الفهرس 
The genus citrusOnly 14 pages are availabe for public view
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Abstract
Citrus are known as one of the major fruit crops in the world. Citrus peel waste though is seasonal, yet, is a big problem to the processing industries and pollution monitoring agencies. However, citrus peels are a potential source of valuable plant secondary metabolites that show insecticidal activity against several insect species. On the other hand, the adverse effects of chemical control have been motivating the demand for alternatives in pest control, such as the use of natural products. So, there is an increased attention in bringing useful products from citrus peel waste to be converted into value-added products to improve the overall economics of processing units and reduce environmental pollution. It is interesting to use such waste products in combating insect pests. In Egypt, studies concerned with the use of native citrus species against local pests are rare. In this thesis, peels of fresh navel orange (Citrus sinensis) and lemon (Citrus limon) fruits were extracted using n-hexane and methanol as solvents differing in their polarity. The aim was to perform physicochemical and chemical analysis on the peel extracts for potential use as botanicals against the cotton leafworm; Spodoptera littoralis (Boisd.).
- Physicochemical characteristics: -
Percentage yield, pH and specific gravity of lemon and orange peel crude extracts were determined using Hex and MeOH as the extraction solvents. The results showed that when MeOH was used for citrus peels extraction, either for lemon or orange, the yield was much higher than Hex. % yield was 7.84 and 11.8 for MeOH extraction of orange and lemon peels, respectively. It was 1.57 and 1.25 for orange and lemon, respectively, extracted by Hex. On the contrary, Hex extracts were more acidic than MeOH ones. PH was 2.85 and 2.32 for orange and lemon peels extracted by Hex, while it was 4.80 and 4.11, respectively, in the case of MeOH extraction. Citrus peels extracted by MeOH characterized by that their specific gravity were > 1, while those extracted by Hex were <1.
- Chemical composition :-
The results emphasized the relatively great ability of MeOH in extracting phytochemicals from lemon and orange peels. It was more efficient than Hex in withdrawing main metabolites (proteins and carbohydrates) or secondary metabolites such as phenols, Flavonoids and alkaloids. Also, it was more able to extract Flavonoids, alkaloids and phenols from orange peel than lemon.
Carbohydrates were the major component extracted by MeOH. They showed 511 and 515 mg/gm crude extract for orange and lemon peels, respectively. Total proteins amount extracted by MeOH were less than 5% as compared to carbohydrates content. They equaled to 19.1 and 25.1 mg/gm, respectively.
Secondary metabolites showed significant differences (P<0.01) between lemon and orange peels extracted by MeOH (Table,3). orange peel total phenols, flavonoids and alkaloids were 14.3 mg/gm, 77.3 mg CE/gm and 14.5 mg AE/gm, respectively, while they were 8.79, 33.6 and 8.3, respectively , for lemon peel. There were no significant difference (P<0.01) in the studied plant secondary metabolites with respect to citrus species, however they were lower in Hex than MeOH extraction.
- Antioxidant activity:-
The results showed over all antioxidant capacity of citrus peel crude extracts. MeOH had the higher ability than Hex to extract antioxidant components especially from orange peel (P<0.01). Total antioxidant capacity was 26.4 and 21.2 μM/gm extract for orange and lemon extracted by MeOH, and was 3.85 and 1.6 μM/gm extract for orange and lemon extracted by Hex, respectively. Toxicity of citrus peel extracts :-
Toxic action of citrus peel extracts was tested against 4th larval instar of the cotton leafworm, S.littoralis using different methods of treatment (contact and oral). In general, treatment by contact method was more efficient than ingestion (feeding), and Hex extracts were more toxic than MeOH extracts in both treatment methods.
Exposure of larvae for 30 min to contact with crude extracts caused satisfactory results. LC50 of MeOH extracts were 4.15 gm% (orange) and 3.32 gm % (lemon), while those of Hex extracts were 2.31 gm % (orange) and only 0.6 gm % (lemon).
Feeding of larvae for 24h on leaves treated by the extracts caused much lower toxicity than contact method. LC50 of MeOH extracts were 58.62 gm% (orange) and 91.11 gm % (lemon), while those of Hex extracts were 7.67 gm % (orange) and 22.24 gm % (lemon).
- Antifeedant activity:
Feeding of 4th larval instar larvae n leaves treated by 1% of lemon and orange peel crude extracts for 24 h caused slight feeding deterrence to larvae, especially for lemon extracts. The data shows that the treated larvae consumed 20.2, 26.2, 21.1, 23.6 and 25.29 mg/larva for MeOH extract of orange and lemon, Hex extracts of orange and lemon, and control, respectively. Orange MeOH and Hex extracts showed significant difference (P<0.01) as compared to control. The calculated antifeedant activity was 20.2, 16.6, 6.7 and 0.00% for orange extracted by MeOH and Hex, and lemon extracted by Hex and MeOH, respectively.
- Repellent action :
All treatments of the cotton leafworm larvae by citrus peel extracts showed moderate repellent action. The larvae in treated filter paper disc half showed 43.3, 60, 36 and 55 % repellency for orange and lemon peel extracted by MeOH , and for orange and lemon peel extracted by Hex, respectively. One percent (gm%) of the tested extracts did not cause significant difference between the different extracts of the same citrus species.
It could be concluded that the extraction solvent type is a principal factor in determining chemical composition and physicochemical properties of citrus extracts. Citrus peel activity is determined by the solvent used for extraction, citrus species and method of treatment. Lemon extracted by Hex, used for repellency or contact action is the most efficient against the cotton leafworm, S. littoralis larvae.
- physical separation of phytoconstituents :
separation by column chromatography:
Amin et al. (2017) indicated that a potent contact larvicide can be obtained from Hex extracts of fresh lemon or orange peel. The extracted phytochemicals are required to be separated and isolated. Physical methods of separation such as column chromatography techniques were used to separate the crude plant extracts. In column chromatography, different fractions were isolated and collected using increasing polarity of solvents. The separated fractions were checked using developed TLC method to separate different constituents from each other for obtaining single phytochemical in pure form. Fractionation of citrus peel Hex crude extract, using silica gel as the stationary phase, eluted 12 fractions for each citrus species; (L1-L12) for lemon peel and (O1-O12) for orange peel extracts.
TLC analysis of fractions:
Fractions separated by the mobile phase were accurately eluted and detected. Similar fractions were added together, so separated fractions of lemon was reduced to be 6 fractions, while that of orange was reduced to be only 4 fractions.
- Toxic bioactive fractions:
Bioassay test using contact method was performed on different fractions to detect the bioactive fractions toxic to 4th larval instar of S. littoralis. The results showed that lemon fractions were more toxic than those of orange.
- GC/MS identification:
Peels from plants of the genus Citrus are obtained as waste by-products of the citrus processing industry. The citrus peels contain many secondary metabolites that are physiologically active against insects and could be used as natural botanical pesticides as ecofriendly products, so identification of such metabolites is of a valuable interest.
The most potent fractions of lemon and orange extracts were selected for further analysis to identify and compare the bioactive secondary metabolites as phytoconstituents present in such fractions. F2 and F3 fractions of both lemon and orange Hex peel extracts were subjected to GC/MS analysis, as they recorded the highest toxicity against the cotton leafworm, S. littoralis.
The identification of individual components was based on comparison of mass spectral fragmentation patterns with those stored in the mass spectral library built up using pure substances, and the mass spectra from the literature. The percentage of composition was computed from gas chromatography peak areas.
The results indicated that secondary metabolites quality and quantity differ according to the citrus spp., and the quantity of d-limonene is much lower in lemon (≈15%) than orange (≈86%) fractions in spite of lemon fractions were more toxic than the orange. However d-limonene was a major compound in both extracts. The active fractions of lemon and orange peel extracts contained d-limonene, α-pinene, linalool, citronllal and citral. Limonene oxide, Geranial and terpineol were only detected in orange peel extract. The results suggest that citrus extract contains many toxic compounds, other than limonene, and /or they could potentiate each other to give the observed toxicity.Peel waste from native citrus species is promising as a natural biopesticide against the cotton leafworm, S. littoralis larvae. TLC developed showed remarkable separation and column chromatography yielded good amount of plant secondary metabolites. Further studies would be useful to identify the specific toxic constituents for each pest species, and to clarify measures of field use such as stability, methods of application and effect on non-target species.