Management of Citrus sinensis peels for protection and treatment against gastric ulcer induced by ethanol in rats

Abstract

Introduction: Stomach ulcer is one of the most prevalent disorders worldwide. The study was aimed to isolate and characterize the major polymethoxylated flavonoids in Citrus sinensis peels petroleum ether extract and investigate its protective and curative effect on gastric ulcer.

Material and methods: Some spectral analyses were used for identification of the isolated compounds from the petroleum ether extract of Citrus sinensis peels. One oral dose (0.5?ml/100?g b.wt.) of absolute ethanol was orally given to rats after starvation for 24?h to induce gastric ulcer. To explore the protective and curative role of the plant extract, it was orally (250?mg/kg b.wt.) given for 1 week either before or post-ulcer induction. A reference drug, ranitidine (100?mg/kg b.wt.), was also evaluated. Stomach acidity, gastric volume, lesion counts, glutathione (GSH), superoxide dismutase (SOD), malondialdehyde (MDA), succinate dehydrogenase (SDH), lactate dehydrogenase (LDH), acid phosphatase (AP), interlukin-10 (IL-10) and prostaglandin E2 (PGE2) were estimated. Stomach histopathological features were monitored.

Results: Nine polymethoxy flavonoids were identified from the extract. Treatment with C. sinensis peels extract recorded amelioration in all parameters.

Conclusion: Citrus sinensis petroleum ether peels extract had protective and curative effects against gastric ulcer. Therefore, the extract recorded anti-secretory, anti-ulcerative, anti-inflammatory, and antioxidant effects. Its healing action exceeded its protective role due to its richness in polymethoxylated flavonoids     

6,7-Dimethoxycoumarin in the peels of Citrus

6,7-Dimethoxycoumarin was isolated and identified from the peels of Citrus sinensis cv. Pineapple orange, of C. paradisi x C. paradisi x C. reticulata cv. Wekiwa and of C. mitis cv. Clamondin. The coumarin was tentatively identified in two other sweet oranges, C. sinensis cv. Mediterranean Sweet and Waite Parson Brown, and in C. aurantifolia cv. West Indian lime and in C. aurantium cv. Bitter Sweet orange.     

Induced parthenogenesis in mandarin for haploid production: induction procedures and genetic analysis of plantlets

This study focused on haploid induction in mandarin through in situ gynogenesis by pollination with irradiated pollen of ‘Meyer’ lemon. Pollination was carried out for three genotypes of mandarin with four levels of gamma-ray-irradiated pollen (150, 300, 600, and 900 Gy). The resulting seeds were characterised by a small size. Embryos were rescued in vitro and the ploidy level of the plantlets was determined by flow cytometry analysis. Haploid, diploid, triploid plantlets were obtained. The haploid parthenogenetic origin was confirmed using microsatellite marker analysis and chromosome count. Diploid and triploid plants were the result of crosses between mandarin and lemon. The induction of gynogenetic haploids of ‘Fortune’ (Citrus clementina Hort ex Tan. × Citrus tangerina Hort ex Tan.) and ‘Ellendale’ (Citrus reticulata Blanco × Citrus sinensis L. Osb) is reported here for the first time.     

Energy Allocation Changes in Overwintering Adults of the Common Pistachio Psylla, Agonoscena pistaciae Burckhardt & Lauterer (Hemiptera: Psyllidae)

The common pistachio psylla, Agonoscena pistaciae Burckhardt & Lauterer (Hemiptera: Psyllidae), is known as the key pest of pistachio orchards in Iran. This pest passes the winter as adults. In this study, energy allocation changes in relation to ambient temperature were investigated in field-collected adults by measuring total body sugar, trehalose, glucose, sorbitol, myoinositol, glycogen, lipid, and protein contents. Glycogen content decreased with decrease in ambient temperature. The decrease in glycogen content was proportional to the increase in total body sugar, trehalose, myoinositol, and sorbitol contents. In January, with mean ambient temperature of 5.4°C, glycogen content was at the lowest level, whereas total body sugar, trehalose, glucose, and sorbitol were at the highest level. Total body sugar, trehalose, myoinositol, and sorbitol contents increased as temperature decreased from 22.7°C in October to 5.4°C in January. In conclusion, low molecular weight carbohydrates and polyols may play a role in winter survival and adaptation to cold of the common pistachio psylla by providing the required cryoprotection. Also, overwintering adults of the common pistachio psylla may store energy in the form of lipid for later utilization during the overwintering.     

Evaluation of phenolic and flavonoid compounds in pollen grains of three Citrus species in response to low temperature

Low temperature during the reproductive stage causes sterility of pollen grains and reduces yield. Phenolic compounds function as stress indicators because they accumulate to high levels in many plant tissues in response to a wide range of biotic and abiotic signals. Branches with unopened flowers were collected from trees of three Citrus species (C. reticulata, C. sinensis and C. paradisi) and exposed to different temperatures (5, 10, 15, 20 and 25 °C) for six hours. The highest amount of polyphenol and flavonoids was observed at 5 °C and then their content reduced with increasing temperature in all species. Total phenols and flavonoids content in control temperature as well as all treatments showed that C. paradisi has lower amounts of these compounds than the two other species. HPLC analysis demonstrated that low temperatures, particularly 5 °C, induce flavonoids accumulation as both peak number and flavonoid levels for retention times in pollen grains of all species. The comparison of the HPLC patterns of the three species showed that flavonoid extracts in pollen of C. paradisi decrease in peak number and flavonoids levels for retention times in both control and treatments. Citrus reticulata showed an increase of eight- and two-fold in peaks of one and four, respectively, at 5 °C. Peaks of one and six in C. sinensis increased about four- and three-fold, respectively, at 5 °C compared to the control. In C. paradisi, the levels of peaks two to four were near to the baseline in control samples. With temperature reduction, peak two showed no significant increase but the levels of peaks one, three and four in samples exposed to 5 °C increased approximately three-, three- and four-fold, respectively. ?     

Toxic effects of pyriproxyfen,neemarin, acetamiprid and Ferula assafoetida essential oil on the common pistachio psylla,Agonoscena pistaciae

The common pistachio psylla, Agonoscena pistaciae, Burckhardt and Lauterer (Hemiptera: Psyllidae) is the key pest of pistachio orchards in Iran. Chemical control is the common method for the control of this pest. In this study, the effects of pyriproxyfen (an insect growth regulator), neemarin (a botanical insecticide), acetamiprid (a neonicotinoid) and Ferula assafoetida essential oil were investigated on the fifth instar nymphs of A. pistaciae. Probit analysis of concentration-mortality data was conducted to estimate the LC₅₀ values. The LC₅₀ values for neemarin, pyriproxyfen, acetamiprid and F. assafoetida essential oil were estimated to be 0.22, 0.52, 290 and 5.62 ppm, respectively. Our results showed that the fifth instar nymphs of A. pistaciae are very sensitive to neemarin and pyriproxyfen.     

Restoration of vigor and rooting competence in stem tissues of mature citrus by repeated grafting of their shoot apices onto freshly germinated seedlings in vitro

Summary Repeated grafting of 0.2-cm shoot tips from fruiting-age trees ofCitrus reticulata Blanco ‘Ponkan’ mandarin andC. sinensis Osbeck ‘Liu Tseng’ sweet orange onto freshly germinated ‘Troyer’ citrange [Poncirus trifoliata (L.) Raf. X.C. sinensis Osbeck] seedlings in vitro resulted in progressive restoration of rooting competence and vigor of regenerated roots and shoots. The restored traits were retained through the course of the investigation and suggested a phase reversal phenomenon.     

Symptomatic expression of tristeza-infected citrus plants in Pakistan

Variable symptoms were recorded during a survey in the citrus trees infected or suspected to be infected with citrus tristeza virus based on ELISA tests. Sweet orange manifested main symptoms like bud union crease, pin holing, bark cracking, incompatibility, yellowing and dropping of leaves. Pin holing was particularly prevalent in Sweet orange (Citrus sinensis Osbeck), Kinnow (Citrus reticulata Blanco) and Grapefruit (Citrus paradisii Mcaf.), while incompatibility was only in Sweet orange and Grapefruit.     

Acaricidal properties of essential oils from agro‐industrial waste products from citric fruit against Tetranychus urticae

Tetranychus urticae is a major agricultural pest with worldwide distribution that has caused considerable damage to vegetable crops in north‐eastern Brazil. The aim of the present study was to investigate the chemical and lethal/sublethal effects of essential oils from the peels of the lime (Citrus aurantiifolia), lemon (C. limon), mandarin orange (C. reticulata) and (C. reticulata × C. sinensis) as well as selected constituents (linalool, α‐terpineol, α‐pinene, β‐pinene, terpinolene and limonene) against T. urticae. The greatest yield was achieved with the mandarin and tangerine peel oils. The chemical analysis (gas chromatography‐mass spectrometry) of the essential oils from the Citrus fruit peels enabled the identification of 127 compounds, revealing a predominance of monoterpenes. Limonene was the major constituent, and α‐pinene, β‐pinene, linalool and α‐terpineol were found in substantial quantities. Regarding the susceptibility of T. urticae, the Citrus oils and selected constituents were more effective by fumigation than residual contact. The C. reticulata oil was the most toxic by fumigation, and the C. limon oil was the most toxic by residual contact. The constituent α‐terpineol exhibited the highest toxicity with both methods. At a sublethal concentration, the oils and selected constituents had significant effects on the fecundity, feeding preference and oviposition of the mite. Citrus oils and their constituents are potentially useful for the future integrated management of T. urticae due to their lethal and sublethal properties. However, further studies are needed to evaluate the action of these essential oils against non‐target organisms and determine the cost–benefit ratio for the formulation of an acaricide harvested from agro‐industrial waste from citric fruit processing activities for use in the integrated control of T. urticae.     

Citrus somatic hybridization with potential for improved blight and CTV resistance

Summary The production of five new somatic hybrids with potential for improved disease resistance is reported herein. Protoplast isolation, fusion, and plant regeneration was achieved from Caipira sweet orange (Citrus sinensis L. Osbeck) as an embryogenic parental source and Volkamer lemon (C. volkameriana Pasquale), Cleopatra mandarin (C. reticulata Blanco), and Rough lemon (C. jambhiri Lushington) as non-embryogenic parental sources. Fusion involving Cleopatra mandarin and Rangpur lime (C. limonia L. Osbeck) as embryogenic parental sources with Sour orange (C. aurantium L.) also resulted in somatic hybrid plants. Somatic hybridization was confirmed by leaf morphology evaluation, chromosome counting, and randomly amplified polymorphic DNA (RAPD) analyses. Somatic hybrids may combine complementary characteristics from both parental sources and have potential for tolerance to blight and citrus tristeza virus (CTV).     

Oviposition preference of swallowtail butterfly,Papilio polytes (Lepidoptera: Papilionidae) on four Rutaceae (Sapindales) host plant species

Abstract Oviposition preference and egg deposition site selection by the butterfly, Papilio polytes L. (Lepidoptera: Papilionidae) on four rutaceous host plants, Citrus aurantofolia (Christm.) Swing., Citrus hystrix DC., Citrus reticulata Blanco, and Murraya koenigii (L.) Sprengel, were assessed in field cages measuring 2 × 2 × 2 m. Simultaneous two-choice and four-choice oviposition tests of whole host plants were conducted. The mean total number of eggs laid per plant on C. reticulata in the two-choice test was significantly higher (P < 0.01) than those on C. aurantifolia, C. hystrix, and M. koenigii. Among the three other host plants, C. aurantifolia was preferred over C. hystrix, and M. koenigii (P < 0.01) and C. hystrix was preferred over M. koenigii (P < 0.01). In the four-choice test, C. reticulata was highly preferred and significantly different from C. hystrix and M. koenigii. However, no oviposition preference was detected with C. aurantifolia. Among the various plant parts in the two- and four-choice tests, eggs on leaves of each plant were the highest, followed by numbers on stems, and negligible numbers on pots. These numbers on leaves of C. reticulata and C. aurantifolia were not significantly different (P > 0.05), but differed significantly (P < 0.01) from those on leaves of C. hystrix and M. koenigii. The quantitative trend of egg-laying on stems was very similar to that observed for the leaves. Papilio polytes showed strong preference to lay eggs on the underside of leaves of all host plants than on the upper side or on the petiole. More eggs were laid on the upper side of each host plant than on its petiole. The four host plants in descending order of preference were C. reticulata≥C. aurantifolia > C. hystrix > M. koenigii. Although M. koenigii was the least preferred, it has the potential to serve as an alternative host plant for P. polytes which can be manipulated when necessary, to alleviate the infestation of this pest to the citrus industry.     

<Emphasis Type="Italic">Camellia sinensis</Emphasis> increased apoptosis on U2OS osteosarcoma cells and wound healing potential on NIH3T3 fibroblast cells

Camellia sinensis (Cs) is a plant which is rich in polyphenols and has antioxidant, antiinflammatory, antimutagenic, anticarcinogenic and antibacterial activities. In this study, two different methanol extracts (Cs-I and Cs-II) were prepared from the leaf of C. sinensis in order to investigate the wound healing and anticancer activities. Total phenolic content and antioxidant activity of the extracts were determined. Wound healing effects of Cs extracts were evaluated by using Masson’s Trichrome Tecnique on NIH3T3 fibroblast cells. Cytotoxic and apoptotic effects of the extracts were determined by MTT and AnnexinV-PI assays on U2OS osteosarcoma cells. Total phenolic contents and antioxidant activities of the extracts were almost the same. The highest concentration (60 µg/mL) of the extracts showed significant cytotoxic and apoptotic effects on U2OS cells. Especially, the highest apoptotic effect was determined with 60 µg/mL Cs-I extract. Significant wound healing potential on NIH3T3 fibroblast cells were determined especially with low extract concentrations (0.5, 1 and 5 µg/mL), while high extract concentrations showed significant anticancer effects. As a result, two Cs leaf extracts exhibited important apoptotic properties and both have wound healing potential. However, the Cs-I extract was found more effective on apoptotic osteosarcoma cell death and has an increased wound healing potential than the Cs-II extract.     

Comparative evaluation of petitgrain oils from six Citrus species alone and in combination as potential functional anti-radicals and antioxidant agents*

Six Citrus petitgrain oils, namely C. × sinensis (L.) Osbeck., C. × limon (L.) Osbeck., C. × clementina Hort., C. × aurantium L., C. reticulata Blanco and C. × bergamia Risso & Poit. (P1-P6), were characterized by means of gas chromatography equipped with flame ionisation detector (GC-FID) and gas chromatography coupled to mass spectrometry (GC-MS) and evaluated for their food functional-related properties. Antioxidant and radical-scavenging properties were tested by means of 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay, β-carotene bleaching test and Ferric Reducing Antioxidant Power (FRAP) assays. The synergistic interactions on antioxidant efficacy of petitgrain oils in combination were studied. The Relative Antioxidant Capacity Index (RACI) was applied from the perspective of statistics by integrating the antioxidant capacity data determined by all used methods. A high content of monoterpenes and a less representative content of sesquiterpenes characterized Citrus petitgrain oils. In the DPPH assay, C. × aurantium showed major effectiveness with an IC₅₀ value of 27.2 μg/ml. Citrus petitgrains oils, a rich source of healthy phytochemicals, should be considered a by-product of interest.     

Carotenoids synthesized in citrus callus of different genotypes

Eight carotenoids, such as phytoene, α-carotene, violaxanthin, etc., synthesized in citrus callus of 31 genotypes were identified and determined. Though varied with genotypes, the carotenoids composition of callus derived from a certain genotype was stable, while carotenoids contents altered between sub-cultures. Some specific carotenoids were produced in calluses of limited genotypes: β-citraurin was only synthesized in calluses of Nianju tangerine (Citrus reticulata Blanco) and Page tangelo (C. reticulata × C. paradisi); while 9-Z-violaxanthin was only detected in Nianju tangerine and Skaggs Bonanza navel orange (C. sinensis L. Osbeck). Notably, the only carotenoid detected in calluses of Natsudaidai (C. aurantium L.) and other two sweet oranges (C. sinensis L. Osbeck) was phytoene. It implied that citrus calluses could be employed to produce specific carotenoids in the future. To further elucidate the characters of callus carotenoids profile, comparisons of carotenoids profiles was made among calluses, fruit tissues and leaves of four selected citrus genotypes. Results showed that lycopene was not detected in leaves and calluses; nevertheless, both citrus fruits and calluses accumulated phytoene, whereas leaves did not except those of Cara Cara navel orange. It is postulated that citrus callus featured its carotenoids profile different from fruit tissues and leaves. In conclusion, the advantages of using citrus callus as an alternative model research system in understanding the regulation of carotenogenesis have been discussed.     

Quantitative trait locus (QTL) analysis of fruit-quality traits for mandarin breeding in Japan

The improvement of fruit quality is an important objective in citrus breeding. Using an F₁ segregating population from a cross between citrus cultivars ‘Harehime’ (‘E647’—‘Kiyomi’ [Citrus unshiu Marcow. ‘Miyagawa Wase’ × Citrus sinensis (L.) Osbeck ‘Trovita’] × ‘Osceola’—a cultivar of clementine [Citrus clementina hort. ex Tanaka] × ‘Orland’ [Citrus paradisi Macfad. ‘Duncan’ × Citrus tangerina hort. ex Tanaka] × ‘Miyagawa Wase’) and ‘Yoshida’ ponkan (Citrus reticulata Blanco ‘Yoshida’), a SNP-based genetic linkage map was constructed and quantitative trait locus (QTL) mapping of four fruit-quality traits (fruit weight, sugar content, peel puffing, and water rot) was performed. The constructed genetic linkage map of ‘Harehime’ consisted of 442 single nucleotide polymorphisms (SNPs) on 9 linkage groups (LGs) and covered 635.8 cM of the genome, while that of ‘Yoshida’ ponkan consisted of 332 SNPs on 9 LGs and covered 892.9 cM of its genome. We identified four QTLs associated with fruit weight, one QTL associated with sugar content, three QTLs associated with peel puffing, and one QTL associated with water rot. For these QTL regions, we estimated the haplotypes of the crossed parents and verified the founding cultivars that these QTLs were originated from and their inheritance in descendant cultivars using pedigree information. QTLs identified in this study provide useful information for marker-assisted breeding of citrus in Japan.     

Nematode–citrus plant interactions: host preference,damage rate and molecular characterization of Citrus root nematode Tylenchulus semipenetrans

• Citrus plants are host to several plant parasitic nematodes (PPNs), which are microscopic organisms. Among PPNs, the citrus root nematode, T. semipenetrans (Cobb 1913) (Tylenchida: Tylenchulidae), causes significant damage to citrus plantations worldwide. Understanding citrus nematode populations, precise identification, host preference among citrus species, and damage threshold are crucial to control T. semipenetrans. The minutiae of citrus plant–nematode interactions, nematode density and molecular nematode identification are not well understood. In this study, nematode species and density in citrus orchards, host specialization, molecular and morphological characteristics of nematodes were assessed.
• Molecular and morphological methods, host–nematode interactions, host (citrus species) preference, damage economic threshold (ET), and economic injury level (EIL) were determined using laboratory methods and field sampling. Citrus plantations in different provinces in the Mediterranean region of Turkey were investigated.
• Nematode species were identified molecularly and morphologically. ITS sequences revealed that samples were infected by citrus root nematode T. semipenetrans. The lowest nematode density was in C. reticulata in Mersin (53 2nd stage juveniles (J2s) 100 g⁻¹ soil), while the highest density was from Hatay in C. sinensis (12173 J2s 100 g⁻¹ soil). Highest citrus nematode population density was on roots of C. reticulata, followed by C. sinensis, C. limon, and C. paradisi.
• The citrus nematode is more common than was thought and population fluctuations change according to specific citrus species. Environmental conditions, host and ecological factors, such as temperature, soil pH, and soil nutrients, might influence nematode populations in citrus orchards. Investigating nematode density in diverse soil ecologies and the responses of different resistant/tolerant citrus species and cultivars to nematode populations is essential in future studies.

     

Production and molecular characterization of two new Citrus somatic hybrids for scion improvement

Somatic hybridization by protoplast fusion from cell suspension cultures and leaf parent has been a well-established technique holding great potential for citrus variety improvement. In this study, somatic hybrid plants were regenerated from the following two fusion combinations: ‘Murcott’ tangor (Citrus reticulata Blanco × C. sinensis (L.) Osbeck) + Hirado Buntan Pink pummelo (HBP) (C. grandis (L.) Osbeck) and ‘Bingtang’ orange (C. sinensis (L.) Osbeck) + Calamondin (C. microcarpa Bunge). Somatic hybrids were selected at an early stage based on their higher capacity for embryogenesis comparing to non-hybrid cells. Flow cytometry analysis showed that all plants from pre-selected lines of the two combinations were tetraploid. SSR analysis confirmed their hybrid nature, with nuclear DNA from both fusion parents, and absence of parental specific bands was also detected. Cytoplasmic compositions of the recovered plants were further revealed by CAPS and cpSSR analysis. The allotetraploid somatic hybrids from the ‘Murcott’ tangor + HBP combination will be applied to develop triploid seedless cultivars by interploid crossing with diploid seedy citrus cultivars, and those from ‘Bingtang’ orange + Calamondin could be valuable for Asiatic citrus canker-tolerant and ornamental citrus breeding.     

Proteomic changes of Citrus roots in response to long-term manganese toxicity

Key message

Fifty-three and thirty-nine differentially expressed protein spots were isolated from Mn-toxic Citrus sinensis and Citrus grandis roots, respectively. Mn-toxicity-induced changes in protein profiles greatly differed between the two species.

Abstract

Limited information is available on the manganese (Mn)-toxicity-responsive proteins in plant roots. ‘Sour pummelo’ (Citrus grandis) and ‘Xuegan’ (Citrus sinensis) seedlings were irrigated for 17 weeks with 2 (control) or 600 μM (Mn-toxic) MnSO₄. C. sinensis displayed more tolerance to Mn-toxicity than C. grandis, which may be related to more Mn accumulation in roots and less Mn distribution in shoots. Using two-dimensional electrophoresis (2-DE), we isolated 11 up-regulated and 42 down-regulated protein spots from Mn-toxic C. sinensis roots, and 25 up-regulated and 14 down-regulated protein spots from Mn-toxic C. grandis roots. This indicated more metabolic flexibility in C. sinensis roots, thus contributing to the Mn-tolerance of C. sinensis. According to the biological functional properties, these differentially expressed proteins in the two species were classified into the following categories: protein metabolism, nucleic acid metabolism, carbohydrate and energy metabolism, stress responses, cell wall and cytoskeleton, cell transport, signal transduction and fatty acid metabolism. Under Mn-toxicity, proteins involved in nucleic acid metabolism, glycolysis and cell transport were up-regulated in nontolerant C. grandis roots, and down-regulated in tolerant C. sinensis roots. The notable down-regulation of proteins in Mn-toxic C. sinensis roots with less accumulation of carbohydrates may provide an advantage to the net carbon balance by lowering related metabolic processes, and enhancing the Mn-tolerance of C. sinensis. To conclude, there are many important differences in Mn-toxicity-induced changes in protein profiles and metabolic responses between the two species.     

Behavioral responses of the parasitoid Psyllaephagus pistaciae (Hymenoptera: Encyrtidae) to host plant volatiles and honeydew

The behavioral responses of the parasitoid Psyllaephagus pistaciae, the major biocontrol agent of the common pistachio psylla, Agonoscena pistaciae, to volatiles emanating from its host plant and host honeydew, were examined using a four‐arm airflow olfactometer. In addition, the arrestment behavior of this parasitoid on clean and honeydew‐treated leaves of the pistachio, Pistacia vera, was monitored. The infested pistachio leaves were the most favored source of the volatile attracting the parasitoids. The parasitoid clearly distinguished and responded to infochemicals emitted by psyllid honeydew but at a lower level than to the volatiles from infested host plants. However, the searching time, locomotory behavior, antennal drumming and ovipositor probing were all affected when they encountered honeydew‐contaminated zones on pistachio leaves. These findings suggest that the psyllid honeydew releases kairomones that stimulate the parasitoids to greater searching activity, as well as providing a directional cue. The intensive searching activities in the presence of the volatiles tested were very similar to responses by the parasitoid females when encountering patches treated with psyllid honeydew. Such behavior could retain the parasitoid in a favorable area, thereby increasing the probability of additional host encounters.