Pomelo Research Articles

Evaluation of genotoxicity of dry powders of pomelo (Citrus maxima) flowers by micronucleus assay

Plants have been used as traditional medicine or health products for several thousands of years. The present study was aimed to evaluate the genotoxicity of pomelo flower powders by micronucleus assay In vivo. During the In vivo genotoxicity-evaluated experiment, the experimental animal’s clinical behavior, body weight (BW), food consumption, and the percentage of RET/RBCs (reticulocytes/red blood cells) and MN-RET/RETs (micronucleated reticulocytes/reticulocytes) after the treatments of pomelo (Citrus maxima) flower powders were evaluated. Both sexes ICR mice were treated three daily treatments by intraperitoneal injection of 2 mg/kg of mitomycin C (genotoxicity induction) or by oral route of 200 μL of PBS (the normal control group). Until 30th hours after the last treatment, K2-EDTA-anticoagulated peripheral blood specimens were collected. These blood samples were processed for the microscopy-based analysis using Giemsa stain and the percentage of reticulocytes and micronucleated reticulocytes was determined. The results were shown that the experimental animal’s clinical behaviors were normal in all groups. The BW and food consumption were no significant difference between all groups. RET/RBCs (%) in male or female ICR mice in the negative control group, the normal control group, the low dose of pomelo (C. maxima) flower powder group, the middle dose of pomelo (C. maxima) flower powder group, and the high dose of pomelo (C. maxima) flower powder group were respectively 8.8 ± 2.3 / 9.6 ± 2.6, 23.0 ± 2.5 / 22.4 ± 2.3, 23.4 ± 2.1 / 23.2 ± 3.8, 24.2 ± 3.6 / 23.0 ± 1.9, and 21.6 ± 3.2 / 21.6 ± 2.4; MN-RET/RETs (‰) in male or female ICR mice in the negative control group, the normal control group, the low dose of pomelo (C. maxima) flower powders group, the middle dose of pomelo (C. maxima) flower powder group, and the high dose of pomelo (C. maxima) flower powder group were 43.0 ± 12.5 / 39.4 ± 9.8, 2.6 ± 1.5 / 2.6 ± 1.5, 2.4 ± 1.1 / 2.2 ± 1.3, 2.2 ± 1.3 / 2.0 ± 1.2, and 1.8 ± 0.8 / 1.8 ± 0.8, respectively. Both RET/RBCs (%) and MN-RET/RETs (%) in male or female ICR mice in the negative control group were significantly difference than the other groups (p < 0.001). Taken all results together, pomelo (C. maxima) flower powders were without genotoxicity. Therefore, pomelo (C. maxima) flower powders were safety.

Product Flow and Challenges in the Pomelo (Citrus maxima) Industry in Northern Mindanao, Philippines

The high demand for Davao pomelo and its suitability to the local climate in the Philippines have given rise to expansion potentials and provided possibilities in curbing poverty levels in the agricultural sector of the region. In order to design intervention projects and policies, the present study investigated the pomelo industry in Northern Mindanao covering specifically the product flow and challenges in the industry. The study area was four provinces of Northern Mindanao, namely Bukidnon, Lanao del Norte, Misamis Occidental, and Misamis Oriental. The data were gathered using a semi-structured questionnaire with 22 respondents composed of 9 pomelo producers and 13 traders. The majority of the pomelo products that were sold in the markets in Northern Mindanao come from outside the region, such as Davao and Cotabato provinces, while pomelo produced in the region were sold outside the region. Among the three marketing chains of pomelo, the producer to consumer is considered the most efficient chain based on the estimated losses. Challenges in the pomelo industry include the lack of technical knowledge among the producers, poor quality of the pomelo, high buying price, and information asymmetry. Improvement in the industry could be enhanced by addressing the challenges identified.

Application of Citrus maxima Burm. Essential Oil in Shower Gel Products

Nowadays, the trend of cosmetics containing natural ingredients is becoming one of the most popular trends in the beauty industries. More and more people are interested and looking for cosmetic products made from natural ingredients such as herbs, essential oils, fruits, vegetables, etc. Essential oils are widely incorporated in cosmetic, perfumery and related household products due to their diverse properties but mainly due to their pleasant aroma and biological activity. Citrus maxima essential oil contains D-limonene as the main compound (85-95%) and other compounds such as α-pinene, sabinene, β-mycrene, α-phellandrene, σ-cymene, β-ocimene, γ- terpinene, β-caryophyllen, β-copaene. These compounds further diversify the applications of the added product. In this study, the factors affecting the production process of Citrus maxima essential oil shower gel including main detergents, moisturizers, emulsifiers, preservatives were investigated to propose production formulas. The results showed that the main detergent accounted for 12%, co- detergent 2.0%, emulsifier 1.5%, foaming agent 7.0%, surfactant, emulsifier 3.0%, moisturizer 2.0%, preservative management 2.0% and Citrus maxima essential oil account 1.0% in the formula. The evaluations of the basic physico-chemical properties are in the permissible range. Thus, it shows that the shower gel products from Citrus maxima essential oil can be produced and marketed.

Numerical optimization of hydrothermal liquefaction process for the production of bio-oil and bio-char from Citrus maxima peel waste and product characterization.

This research study aims to potential utilization of Citrus maxima peel waste and optimize the hydrothermal liquefaction process for the production of bio-oil (BO) and bio-char (BC). The effect of several HTL processing variables on BO yield (%) and BC yield (%), including temperature, retention period, and slurry concentration, has been examined using central composite design (CCD) (a three-level three-factor design). The optimized values of HTL process variables were found to be 240°C (temperature), 52min (retention time), and 7% (slurry concentration) and the corresponding responses were 5.794% (BO yield) and 29.450% (BC yield). The values obtained from the RSM-CCD model as the predicted values agreed with the experimental values (5.93% and 30.14%). Further the BO and BC obtained under optimized conditions and CPP were analyzed to identify the variations by 1H-NMR, GC-MS, FT-IR, and CHNO-S.

Analysis of Pharmacological Activities and Mechanisms of Essential Oil in Leaves of C. grandis 'Tomentosa' by GC-MS/MS and Network Pharmacology.

Citrus grandis 'Tomentosa,' a fruit epicarp of C. grandis 'Tomentosa' or C. grandis (L.) Osbeck is widely used in health food and medicine. Based on our survey results, there are also rich essential oils with bioactivities in leaves, but the chemical compounds in this part and relevant pharmacological activities have never been studied systematically. Therefore, this study was to preliminarily decipher the pharmacological activities and mechanisms of the essential oil in leaves of C. grandis 'Tomentosa' by an integrated network pharmacology approach. Essential oil compositions from leaves ofC. grandis 'Tomentosa' were identified using GC-MS/MS. And then, the targets of these oil compositions were predicted and screened from TCMSP, SwissTargetPrediction, STITCH and SEA databases. STRING database was used to construct the protein-protein interaction networks, and the eligible protein targets were input into WebGestalt 2019 to carry out GO enrichment and KEGG pathway enrichment analysis. Based on the potential targets, disease enrichment information was obtained by TTD databases. Cytoscape software was used to construct the component-target-disease network diagrams. Finally, 61 essential oil chemical components were identified by GC-MS/MS, which correspond to 679 potential targets. Biological function analysis showed 12, 19, and 12 GO entries related to biological processes, cell components and molecular functions, respectively. 43 KEGG pathways were identified, of which the most significant categories were terpenoid backbone biosynthesis, TNF signaling pathway and leishmaniasis. The component-target-disease network diagram revealed that the essential oil compositions in leaves of C. grandis 'Tomentosa' could treat tumors, immune diseases, neurodegenerative diseases and respiratory diseases, which were highly related to CHRM1, PTGS2, CASP3, MAP2K1 and CDC25B. This study may provide new insight into C. grandis 'Tomentosa' or C. grandis (L.) Osbeck and may provide useful information for future utilization and development.

Volatile compounds of grapefruit (Citrus Grandis (L.) Osbeck) peel essential oil by cold pressing and hydrodistillation methods

The study of the essential oils (EOs) extraction with high medicinal properties is necessary to contribute to the use of natural compounds. By using cold pressing and hydrodistillation methods, the content of grapefruit peel (Citrus grandis (L.) Osbeck) EOs was extracted. The chemical components of EOs was determined by GC-MS method. Results have shown that 10-12 compounds were detected in the grapefruit peel EOs, which are mainly monoterpenes such as D-Limonene (71.21%-74.83% and γ-Terpinene (15.70%-17.68%). The findings from the present study demonstrated a new research direction on chemical composition of the grapefruit EO, thereby contributing to applications ofthe natural compounds with high medicinal properties from grapefruit.

Pomelo (Citrus maxima. Merr) essential oil extraction by cold pressing and hydrodistillation methods and comparison of chemical composition

Essential oils extraction from the shell bring many applications in life, as well as contribute to the development of the food flavoring and cosmetic industry. Pomelo (Citrus maxima Merr) essential oil (EO) was extracted by cold pressing and hydrodistillation methods. The EO content obtained was 0.067% and 0.33%, respectively. The chemical composition was determined by the GC-MS analysis method, with 4 main components being D-Limonene, β-Myrcene, α-Pinene, and α-Phellandrene which exhibited high antibacterial and antioxidant capacity). The EO obtained from pomelo peels by cold pressing conditions was limited and less effective with more natural aroma, as compared to the hydrodistillation methods. Therefore, further research on EO pressing equipment and methods that can be used for effective oil extraction and recovery are necessary.

Green synthesis of nano-silver and its antibacterial activity against methicillin-resistant Staphylococcus aureus

Superbugs resistant to antimicrobials are swiftly expanding worldwide. One of these superbugs for which effective antibiotics are urgently needed is methicillin-resistant Staphylococcus aureus (MRSA). Thus, the creation of novel antimicrobial substances is necessary, one of the appealing antibacterial inorganic materials is silver nanoparticles (AgNPs) for application of treating bacterial infectious illnesses. In order to investigate novel, potent, and economically feasible therapeutic approaches, the current study presents a cost-effective and environmentally friendly technique for synthesizing AgNPs using Citrus maxima peel (CMP) extract as a reducing agent. UV–Vis Spectroscopy confirmed the formation of AgNPs in the 400–––500 nm wavelength range. The Powder X-ray diffr action (PXRD) and high-resolution transmission electron microscopy (HRTEM) results at the optimized synthesis conditions revealed highly crystalline AgNPs (face-centered cubic structure) with particles the size of 10–––20 nm. The Fourier transform infrared spectroscopy (FT-IR) revealed the presence of flavonoids, terpenoids, phenolics, and glycosides in the phytochemical compositions of the CMP extract, which can serve as reducing agents for the formation of the spherical AgNPs. Minimum inhibitory concentration (MIC) values of 8.27 µg/mL, minimum bactericidal concentration (MBC) values of 16.54 µg/mL, and an inhibition zone of 11.7 mm were indicative of the potent antibacterial activity of as-prepared AgNPs against MRSA. The findings suggest biogenic silver nanoparticles could be an effective antimicrobial agent against nosocomial infections.

Development and pathogenicity analysis of full-length infectious cDNA clones of citrus yellow mottle-associated virus in citrus plants

Citrus yellow mottle-associated virus (CiYMaV) belonging to the subgenus Mandarivirus within the genus Potexvirus, was first identified in 2018 from Pakistan (CiYMaV-PK), where it is endemic in several regions. Here, three full-length cDNA clones (pCiYMaV-FL-1, pCiYMaV-FL-18, and pCiYMaV-FL-22) corresponding to the genomic RNA of CiYMaV were constructed and then agroinfiltrated on Chandler pummelo (Citrus grandis) seedlings using the vacuum infiltration method. All the inoculated plants developed severe vein yellowing, leaf mottling, and dwarfing symptoms by 40 days post-infiltration (dpi). The results of a direct tissue blot immunoassay and reverse transcription polymerase chain reaction detection showed 94.7–100% infection rates of pCiYMaV-FL at 60 dpi. Despite there being no observed difference among the three clones in the severity of symptom, pCiYMaV-FL-22 showed the highest accumulation levels of viral RNA and coat proteins. Moreover, pCiYMaV-FL-22 successfully infected seven other citrus varieties and induced symptoms in five of them. Transmission electron microscopy identified the presence of filamentous virus particles in extracts from systemic leaves of the plants infected with pCiYMaV-FL-22 at 6-months post-infiltration. Taken together, the results indicate that Koch's postulates were fulfilled for CiYMaV in citrus plants. This is the first report of full-length infectious cDNA clones of CiYMaV, and thus, the data provide a basis for further study of the molecular mechanisms of virus infection and the development of a viral vector to express foreign genes in citrus plants.

Influence of the loading regime on the uniaxial compressive behaviour of density graded Citrus Maxima peel

To conceive more efficient protective structures, it is possible to draw inspiration from natural structures. However, the origin of the mechanical absorption properties of natural structures is not always clear. Among the multitude of existing natural structures, the density graded peel of the Citrus Maxima was studied in this work to characterize its mechanical behaviour under uniaxial compression, from the quasi-static regime to the dynamic regime. The resulting behaviour is very different from a classical foam behaviour as no linear part is observed for small strains. Furthermore, the mechanical behaviour is deeply influenced by the loading regime: a stair-case response appears under dynamic loading.

Determination of the chemical and mineral composition of citrus by-products in relation to its utilization as a feed raw material

The global food industry and especially fruit juice production generates millions of tons of organic waste annually. Citrus waste, also known as citrus pulp оr citrus pomace, is a residue obtained during various citrus juices production. This by-product has the potential to become an excellent source of essential nutrients in animal feeding. In the current study, the chemical and mineral composition were determined in the citrus waste by Orange (Citrus sinensis), Lemon (Citrus limon), Red grapefruit (Citrus paradisi), Mandarin (Citrus reticulata), Lime (Citrus aurantifolia), Pomelo (Citrus maxima). Waste products are derived from the production factory of freshly squeezed fruit and vegetable juices located in the village of Musachevo, Western Bulgaria. For all citrus waste, the following indicators were determined: dry matter (DM), moisture (RH), crude protein (CP), crude fibre (CF), crude ash (CA), ether extract (EE), manganese (Mn), zinc (Zn), magnesium (Mg), calcium (Ca), phosphorus (P), sodium (Na), potassium (K). The mineral content was determined by Atomic Absorption Spectrophotometer. Their chemical and mineral composition analyses showed differences in the distribution of nutrients and minerals. The data obtained from the conducted analyses showed that citrus pulp could successfully replace some of the main raw materials in the animal’s ration, as it is a source of essential nutrients for it.

Dependency of Crops on Pollinators and Pollination Deficits: An Approach to Measurement Considering the Influence of Various Reproductive Traits

Pollinators provide crucial ecosystem services, i.e., pollination, which determines crops’ reproductive fitness and yield. As pollinators decline, flowering crops might face pollination-deficit stress depending on their dependence on pollinators and pollinator availability. Here, we assessed the dependency of some crops (belonging to diverse plant families) on biotic pollinators based on their maximum reproductive potential in a supplementary pollination treatment and minimum reproductive success in a pollinator exclusion treatment. Additionally, we determined the pollen transfer limitation of the crops in open field conditions. We also determined the influence of the different reproductive traits with the index of dependency of crops on pollinators (IDP) and the coefficient of pollination deficit (D). Based on the values of IDP, members of Cucurbitaceae are obligatorily dependent on pollinators for their fruit set. Members of Brassicaceae and Rutaceae are highly reliant on pollinators. A few crops, like Lablab purpureus and Nigella sativa, are less dependent on pollinators. In open field conditions, most crops have a low pollination deficit, some without pollen transfer limitations, and only a few crops (Citrus × limon and Citrus maxima) show a higher pollination deficit. The IDP is negatively influenced by the pollen–ovule ratio, which also negatively affects the pollination deficit of the crops. This study will be useful in understanding and mitigating the effects of pollinator losses, as well as in choosing crops (those under pollination deficit stress and largely dependent on pollinators for fruit set) for supplemental pollination services to increase agricultural production.

Aqueous extract of pummelo pulp (Citrus maxima) improves the biochemical profile and reduces the inflammation process in Wistar rats with non-alcoholic fatty liver disease.

This study investigated the effect of pummelo extract (Citrus maxima) on biochemical, inflammatory, antioxidant and histological changes in NAFLD rats. Forty male Wistar rats divided into four groups were used: (1) control group; (2) fructose associated with high-fat diet - DHF; (3) normal diet+pummelo extract (50mg/kg); and (4) FHD+pummelo extract. This was administered at dose of 50mg/kg of the animal's weight, by gavage, for 45 days. Significant improvement in lipid profile, liver and kidney function, inflammation, oxidative stress markers was identified in group 4 compared to group 2. Regarding TNF-α and IL-1β, group 2 showed higher values (respectively 142, 5±0.7 and 560.5±2.7pg/mg protein) compared to group 4 (respectively 91.4±0.9 and 402.1.4±0.9pg/mg protein), p<0.05. Significant increases were found in SOD and CAT activities, respectively 0.10±0.06 and 8.62±1.67 U/mg protein for group 2 and respectively 0.28±0.08 and 21.52±2.28 U/mg of protein for group 4. Decreases in triglycerides, hepatic cholesterol and fat droplets in hepatic tissue were observed in group 4 compared to group 2. Results highlight that pummelo extract may be useful for prevent the development of NAFLD.

Green synthesis of CuO nanoparticles using citrus maxima peel aqueous extract

Discarded fruit peels caused pollution problem and reused of waste product reduces pollution and sustain environmental balance. In most cases, fruits peels are land filled or composted which lead to environmental problems like methane emission to the atmosphere due to anaerobic digestion and leachate formation, which percolates into the bottom of the earth during rainfall. This communication reports on the synthesis of copper oxide nanoparticles (CuO NPs) using citrus maxima peel. The prepared CuO NPs were characterized by XRD, SEM, EDS, UV–visible spectroscopy, FTIR and PL. The characterization results confirmed that the prepared CuO NPs have a monoclinic crystalline structure with a spherical shape and the average crystallite size of CuO NPs was about 20 nm. The optical band gap (Eg) of CuO NPs calculated from UV–Visible diffuse reflectance spectroscopy was 1.5eV. The photo response V-I characteristics of CuO NPs showed good photo response under illumination conditions, which might be due to the maximum number of exciting photoelectrons due to illumination.

Species Account and Diversity of Indigenous Medicinal Plants in Rogongon, Iligan City, Lanao Del Norte

The documentation of indigenous medicinal plants in Rogongon, Iligan City, Lanao del Norte was assessed for species composition, diversity, and dominance using the transect method.The study aimed to determine the species accounts of indigenous medicinal plants in the area of Rogongon, Iligan City. A total of twenty (20)medicinal plant species were counted and identified namely, Annona muricata, Carica papaya, Ipomoea batatas, Senna alota, Leucaenaleucocephala, Gliricidiasepium, Origanumvulgare, Plectranthusscutellarioides, Perseaamericana, Theobroma cacao, Ficusseptica, Musa spp., Psidiumguajava, Peperomiapellucida, Eleusineindica, Cymbopogoncitratus, Coffeaarabica, Citrus maxima, Capsicum annuum and Curcuma longa. The botanical families with the highest numberof species were represented by the families Fabaceae (20.39%)followed byMoraceae(15.53%) andMusaceae (13.59%). The information recorded and presented in this study may be useful for achieving ecologically-stable medicinal flora associates in the future that may provide economic benefits to the community.

The adaptation of root cell wall pectin to copper toxicity in two citrus species differing in copper tolerance: remodeling and responding.

Citrus species are prone to suffer from copper (Cu) toxicity because of improper application of Cu-based agrochemicals. Copper immobilization mediated by pectin methylesterase (PME) in the root cell wall (CW) is effective for Cu detoxification. However, the underlying mechanisms of the structural modification and stress responses of citrus root CW pectin to Cu toxicity have been less discussed. In the present study, seedlings of 'Shatian pummelo' (Citrus grandis L. Osbeck) and 'Xuegan' (Citrus sinensis L. Osbeck), which differ in Cu tolerance, were irrigated with nutrient solution containing 0.5 (as control), 100, 300 or 500μM Cu for 18 weeks in sandy culture or 24h in hydroponics. At the end of treatments in the 18-week sandy culture, Cu toxicity on CW pectin content, Cu distribution, degree of pectin methylesterification (DPM) and the PME enzyme activity were discussed. At the genome-wide level, PME gene family was identified from the two citrus species, and qRT-PCR array of citrus PMEs under control and 300 μM Cu stress for 18 weeks were performed to screen the Cu-responsive PME genes. Moreover, the candidate genes that responded to Cu toxicity were further examined within 24h. The results showed that Cu toxicity increased the root CW pectin content. The root CW pectin under Cu toxicity was remodeled by upregulation of the expression of the Cu-responsive PME genes followed by increasing PME activity, which mainly promoted low methylesterased pectin level and the Cu content on root CW pectin. Compared with C. sinensis, C. grandis root CW had a lower DPM and higher Cu content on the Cu-stressed root CW pectin, contributing to its higher Cu tolerance. Our present study provided theoretical evidence for root CW pectin remodeling in response to Cu toxicity of citrus species.

Involvement of Vacuolar Processing Enzyme CgVPE1 in Vacuole Rupture in the Programmed Cell Death during the Development of the Secretory Cavity in Citrus grandis 'Tomentosa' Fruits.

Vacuolar processing enzymes (VPEs) with caspase-1-like activity are closely associated with vacuole rupture. The destruction of vacuoles is one of the characteristics of programmed cell death (PCD) in plants. However, whether VPE is involved in the vacuole destruction of cells during secretory cavity formation in Citrus plants remains unclear. This research identified a CgVPE1 gene that encoded the VPE and utilized cytology and molecular biology techniques to explore its temporal and spatial expression characteristics during the PCD process of secretory cavity cells in the Citrus grandis 'Tomentosa' fruit. The results showed that CgVPE1 is an enzyme with VPE and caspase-1-like activity that can self-cleave into a mature enzyme in an acidic environment. CgVPE1 is specifically expressed in the epithelial cells of secretory cavities. In addition, it mainly accumulates in vacuoles before it is ruptured in the secretory cavity cells. The spatial and temporal immunolocalization of CgVPE1 showed a strong relationship with the change in vacuole structure during PCD in secretory cavity cells. In addition, the change in the two types of VPE proteins from proenzymes to mature enzymes was closely related to the change in CgVPE1 localization. Our results indicate that CgVPE1 plays a vital role in PCD, causing vacuole rupture in cells during the development of the secretory cavity in C. grandis 'Tomentosa' fruits.

Ethanol Extract of Citrus grandis 'Tomentosa' Exerts Anticancer Effects by Targeting Skp2/p27 Pathway in Non-Small Cell Lung Cancer.

This study aims to investigate the anticancer properties of Citrus grandis 'Tomentosa' (CGT) in non-small cell lung cancer (NSCLC). The ethanol extract of CGT (CGTE) is prepared by using anhydrous ethanol and analyzed by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS), revealing that the main chemical components in CGTE are flavonoids and coumarins, such as naringin, rhoifolin, apigenin, bergaptol, and osthole. CGTE at concentrations without inducing cell death significantly inhibits cell proliferation via inducing cell cycle G1 phase arrest by MTT, colony formation, and flow cytometry assays, implying that CGT has anticancer potential. CGTE markedly inhibits the activity of Skp2-SCF E3 ubiquitin ligase, decreases the protein level of Skp2, and promotes the accumulation of p27 by co-immunoprecipitation (co-IP) and in vivo ubiquitination assay; whereas Skp2 overexpression rescues the effects of CGTE in NSCLC cells. In subcutaneous LLC allograft and A549 xenograft mouse models, CGTE, without causing obvious side effects in mice, significantly inhibits lung tumor growth by targeting the Skp2/p27 signaling pathway. These findings demonstrate that CGTE efficiently inhibits NSCLC proliferation both in vitro and in vivo by targeting the Skp2/p27 signaling pathway, suggesting that CGTE may serve as a therapeutic candidate for NSCLC treatment.

Elucidation of flavanones, phenols and antioxidant capacity influenced by drying methods from physiologically dropped underutilized Citrus grandis fruits.

Nutritional content in citrus fruit is enormous. Citrus grandis (L.) Osbeck is underutilised citrus crop that receives little attention due to the lack of knowledge regarding its nutritional value. Citrus waste disposal poses a problem due to economic and environmental factors. The metabolites flavonoids, phenols and antioxidant capacity in the dropped fruits of the underutilised citrus species pomelo (Citrus grandis (L.) Osbeck) were examined. Hesperidin varied from 1.22 to 2.83% and 1.08 to 1.16% from 10 mm to 14 mm whereas naringin dominates in fruits measuring 10 mm and 12mm with 60.61%, 60.77%, and 47.76%, 45.87% in freeze dried (FD) and hot air oven dried (HAOD) samples. According to the results of the antioxidant assays, the highest concentrations of ABTS azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) and DPPH (2, 2-diphenyl-1-picrylhydrazyl radical) were found in freeze dried samples, ranging from 9.679 to 10.416 mmol L-1 Trolox and 14.825 to 16.432 mmol L-1 Trolox, respectively. However, the Ferric Reducing Antioxidant Power (FRAP) assay revealed higher content in samples of both FD and HAOD that were 10mm in size (4.578 mmol L-1 Trolox and 3.730 mmol L-1 Trolox). Total phenol content was measured, and the highest concentrations were found in fruits with a diameter between 10 mm and 18 mm. It ranged from 48.479 to 54.498 mg GAE L-1 in FD samples and from 45.757 to 51.159 mg GAE L-1 in HAOD samples. The smallest fruits, or those that were still in the immature stage, had the highest content. It was found that when the immature dropped fruits were dried by HAOD, the content decreased. At p<0.01 and p<0.05, there was a significant positive correlation between the flavonoids, antioxidants, and total phenols. The results showed that the immature dropped immature fruits of lesser known underutilised citrus sp. Citrus grandis can act as potential source of flavonoids, total phenol concentration, and antioxidant potential. Freeze drying can be recommended to recover the most bioactive substances from physiologically dropped fruits of Citrus grandis for use in the pharmaceutical and nutraceutical sectors. This study will help in reducing the environmental impact caused due to citrus dropped fruits and its responsible management.

Comprehensive genome-wide identification and transferability of chromosome-specific highly variable microsatellite markers from citrus species

Citrus species among the most important and widely consumed fruit in the world due to Vitamin C, essential oil glands, and flavonoids. Highly variable simple sequence repeats (SSR) markers are one of the most informative and versatile molecular markers used in perennial tree genetic research. SSR survey of Citrus sinensis and Citrus maxima were identified perfect SSRs spanning nine chromosomes. Furthermore, we categorized all SSR motifs into three major classes based on their tract lengths. We designed and validated a class I SSRs in the C. sinensis and C. maxima genome through electronic polymerase chain reaction (ePCR) and found 83.89% in C. sinensis and 78.52% in C. maxima SSRs producing a single amplicon. Then, we selected extremely variable SSRs (> 40 nt) from the ePCR-verified class I SSRs and in silico validated across seven draft genomes of citrus, which provided us a subset of 84.74% in C. sinensis and 77.53% in C. maxima highly polymorphic SSRs. Out of these, 129 primers were validated on 24 citrus genotypes through wet-lab experiment. We found 127 (98.45%) polymorphic HvSSRs on 24 genotypes. The utility of the developed HvSSRs was demonstrated by analysing genetic diversity of 181 citrus genotypes using 17 HvSSRs spanning nine citrus chromosomes and were divided into 11 main groups through 17 HvSSRs. These chromosome-specific SSRs will serve as a powerful genomic tool used for future QTL mapping, molecular breeding, investigation of population genetic diversity, comparative mapping, and evolutionary studies among citrus and other relative genera/species.