Valencia Orange Peel Research Articles

Identification of Polymethoxyflavones (PMFs) from Orange Peel and Their Inhibitory Effects on the Formation of Trimethylamine (TMA) and Trimethylamine-N-oxide (TMAO) Using cntA/B and cutC/D Enzymes and Molecular Docking.

This study investigates the inhibitory effects of polymethoxyflavones (PMFs) on enzymes involved in the production of trimethylamine (TMA) and trimethylamine-N-oxide (TMAO). PMFs were isolated from Valencia orange peel and identified using column separation and NMR techniques. The findings reveal that nobiletin and 3,6,7,8,2',5'-hexamethoxyflavone significantly suppress cntA/B and cutC/D, respectively. Furthermore, 3,6,7,8,2',5'-hexamethoxyflavone decreases the level of TMAO formation by suppressing the FMO3 mRNA level. This study elucidates that specific structural features of PMFs can contribute to their interactions with enzymes. Our study represents the first demonstration of the ability of PMFs to mitigate the risk of cardiovascular disease (CVD) by inhibiting enzymes responsible for TMA production, which are generated by gut microbiomes. Furthermore, we introduce a novel model system utilizing TMA-induced HepG2 cells to assess and compare the inhibitory effects of PMFs on TMAO production. These findings could pave the way for the development of novel therapeutic approaches to manage CVD.

Comparative GC Analysis of Valencia orange Ripe and Unripe Peel Essential Oils, Nano-formulation, Anti-Helicobacter Pylori and Anti-inflammatoryEvaluation: in vitro and in silico

Peptic ulcer associated with H. pylori infection and antibiotics resistance warranted the execution of the foregoing study to produce natural and safe alternatives. GC-MS analysis of the essential oil derived from the ripe and unripe peel of Citrus sinensis var. Valencia grown in Egypt was investigated besides its anti-Helicobacter pylori and COX-2 inhibition actions. Ripe mature peels displayed a higher oil yield than unripe ones (0.8 and 0.6%, respectively). Twenty-nine compounds were identified in all samples with volatile monoterpenes as the predominant class, (88.90% and 90.62% in fresh ripe peel (P1) and dried ripe peel (P2), respectively, however unripe fresh orange peel (P3) showed a lower quantity (83.95%). D-Limonene was the major compound in P1 and P2 (85% and 88 %, respectively), while 73% were found in P3. In comparison to standard Clarithromycin (MIC90 = 1.95 ± 0.002 μg/ml), the nano-hexosomal formula of P1 reduced H. pylori action more than P1 alone with MIC90= 1.95 ± 0.0029 μg/ml and MIC90= 3.90 ± 0.006 μg/ml for formulated and unformulated P1, respectively. An anti-inflammatory assay via COX-2 inhibition revealed that the formulated P1 is more potent (IC50= 0.85 μg/ml) than P1 alone (IC50= 2.04 μg/ml) in comparison with Celecoxib® (IC50= 0.28 μg/ml). To unravel the mechanism of the anti-Helicobacter pylori activity, an in silico study on D-Limonene has been performed, using H. pylori urease enzyme, which revealed that D-Limonene showed promising binding scores. Thus, Valencia orange peel essential oil offers a natural, cheap, and relatively safe candidate to combat Helicobacter pylori infection.

Potential Use of Low-Cost Agri-Food Waste as Biosorbents for the Removal of Cd(II), Co(II), Ni(II) and Pb(II) from Aqueous Solutions

We evaluated the potential use of agri-food waste for the removal of heavy metal ions from aqueous solutions and its application in different processes (e.g., water remediation, in the production of biomass enriched in nutritionally significant elements, etc.). Biomasses from grape seed, grape pomace, loquat seed, Calabrese broccoli stem, empty pods of carob and broad bean pods, unripe bitter orange peel, kumquat, orange pulp and Canary Island banana pulp were prepared. The percentages and biosorption capacities were evaluated and compared with those refe-renced using Valencia orange peel (Citrus sinensis Valencia late). These studies allow for easily providing added value to different agri-food wastes. The results show that the proposed biomasses were able to retain the studied metal ions and obtained different percentages, being in some cases above 90%. The highest values were obtained using broad bean pod (Pb(II) (91.5%), Cd(II) (61.7%), Co(II) (40.7%) and Ni(II) (39.7%)). Similar values were observed using grape seed, broccoli stem, carob pod and unripe bitter orange peel. Carob pod for biosorption of Cd(II) is also of great interest. These studies suggest that the agri-food residues evaluated can be applied to prepare effective biosorbents of divalent metal ions from aqueous solutions.

Commercial pectin production from dried florida orange peel: Effect of process conditions on pectin structure and function

To determine the feasibility of using Florida, USA, oranges for pectin production, we evaluated the effects of time, temperature, and pH on pectin properties extracted from dried Valencia orange peel using a hot acid process. Juice extracted peel was collected weekly, and following pretreatment, the dried peel was pooled and used to extract pectin with time (4, 5, and 6 h), temperature (65 and 75°C), and pH (1.6 and 1.8) as process variables. Pectin properties, including yield, sugar composition, molecular weight, intrinsic viscosity, degree of methylesterification, degree of branching, degree of blockiness, absolute degree of blockiness, and Standard Acid in Glass (SAG), were determined. In addition, we evaluated the response of each variable to the process conditions using response surface methodology. The analysis of variance (anova) models for molecular weight, intrinsic viscosity, and yield were highly significant, with actual values decreasing with increasing temperature. Increasing reaction time also decreased values for molecular weight and intrinsic viscosity. Novelty impact statement This study indicates that process time and temperature affected pectin quality, functionality, and yield. Increasing temperature had the most detrimental effect on desirability in an optimization analysis.

Biowaste-based biochar: A new strategy for fermentative bioethanol overproduction via whole-cell immobilization

This work explores the potential use of biochar as a microbial cell carrier enhancing the efficiency of alcoholic fermentations. Olive kernels, vineyard prunings, sewage sludge and seagrass residues were applied as biowaste for biochar production through pyrolysis at two different temperatures (250 °C and 500 °C), while a commercial type of non-biomass char was also employed for benchmarking purposes. Apart from vineyard prunings pyrolyzed at 250 °C, all other carbonaceous materials presented crystalline phases including halite, calcite, sylvite and/or silicon. Moreover, increase in pyrolysis temperature enhanced biochar’s porosity and BET-specific surface area, which reached 41.7 m2 g−1 for VP-based biochar remaining at lower levels (0.15–5.3 m2 g−1) in other specimens tested. Elemental analysis demonstrated reduction in oxygen and increase in the carbon content of biochars produced at elevated temperatures, while biochar from seagrass included residues of chloride (0.3–5.14%). Three major yeasts were immobilized on materials exhibiting the highest surface areas and applied in repeated batch fermentations using Valencia orange peel hydrolyzates as feedstock. The biocatalysts developed using S. cerevisiae and K. marxianus immobilized on vineyard prunings-based biochar exhibited exceptional ethanol productivities as compared to the relevant literature, which reached 7.2 g L−1 h−1 and 7.3 g L−1 h−1 respectively. Although the aforementioned strains improved biofuel production by 36–52% compared to the conventional process, P. kudriavzevii KVMP10 was not efficient following immobilization on biochar. The approach constitutes an innovative method for bioenergy production, demonstrating a novel application of biochar in industrial biotechnology which incorporates important technological advances such as enhanced biofuel production and biomass recycling.

High temperature alcoholic fermentation of orange peel by the newly isolated thermotolerant Pichia kudriavzevii KVMP10.

This work explores the potential for the development of orange peel based ethanol bioprocesses through isolation of the thermotolerant Pichia kudriavzevii KVMP10. A model solution of hydrolysed Valencia orange peel was employed to determine the ethanologenic potential of the yeast, which was maximized at 42°C producing 54gl(-1) of ethanol. The effect of orange peel oil on bioethanol formation was investigated at 30 and 42°C confirming that the minimum inhibitory peel oil content was 0·01% (v/v). Pichia kudriavzevii KVMP10 demonstrated significant technological advantages for the production of sustainable bioenergy, such as utilization of both hexoses (glucose, sucrose, fructose and galactose) and pentoses (xylose) at high temperatures, exemplifying its great potential for application in orange peel based biorefineries for ethanol production. Citrus peel waste is one of the most underutilized and geographically diverse residues in the planet. In attempt to develop a citrus peel based biorefinery we report here the isolation of a yeast which exhibited favourable technological characteristics for the production of ethanol through utilization of the specific food waste. Pichia kudriavzevii KVMP10 was highly thermotolerant and utilized both hexoses and pentoses for ethanol production, which was achieved at elevated rates, highlighting its great potential for application in ethanol production processes from citrus peel.

Comparative Structural Changes and Inactivation Kinetics of Pectin Methylesterases from Different Orange Cultivars Processed by High Pressure

Pectin methylesterases (PMEs) from two different orange sources, Valencia and Navel cv., were extracted and purified using cation exchange, heparin chromatography, and finally, gel filtration chromatography, yielding a single peak corresponding to a protein of molecular weight 34 and 35 kDa for Valencia and Navel PMEs, respectively. Effects of high pressure (HP) and thermal processing for various treatment durations on the activity of PMEs in Tris–HCl buffer solution (pH 7.5) were explored. Higher levels of pressure, temperature, and treatment duration resulted in an analogous reduction of the PME residual activity. HP-induced inactivation of both purified PMEs was described by a first-order kinetic model. Kinetic parameters were estimated and a multiparameter equation was developed to predict the PME inactivation rate constant at any combination of pressure and temperature conditions for both enzymes. The PME from Valencia orange peel appeared to be more heat- and pressure-sensitive compared to Navel PME. HP-induced conformational changes of the PME molecules were also investigated using circular dichroism (CD) spectroscopy. A direct comparison of the CD results for treated and untreated proteins reveals that pressure treatment has negligible effects upon far-UV CD spectra, while significant irreversible changes are depicted in near UV for both PMEs. It is, thus, evidenced that exposure to HP may lead to a structurally molten globulelike state, where the PME maintains a secondary structure of untreated protein molecules, while a tertiary structure is substantially affected bearing subsequent impact on substrate–enzyme binding interaction, leading to reduction of enzyme activity.

Citric acid production from orange peel wastes by solid-state fermentation

Valencia orange (Citrus sinensis) peel was employed in this work as raw material for the production of citric acid (CA) by solid-state fermentation (SSF) of Aspergillus niger CECT-2090 (ATCC 9142, NRRL 599) in Erlenmeyer flasks. To investigate the effects of the main operating variables, the inoculum concentration was varied in the range 0.5·103 to 0.7·108 spores/g dry orange peel, the bed loading from 1.0 to 4.8 g of dry orange peel (corresponding to 35-80 % of the total volume), and the moisture content between 50 and 100 % of the maximum water retention capacity (MWRC) of the material. Moreover, additional experiments were done adding methanol or water in different proportions and ways. The optimal conditions for CA production revealed to be an inoculum of 0.5·106 spores/g dry orange peel, a bed loading of 1.0 g of dry orange peel, and a humidification pattern of 70 % MWRC at the beginning of the incubation with posterior addition of 0.12 mL H2O/g dry orange peel (corresponding to 3.3 % of the MWRC) every 12 h starting from 62 h. The addition of methanol was detrimental for the CA production. Under these conditions, the SSF ensured an effective specific production of CA (193 mg CA/g dry orange peel), corresponding to yields of product on total initial and consumed sugars (glucose, fructose and sucrose) of 376 and 383 mg CA/g, respectively. These results, which demonstrate the viability of the CA production by SSF from orange peel without addition of other nutrients, could be of interest to possible, future industrial applications.

Ethanol production by Saccharomyces cerevisiae and Kluyveromyces marxianus in the presence of orange-peel oil

Two ethanologenic yeasts, Saccharomyces cerevisiae and Kluyveromyces marxianus, were used to ferment sugar solutions modeling hydrolyzed Valencia orange peel waste at 37°C. Orange stripper oil produced from orange peel was added in various amounts to determine its effect on ethanol production. The minimum peel oil concentration that inhibited ethanol production was determined after 24, 48 and 72 h and the two yeasts were compared to one another in terms of ethanol yield. Minimum inhibitory peel oil concentrations for ethanol production were 0.05% at 24 h, 0.10% at 48 h, and 0.15% at 72 h for both yeasts. S. cerevisiae produced more ethanol than K. marxianus at each time point.

Biological activity of carotenoids in red paprika, Valencia orange and Golden delicious apple

Carotenoid fractions were extracted from red paprika, Valencia orange peel and the peel of Golden delicious apple. Thus, hypophasic carotenoids of paprika (PM1), orange (PM3) and apple (PM4), and epiphasic extractions of paprika (PM2) and apple (PM5) were obtained by extraction, saponification and partition between MeOH-H(2)O (9:1) (hypophasic) and hexane (epiphasic). A high content of capsanthin was quantified in hypophasic carotenoids (PM1) from red spice paprika, whereas the hypophasic fractions from orange (PM3) and apple (PM4) were mainly composed of violaxanthin, zeaxanthin and lutein. On the other hand, a high content of beta,beta-carotene and beta-cryptoxanthin was found in epiphasic fractions (PM2 and PM5). The extracts were studied for their anti-Helicobacter pylori (H. pylori), anti-human immunodeficiency virus (HIV), cytotoxic, multidrug resistance (MDR) reversal and radical scavenging activity. Among five PM extracts and beta,betacarotene, PM4 showed potent anti-H. pylori activity (MIC(50) = 36 microg/mL), comparable to metronidazole (MIC(50) = 45 microg/mL). The extracts were inactive against HIV. PM3 and PM4 showed slightly higher cytotoxic activity against three human tumor cell lines (squamous cell carcinoma HSC-2, HSC-3, submandibular gland carcinoma HSG) and human promyelocytic leukemic HL-60 cells than against three normal human oral cells (gingival fibroblast HGF, pulp cell HPC, periodontal ligament fibroblast HPLF), suggesting a tumor-specific cytotoxic activity. PM1, PM3 and PM4 displayed much higher MDR-reversing activity than (+/-)-verapamil. ESR spectroscopy demonstrated that PM1-5 and beta,beta-carotene produced little or no detectable radical under alkaline conditions and did not scavenge the O(2) (-) produced by the hypoxanthine and xanthine oxidase reaction. On the other hand, PM1 and PM2 scavenged efficiently 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical, whereas singlet oxygen was also quenched efficiently by PM5 and PM2. The data suggest the potential importance of carotenoids as possible anti-H. pylori and MDR reversal agents. The active principles in the carotenoid extract might differ, depending upon the types of fruits and vegetables.

Characterization of a salt-independent pectin methylesterase purified from valencia orange peel.

The pectin methylesterase (PME; EC 3.1.1.11) present in a commercial orange peel enzyme preparation was characterized to establish its identity among the multiple PME isozymes present in Valencia orange (Citrus sinensis L.) peel. We show the commercial enzyme corresponds to the major peak 2 PME previously separated by heparin-Sepharose chromatography (Cameron et al., J. Food Sci. 1998, 63, 253). Both PMEs have comparable elution profiles on cation-exchange and hydrophobic-interaction perfusion chromatography columns, molecular weights (ca. 34 kDa) and pI (pH 9.2), and biochemical properties, including a broad pH activity range and activity in the absence of added cations. An identical partial amino terminal peptide sequence was also obtained for the PMEs, which further demonstrated a structural identity with other plant PMEs. The biochemical and structural properties readily distinguish this Valencia orange PME from salt-dependent isozymes and further suggest that it is an ortholog to the salt-independent fruit-specific isozyme of tomato. This work provides a well-defined, enzymatically homogeneous, salt-independent (type 1) plant PME isozyme that is suitable for studying details of the enzyme's mode of action and for use in modifying methylester patterns for studying the structure-functional property relationships in pectin.

Enzymatic modification of pectin to increase its calcium sensitivity while preserving its molecular weight.

A commercial high-methoxy citrus pectin was treated with a purified salt-independent pectin methylesterase (PME) isozyme isolated from Valencia orange peel to prepare a series of deesterified pectins. A series of alkali-deesterified pectins was also prepared at pH 10 under conditions permitting beta-elimination. Analysis of these pectins using high-performance size exclusion chromatography (HPSEC) with on-line multiangle laser light-scattering, differential viscometer, and refractive index (RI) detectors revealed no reduction in weight-average molecular weight (M(w); 150000) in the PME-treated pectin series, whereas a 16% reduction in intrinsic viscosity (IV) occurred below a degree of esterification (DE) of 47%. In contrast, alkali deesterification rapidly reduced both M(w) and IV to less than half of that observed for untreated pectin. PME treatment of a non-calcium-sensitive citrus pectin introduced calcium sensitivity with only a 6% reduction in the DE. Triad blocks of unesterified galacturonic acid were observed in (1)H nuclear magnetic resonance spectra of this calcium-sensitive pectin (CSP). These results demonstrate that the orange salt-independent PME isozyme utilizes a blockwise mode of action. This is the first report of the preparation of a CSP by PME treatment without significant loss of the pectin's M(w) due to depolymerization.

MECHANICAL PROPERTIES OF ORANGE PEEL AND FRUIT TREATED PRE–HARVEST WITH GIBBERELLIC ACID

Mechanical property data of citrus peel and fruit are limited. Gibberellic acid (GA3), a plant growth regulator,is applied to citrus fruit, in part, because it is believed to improve fruit integrity, yet studies characterizing effects ofGA3treatment on mechanical properties of citrus peel and fruit are lacking. The effect of GA3 treatment on the mechanicalproperties of Valencia orange (Citrus sinensis) peel and fruit was tested. Trees were treated with GA3 before, at, or after peelcolor break in the fall of 1998, or nontreated, and their fruit was harvested on 23 April (midseason) and 13 May(lateseason) 1999. Peel samples were subjected to shear and tensile tests, and whole fruits were subjected to compressionand cutting tests. The magnitude of peel and wholefruit mechanical properties decreased between harvests, but moretreatment effects were observed on the second harvest than on the first.<br><br>Treatments affected more mechanical properties of the peel than the whole fruit. Mechanical properties of the peel werepoorly correlated with thickness, probably because the mechanical properties of the tissues composing the peel are notequivalent. The earliest GA3 application date was the most effective. On the second harvest, the peel of fruit treated beforecolor break had 10% greater rupture force (in shear) and 15% greater tensile strength than the peel of nontreated fruit, whilethe properties of treated and nontreated whole fruits were similar. On the later harvest, the peel of fruit treated with GA3before color break had 10% greater rupture force than the peel of nontreated fruit, while the peel of fruit treated at colorbreak had 15% greater shear strength than nontreated fruit. The peel of treated fruit generally had tensile properties thatwere 10% to 20% greater than the peel of control fruit, especially on the second harvest. Fruit treated before color break alsohad greater apparent modulus of elasticity and higher cutting force than other fruit.

THERMOSTABLE AND THERIVIOLABILEISOFORMS IN COMMERCIAL ORANGE PEEL PECTINESTERASE

Thermostable and thermolabile pectinesterase isoforms in commercial Valencia orange peel pectinesterase (Sigma Chemical Co., St. Louis, MO) were separated by affinity chromatography on a Heparin-Sepharose CL-6B column. Among the seven putative PE isoforms separated, three (P1, P2 and P3 with 70 kDa, 60 kDa and 27 kDa, respectively) were thermostable and four [P4, P5, P6 (35 kDa) and P7 (35 kDa)J were thermolabile, according to the qualitative and quantitative studies of thermal stability. The commercial PE showed 83.4% and 98.3% inactivation after heat treatment at 70C for 5 min and at 90C for I min, respectively. P2 was the most heat stable, with 85. 5% inactivation at 70C for 5 min and 93. 0% inactivation at 90C for 1 min. The thermolabile P6 and P7 could be inactivated at 70C for 5 min or at 90C for 1 min. Thermostable PE activity was 16.6% of total activity of the commercial PE based on heat treatment at 70C for 5 min, but only 1.5% based on heating at 90C for I min. The 90C for 1 min heat treatment might be better than the 70C for 5 min heat treatment to distinguish thermostable PE and thermolabile PE isoforms in Valencia oranges.

Chlorophyll breakdown by chlorophyllase: isolation and functional expression of the Chlase1 gene from ethylene-treated Citrus fruit and its regulation during development.

We report on the isolation, functional expression and characterization of a cDNA encoding chlorophyllase, the enzyme catalyzing the first step in the chlorophyll breakdown pathway. The Chlase1 cDNA from Valencia Orange (Citrus sinensis cv. Valencia) was obtained by RT-PCR using degenerate primers based on the amino acid sequence of the previously purified protein. Chlase1 encodes a protein of 329 amino acids, including a sequence domain characterizing serine-lipases and a putative chloroplast-directing transit peptide. The Chlase1 gene encodes an active chlorophyllase enzyme which catalyzes the dephytylation of chlorophyll as shown by in vitro recombinant enzyme assays. Chlorophyllase expression at the transcript level in Valencia orange peel was found to be low and constitutive during natural fruit development without significant increase towards color-break and ripening. However, ethylene treatment induced an increase in chlorophyllase transcript at all stages of development. An enhanced response to ethylene treatment was observed during the months of October and November, corresponding to the time of natural color-break. The senescence-delaying regulator gibberellin-A3 (GA3) inhibited the effect of ethylene on chlorophyllase transcript accumulation. The data presented suggest that chlorophyllase may not be the regulator of chlorophyll breakdown during natural fruit ripening but is consistent with the notion that chlorophyll is gradually degraded during ripening due to a negative balance between synthesis and breakdown. According to this model, exogenous application of ethylene accelerates chlorophyll breakdown due to increased de novo synthesis of chlorophyllase. Further experimentation on the regulation and role of chlorophyllase in planta will be facilitated by the gene tools established in this work.

CELLULOSE‐HEMICELLULOSE FRACTIONS IN THE ALCOHOL‐INSOLUBLE SOLIDS OF VALENCIA ORANGE PEEL

ABSTRACTThe hemicellulose fraction of immature and mature Valencia orange peel was fractionally extracted and analyzed after removing the interfering polymeric materials consisting of proteins and large amounts of pectic substances. A procedure for the fractionation of the alcohol‐insoluble solids (AIS) is presented for citrus fruits, which may be used for other plant materials. The monosaccharide constituents of the hemi‐cellulose were determined after hydrolysis and paper chromatography. Comparisons of the pectic substances, protein, hemicellulose, and cellulose components were made between immature and mature Valencia orange peel.

β-Citraurin epoxide, a new carotenoid from valencia orange peel

A new minor carotenoid, β-citraurin epoxide (3-hydroxy-5,6-epoxy-5,6-dihydro-8′-apo-β-caroten-8′-al) and several isomers of violaxanthin (5,6,5′,6′-diepoxy-5,6,5′,6′-tetrahydro-β,β-caroten-3,3′-diol) have been identified in Valencia orange peel. The previously reported occurrence of apo-10′-violaxanthal (3-hydroxy- 5,6-epoxy-5,6-dihydro-10′-apo-β-caroten-10′-al) and apo-12′-violaxanthal (3-hydroxy-5,6-epoxy-5,6-dihydro- 12′-apo-β-caroten-12′-al) has been confirmed, and their syntheses are described. The quantitative determination of the carotenoids has also been performed.

Nootkatene from Citrus sinensis

The sesquiterpene hydrocarbon nootkatene and an unidentified sesquiterpene hydrocarbon, MW 204, have been isolated from Valencia orange peel.

Aldehydes, Ketones and Esters in Valencia Orange Peel Oil

SUMMARY: The aldehydes, ketones and esters in the volatile portion of Valencia orange oil have been isolated and identified. Seventeen aldehydes and ketones and three esters are included, 10 of which have never been identified as constituents of Valencia oil. Cis‐ and trans‐limonene oxide have also been found to occur in the same fraction. This is the first time that these compounds have been identified in cold‐pressed orange oil. A series of five homologous α,β‐unsaturated aldehydes were isolated and partially characterized.

Isolation and identification of a series of .alpha.,.beta.-unsaturated aldehydes from Valencia orange peel oil

Isolation and identification of a series of .alpha.,.beta.-unsaturated aldehydes from Valencia orange peel oil