of U. severtzovii against the female aphids was 0.235% (2.35 g/l). There was 100% mortality of the subsequent nymphs of the treated females at 0.25-1.0%. The extract was equally active against the nymphs at the highest concentrations. Hexane and ethanolic extracts were the most active against the aphids of all the extracts. Fractionation of the hexane extract by TLC yielded three fractions. Fraction II was the most active (>90% mortality) against the aphids. Xanthoxylin was identified as the major constituent in fraction II of the hexane extract and may be responsible for the insecticidal effect of U. severtzovii.     

Effects of crude seed extracts of Annona atemoya and Annona squamosa L. against the cabbage looper, Trichoplusia ni in the laboratory and greenhouse

Antifeedant, growth inhibitory and toxic effects of crude seed extracts of Annona squamosa and Annona atemoya from Fazenda Viveiro Bona, Parasisópolis – Minas Gerais, Brazil, were evaluated against the cabbage looper, Trichoplusia ni (Hübner) (Lepidoptera: Noctuidae) using different bioassays. Crude methanolic seed extracts deterred feeding of third instar T. ni larvae in a leaf disc choice bioassay. A. squamosa was ∼10 times more active as a feeding deterrent than A. atemoya (DC₅₀ = 2.3 mg/ml vs. 20.1 mg/ml). A. squamosa was ∼three times more active as a growth inhibitor than A. atemoya (EC₅₀ = 38.0 ppm vs. 117.0 ppm). Methanolic seed extracts of A. squamosa and A. atemoya were toxic to third instar T. ni larvae both through topical and oral application. A. squamosa was more toxic through feeding (LC₅₀ = 167.5 ppm vs. 382.4 ppm) whereas, A. atemoya exerted greater toxicity via topical application (LC₅₀ = 301.3 μg/larva vs. 197.7 μg/larva). Both A. squamosa and A. atemoya extracts reduced leaf area consumption and larval growth in a greenhouse experiment. Our results indicate that both A. squamosa and A. atemoya have potential for development as botanical insecticides, especially for local use in Brazil.     

Comparative assessment of antioxidant and cholinesterase inhibitory properties of the marigold extracts from Calendula arvensis L. and Calendula officinalis L.

Calendula sp. is an important medicinal and industrial plant with various bioactivities. In this study, we examined enzyme inhibitory effects of the n-hexane, dichloromethane, acetone, ethyl acetate, methanol, and water extracts of the leaf and flowers of Calendula arvensis L. and C. officinalis L. against acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). The extracts were screened for their antioxidant activity using 2,2-diphenyl-1-picrylhydrazyl (DPPH), ferric ion-chelating capacity and ferric-reducing antioxidant power (FRAP) assays at 250, 500, and 1000 μg mL⁻¹. Total phenol and flavonoid quantification of the extracts was achieved using Folin-Ciocalteau and AlCl₃ reagents, respectively. The ethyl acetate extract of C. arvensis flowers was the most active in AChE inhibition assay (31.24 ± 1.29%), while the n-hexane extract of C. officinalis leaves exerted the highest ferric ion-chelating capacity (74.27 ± 2.25%). Thin layer chromatographic analysis indicated presence of flavonoid and triterpene derivatives mainly in the extracts.     

Antioxidant, antibacterial, and antiviral effects of Lactuca sativa extracts

Antioxidant, antibacterial and antiviral effects of aqueous and methanol extracts of Lactuca sativa var longifolia leaves were investigated. The antioxidant activity was evaluated using the DPPH assay. The effect of the extracts against 5 Gram-positive and 6 Gram-negative bacteria was tested. The antiviral activity was determined against human cytomegalovirus (HCMV) strain AD-169 (ATCC Ref. VR 538) and coxsackie B virus type 3 (CoxB-3) using a cytopathic effect (CPE) reduction assay. The methanol extract had the highest total phenolic contents (235.31 mg CE/g extract). It exhibited a significantly (p < 0.05) greater hydroxyl radical-scavenging activity (IC₅₀ = 3.5 μg/ml) than the aqueous extract (4.1 μg/ml). It was also the most effective extract with the lowest MIC (2.5 mg/ml) against all Gram negative and Gram positive bacteria. Methanol and aqueous extracts exhibited antiviral activity against HCMV and Cox-B3 viruses with IC₅₀ of 200 μg/ml.     

Mosquitocidal activity of two Bacillus bacterial endotoxins combined with plant oils and conventional insecticides

The full whole culture (FWC), containing parasporal protein toxins of Bacillus thuringiensis israelinsis (Bti) and Bacillus sphaericus 2362 (Bs), either singly or in combination with plant oils and commercial insecticides, was tested against larval and adult stages of Culex pipiens mosquitoes under controlled laboratory conditions. In terms of LC₅₀ values recorded after 24, 48, 72 and 96 h, the bacterial toxins showed high potency towards both larvae and adults of mosquitoes in a dose-dependent manner. Generally, the Bti toxin seemed to be more potent than the Bs toxin. For example, the Bti toxin showed a 24 h LC₅₀ of 8.2 ppm against mosquito larvae compared to 13.6 ppm for the Bs toxin. In the adult bioassay, the obtained 24 h LC₅₀ values were 0.064 and 0.085 mg/cm², respectively for the two bacterial toxins. The bacterial toxins mixed with plant oils or insecticides at equitoxic doses (e.g., LC₂₅ values) mostly showed potentiation effects, either against larvae or adults of the tested insect. Among a total of 14 paired mixtures, only the joint action estimated for the mixture of malathion + Bti or Bs was accounted as additively. Combining Bti or Bs endotoxins at LC₀ with different plant oils and insecticides at LC₅₀ concentration levels each, has resulted in considerable synergism against either larvae or adults. In the case of larval bioassays, the maximum synergistic factor (SF) obtained (ca. 2.0) was entitled to the mixture of Bti + spinosad. In the adult bioassays, the mixtures containing Curcuma longa or Melia azedarach oil extracts with Bti or Bs toxins achieved a SF accounted to 2.0. The results of the present study may be considered as an additional contribution to the area of joint toxicity of biocidal agents combining bacterial toxins, plant oils and traditional insecticides. The reached findings may encourage future research to elucidate its performance under practical field conditions.     

Enzyme inhibitory and antioxidant activity of Melia azedarach L. naturalized in Anatolia and its phenolic acid and fatty acid composition

In the present study, acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and tyrosinase inhibitory and antioxidant activities of the fruit and leaf extracts of Melia azedarach L. (Meliaceae) of Turkish origin were evaluated. Enzyme inhibitory activity of the extracts was tested in vitro using ELISA microplate reader. Antioxidant activity of the extracts was tested using 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, ferrous ion-chelation, and ferric-reducing antioxidant power (FRAP) assays. Phenolic composition of the extracts was elucidated by high performance liquid chromatography (HPLC) and fatty acid compositions of the fatty oils of the fruits and leaves were elucidated by gas chromatography-mass spectrometry (GC-MS). The ethyl acetate extract from the leaves showed the highest inhibition against AChE (33.63 ± 1.40%) and BChE (92.89 ± 3.05%). The methanol extract from the leaves exerted the best antioxidant activity in DPPH radical scavenging and FRAP assays, while the ethyl acetate extracts of the fruits and leaves had the most notable effect in metal-chelation assay.     

Bioactive properties and chemical constituents of methanolic extract and its fractions from Jatropha curcas oil

The present work is designed to evaluate the bioactive properties of the crude methanolic extract of Jatropha curcas oil and its solvent fractions. The crude methanolic extract obtained was fractionated using a hydrophilic lipophilic balanced (HLB) cartridge and then eluted with different solvents in the order of hexane (F1), dichloromethane (F2), chloroform (F3), ethyl acetate (F4) and methanol (F5), respectively. Total phenolic content of the crude methanolic extract and its fractions was in the range of 0.19-4.5 mg/g as gallic acid equivalent. Antioxidant activity of the crude methanolic extract and its fractions were determined by two complementary test methods, namely, phenanthroline method and 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging method. All samples demonstrated weak antioxidant activity (150-851 μmol Fe/100 g of the extract and IC₅₀ of 1.05-13.5 mg/mL). When compared to butylated hydroxytoluene (BHT), a reference synthetic antioxidant, both showed weaker antioxidative potential. The evaluation of antimicrobial activity of the extracts was performed using a disc diffusion method and a micro-well dilution method against six economic plant disease bacteria. The results showed that all extracts possessed strong to moderate antibacterial activity with varying degrees of growth inhibition against the test bacteria. The minimum inhibitory concentrations (MIC) were in the range of 14.92-428.6 μg/mL. In addition, the chemical constituents in each fraction of the extract were subjected to analyze by gas chromatography-mass spectrometry (GC-MS). The eleven constituents were identified. Among them, 2,4-di-tert-butylphenol, methyl 3-(3,5-di-tert-butyl-4-hydroxyphenyl)propionate and linoleic acid may be the main cause of its strong antibacterial activity. Therefore, this oil present in the methanolic extract had great potential as effective antibacterial sources.     

Potential of extracts of the tropical plant Balanites aegyptiaca (L) Del. (Balanitaceae) to control the mealy bug, Maconellicoccus hirsutus (Homoptera: Pseudococcidae)

The effects of extracts of different parts of the perennial tropical plant Balanites aegyptiaca (L) Del., including various solvent extracts of roots, methanol extracts from leaves, fruits, flowers and roots, partially purified saponins obtained from its roots and a standard saponin were studied on the life cycle (adult longevity, number of eggs, crawlers, adults, weight of adults and % wax content) of a laboratory-reared parthenogenic line of the mealy bug, Maconellicoccus hirsutus (Homoptera: Pseudococcidae). Extracts derived from various parts of B. aegyptiaca (leaves, fruits, flowers, and roots in methanol) affected the life cycle of M. hirsutus with a methanol root extract being the most effective at a concentration of 500 μg ml⁻¹. Partially purified saponin of B. aegyptiaca and the commercial bark saponin extract (Sigma) from Quillaja saponaria at a concentration of 500 μg ml⁻¹ were effective in reducing the longevity of M. hirsutus. Significant reductions in oviposition by M. hirsutus were found for all the extracts at a concentration of 500 μg ml⁻¹. Extracts also affected the number of emerging crawlers, number of adults as well as the weight and wax content of emerging adults. These studies suggest that B. aegyptiaca plant extracts and saponins can be useful botanical insecticides for the protection of crops from mealy bugs.     

The effect of different extraction techniques on extraction yield, total phenolic, and anti-radical capacity of extracts from Pinus radiata Bark

The performance of four techniques, conventional maceration, Soxhlet extraction, microwave assisted extraction (MAE), and ultrasound assisted extraction (UAE), for extraction of Pinus radiata bark, in one and several stages, were evaluated. For each technique, the mass extracted (g extract/g bark), total phenols (by Folin-Cicalteau), and tannin (by precipitation) concentration and anti-radical capacity (diphenyl-picrylhydrazyl, DPPH) were quantified. In one stage, the extracted mass increased in the following order: maceration < UAE < MAE < Soxhlet (p < 0.05). The total phenols and tannin levels were also higher with the Soxhlet technique. With additional extraction stages, only the samples produced with MAE and UAE techniques improved their parameters. Additionally, MAE extracts presented a higher anti-radical capacity than does Soxhlet and Pycnogenol^® extracts. Therefore, MAE was a simple and rapid method that was useful for extraction of P. radiata bark. Scanning electron micrographs (SEM) provided evidence of the mechanical effects on cell walls, mainly evidenced by cell destruction produced by Soxhlet, MAE, and UAE on the bark. In contrast, maceration only results in slightly ruptured cell pores, which could explain its low extraction yield.     

Feeding perturbation and toxic activity of five Chrysanthemum species crude extracts against Spodoptera littoralis (Boisduval) (Lepidoptera; Noctuidae)

The effect of the whole methanol extracts of five Chrysanthemum species on feeding and performance of Spodoptera littoralis (Boisduval) larvae has been investigated in vitro. The extracts exhibited an anti-feeding and phagostimulating activities against cotton leafworm larvae when applied either on leaf discs or incorporated into an artificial diet. Under chosen conditions, the antifeedant index calculated over 24 h for sixth instar larvae significantly varied from 78.55 for Chrysanthemum segetum L. to −44.18 for Chrysanthemum fuscatum Desf. at the dose of 1000 ppm. Toxicity of the extracts was manifested by a high mortality, reduced growth rates, and low weight gain by larvae fed on diets containing 1000–10,000 ppm of the extract. Anyone of the larvae treated with Chrysanthemum macrotum (D.R.) Ball. leaves crude extract survived to pupation at the two higher concentrations. The time to pupation increased for Chrysanthemum grandiflorum flowers crude extract from 11.40 ± 0.93 to 28.93 ± 10.92 days as the extract concentration in the diet increased from 0 to 10,000 ppm. The ingestion of crude extract by the third instar larvae reduced significantly the consumption, growth and utilisation of the ingested and digested food, and reduced digestibility.     

Monitoring resistance of Helicoverpa armigera to different insecticides used in cotton in Spain

Helicoverpa armigera is the key pest of cotton in Spain, resulting in many insecticide treatments against it. The resistance status of H. armigera to different insecticides currently used in cotton was evaluated in Spain in two different seasons, 1999 and 2004. Four populations were tested in total, two in each season. Toxicological bioassays were conducted in the laboratory, and performed on third instar larvae by topical application of the insecticides. LD₅₀'s were estimated by probit analysis and resistance factors (RF) were calculated at the LD₅₀ level. Four insecticides were evaluated, but only endosulfan reached a moderate resistance level (RF = 11.4), and the others (methomyl, chlorpyrifos and lambda-cyhalothrin) showed low resistance (RF between 1.9 and 6.0). Such results indicate the generally low resistance of H. armigera to most of the insecticides used against this pest in cotton in Spain. Possible explanations for this situation are discussed.     

Insecticidal and repellent activity of selected essential oils against of the pollen beetle, Meligethes aeneus (Fabricius) adults

The essential oils from 9 aromatic plants were tested on repellency and mortality of Meligethes aeneus adults. All the tested essential oils caused high mortality of M. aeneus adults in the tarsal tests. The lethal doses after 6 h exposure were ranged between 197 and 1508 μg cm⁻². Essential oils obtained from Carum carvi and Thymus vulgaris were most efficient where LD₅₀ was estimated as 197 and 250 μg cm⁻², respectively.Repellency declined in all the essential oils as a function of time. The longest persistence time was determined for essences obtained from C. carvi and T. vulgaris where significantly the highest repellent index of 65.6% and 63.8%, respectively, was determined. Repellent index lower than 15% was determined for the remaining essential oils.     

Efficacy of Plectranthus glandulosus (Lamiaceae) and Callistemon rigidus (Myrtaceae) Leaf Extract Fractions to Callosobruchus maculatus (Coleoptera: Bruchidae)

As part of on-going efforts to use eco-friendly alternatives to chemical pesticides, methanol crude extracts of Plectranthus glandulosus and Callistemon rigidus leaves were sequentially fractionated in hexane, chloroform, ethyl acetate, and methanol to establish the most active fraction(s) against Callosobruchus maculatus in cowpea. Cowpea seeds (25 g) were treated with 0.5, 1, 2, and 4 g/kg of extract to evaluate the contact toxicity and F₁ progeny production of the beetles in the laboratory. Mortality was recorded 1, 3, and 7 d postexposure. P. glandulosus hexane fraction was more toxic than the other fractions recording 100% mortality at 4 g/kg, within 7 d with LC₅₀ of 0.39 g/kg. Hexane fraction of C. rigidus showed superior toxicity, causing 100% mortality at 4 g/kg within only 1 d of exposure with LC₅₀ of 1.02 g/kg. All the fractions greatly reduced progeny emergence, with C. rigidus hexane fraction being the best progeny inhibitor. Fractions of P. glandulosus and C. rigidus leaves had sufficient efficacy to be a component of storage pest management package for C. maculatus.     

Insecticidal Activity and Insect Repellency of Four Species of Sea Lily (Comatulida: Comatulidae) From Taiwan

The methanol and ethyl acetate (EA) extracts of four species of sea lily (Himerometra magnipinna, Comaster multifidus, Comanthina sp., and Comatella maculata) were evaluated for their insecticidal activity against Yellow-fever mosquito larvae (Aedes aegypti) and their repellency against adult Asian Tiger mosquitoes (Aedes albopictus). The 24-hr minimum inhibition concentration (MIC) data revealed that the extracts from H. magnipinna and the C. maculata were the most active, killing mosquito larvae at 12.5 ppm. The toxicity of the extracts from these four sea lilies in descending order was H. magnipinna (12.5 ppm), C. maculata (12.5 ppm), C. multifidus (100 ppm), and Comanthina sp. (200 ppm). Furthermore, no significant difference in toxicity was found using either EA or methanol as the extraction solvent. The MIC at 12.5 ppm is promising as an insecticide lead. The repellency study results show that EA is a better solvent for one species (H. magnipinna), but the methanol is a better solvent overall. The repellency of these sea lily extracts in descending order was Comanthina sp. MeOH (ED₅₀ at 0.32%), followed by H. magnipinna EA (ED₅₀ at 0.38%), C. multifidus MeOH (ED₅₀ at 0.57%), C. maculata MeOH (ED₅₀ at 0.76%), C. multifidus EA (ED₅₀ at 1.25%), and H. magnipinna MeOH (ED₅₀ at 1.67%). A compound with ED₅₀ <0.5% is considered to be a promising repellant. Among the studied sea lilies, both Comanthina sp. and H. magnipinna have potential to be further developed as mosquito control agents due to their favorable toxicity and repellency.     

Variation in chemical composition and biological activities of two species of Opuntia flowers at four stages of flowering

The chemical composition of hexane extracts from flowers belonging to two species of prickly pear, Opuntia ficus-indica (L.) Miller and Opuntia stricta (Haw.) Haworth has been studied by gas chromatography-mass spectrometry in four developmental stages of flower: vegetative, initial flowering, full flowering, and post-flowering stages. Remarkable differences were noted between the flowers’ compositions. The main compounds were carboxylic acid (28-97%), terpenes (0.2-57%), esters (0.2-27%), and alcohols (<1.8%).The study of the biological activities showed that extracts were active in vitro towards four bacteria and two fungal strains. It exhibited remarkable activity against Pseudomonas aeruginosa, Staphylococcus aureus, and Escherichia coli.Antioxidant activity of the flowers extracts was evaluated by the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical method.Our findings demonstrate the interest of Opuntia flowers extract as a source of bioactive substances and its potential preservative use in food.     

Toxicity of quinones against two-spotted spider mite and three species of aphids in laboratory and greenhouse conditions

Toxicities of the eight quinones were evaluated through leaf dip bioassays conducted against Tetranychus urticae, Myzus persicae, Myzocallis walshii, and Illinoia liriodendri. Based on LC₅₀ values, plumbagin (LC₅₀ = 0.001%) was the most active compound against T. urticae and ubiquinone Q₀ (LC₅₀ = 0.005%), plumbagin (LC₅₀ = 0.010%), and dibromothymoquinone (LC₅₀ = 0.012%) were the most active compounds against M. persicae. The most active compounds against M. walshii were juglone (LC₅₀ = 0.011%) and ubiquinone Q₀ (LC₅₀ = 0.019%), whereas dibromothymoquinone (LC₅₀ = 0.030%), plumbagin (LC₅₀ = 0.033%) and ubiquinone Q₀ (LC₅₀ = 0.058%) were the most toxic to I. liriodendri. Ecotrol (positive control) was the least toxic compound (LC₅₀ = 0.39%) against T. urticae and M. persicae (LC₅₀ = 0.447%). Although the majority of the compounds tested were toxic to all four test species in residual bioassays, there was little overlap among the test species in terms of susceptibility to the compounds and interspecific differences were observed. Regarding structure-activity relationships for quinones, the addition of a hydroxyl group resulted in a significant increase in the toxicity of the 1,4-naphthoquinones, and those possessing a methyl group exhibited the highest levels of activity in T. urticae. The bromine atom at the 2- and 5-positions of the benzoquinone ring is crucial to the toxicity of the compounds against I. liriodendri. Toxicity was greatly affected not only by the number of hydroxyl groups, but also by their positions in the ring in the case of M. walshii. Juglone and plumbagin as residual toxins in the laboratory also reduced the population of two-spotted spider mites compared to EcoTrol™ (positive control) and the negative control in the greenhouse experiment. Some quinones tested may have potential as commercial insecticides and miticides, or alternatively, could serve as lead compounds for the development of more potent crop protection agents.     

Maritime Halophyte Species from Southern Portugal as Sources of Bioactive Molecules

Extracts of five halophytes from southern Portugal (Arthrocnemum macrostachyum, Mesembryanthemum edule, Juncus acutus, Plantago coronopus and Halimione portulacoides), were studied for antioxidant, anti-inflammatory and in vitro antitumor properties. The most active extracts towards the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical were the methanol extracts of M. edule (IC₅₀ = 0.1 mg/mL) and J. acutus (IC₅₀ = 0.4 mg/mL), and the ether extracts of J. acutus (IC₅₀ = 0.2 mg/mL) and A. macrostachyum (IC₅₀ = 0.3 mg/mL). The highest radical scavenging activity (RSA) against the 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) radical was obtained in the ether extract of J. acutus (IC₅₀ = 0.4 mg/mL) and H. portulacoides (IC₅₀ = 0.9 mg/mL). The maximum total phenolic content (TPC) was found in the methanol extract of M. edule (147 mg gallic acid equivalents (GAE)/g) and in the ether extract of J. acutus (94 mg GAE/g). Significant decreases in nitric oxide (NO) production were observed after incubation of macrophages with lipopolysaccharide (LPS) and the chloroform extract of H. portulacoides (IC₅₀ = 109 µg/mL) and the hexane extract of P. coronopus (IC₅₀ = 98.0 µg/mL). High in vitro cytotoxic activity and selectivity was obtained with the ether extract of J. acutus. Juncunol was identified as the active compound and for the first time was shown to display selective in vitro cytotoxicity towards various human cancer cells.     

Genetic structure of natural Tunisian Hypericum humifusum L. (Hypericacae) populations as assessed by allozymes and RAPDs

The genetic diversity within and among seven Tunisian natural populations of Hypericum humifusum L., from different geographic regions and bioclimates, was assessed using 11 isozymic polymorphic loci, and 166 RAPD markers amplified by 8 primers. The genetic diversity within populations based on allozymes was higher (P = 72.46%, Ap = 2.01 and H_e = 0.29), than that revealed by RAPDs (29.52 < P < 39.16% and 0.150 < H < 0.200). Both markers yielded high estimates of genetic differentiation and low gene flow (Nm = 0.257 and 0.508 for RAPD and allozymes, respectively) among populations at all space scales. However, the level of differentiation revealed by RAPDs (Φ_ST = 0.494; G_ST = 0.561) was higher than that based on allozymes (F_ST = 0.117). No correlation (Mantel test) among genetic (F_ST and Φ_ST matrices) and geographical distance matrices was observed indicating no isolation by distance. Cluster analyses from allozyme and RAPD loci did not completely agree. The dendrogram based on allozymes yielded higher separation among most populations, while that from RAPDs separated populations into three distinct subclusters. Groupings of populations, in both dendrograms, did not reflect spatial geographic or bioclimatic patterns, indicating specific adaptation of populations to local environments. The correlation between matrices of allozyme and RAPD band frequencies was not significant (Mantel test). The dendrogram obtained from combined data yielded similar population groupings to that probed by RAPDs suggesting higher accuracy of these markers. Given the high differentiation among all populations even at a low geographic distance, ex situ conservation should involve extensive seed collection from all populations from all bioclimatic zones. The populations from the upper semi-arid bioclimate exhibiting relatively high level of genetic diversity should be first protected.     

Antifungal and antiaflatoxigenic efficacy of Caesulia axillaris Roxb. essential oil against fungi deteriorating some herbal raw materials, and its antioxidant activity

The study deals with evaluation of antifungal and antiaflatoxigenic Caesulia axillaris Roxb. essential oil (EO) against herbal raw materials deteriorating fungi and its free radical scavenging activity. During mycoflora analysis these herbal raw materials were found to be severely contaminated by different fungi and aflatoxins. A total of nine different fungal species were isolated from three herbal raw materials. Aspergillus flavus LHPtc was recorded as the highest aflatoxin B1 producing strain. EOs of some plants were tested for their fungitoxicity against the toxigenic strain A. flavus LHPtc, and C. axillaris EO was found as potent fungitoxicant. C. axillaris EO was chemically characterized through GC-MS analysis which depicted the presence of 18 compounds, dl-limonene and Euasarone being the major components. The EO exhibited broad spectrum of fungitoxicity against fungi causing postharvest deterioration of herbal raw materials. At 1.0 μl ml⁻¹ the oil showed complete inhibition of fungal growth and aflatoxin B₁ production was inhibited at 0.8 μl ml⁻¹. Free radical scavenging activity of the oil was also recorded by 2,2-diphenyl-1-picrylhydrazyl assay, and its IC₅₀ value was found 18 μl ml⁻¹. The safety limit of the EO was determined in terms of LD₅₀ on mice, which was 9166.6 μl kg⁻¹, suggesting its non mammalian toxicity. The EO of C. axillaris may be recommended as a plant based preservative in enhancement of shelf life of herbal raw materials by preventing their lipid peroxidation as well as biodeterioration due to fungal and aflatoxin contamination.