Juices and By-Products of Red-Fleshed Sweet Oranges: Assessment of Bioactive and Nutritional Compounds.

The content of nutrients and bioactive compounds, and antioxidant capacity were assessed in the juices from two red-fleshed oranges, Cara Cara and Kirkwood, and compared with that of a standard Navel orange. Two juice extraction procedures, hand-squeezing and industrial, and two treatments, pasteurization (85 °C/30 s) and high-pressure homogenization (HPH, 150 MPa/55 °C/1 min), were evaluated. For most of the nutrients and bioactive compounds, the hand and industrial juice squeezing rendered similar extraction efficiency. Individual composition of carotenoids in the juices were differentially affected by the extraction procedure and the treatments, but the red-fleshed orange juices contained between 3- to 6-times higher total carotenoids than the standard Navel juices, being phytoene and phytofluene the main carotenoids. The industrial and treated juices of both red-fleshed oranges contained 20-30% higher amounts of tocopherols but about 20% lower levels of vitamin C than Navel juices. Navel juices exhibited higher hydrophilic antioxidant capacity, while the red-fleshed orange juices showed an improved lipophilic antioxidant capacity. The main distinctive characteristic of the industrial juice by-product of the red-fleshed oranges was a higher content of carotenoids (×10) and singlet oxygen antioxidant capacity (×1.5-2) than the Navel by-product.

A comprehensive analysis of carotenoids metabolism in two red-fleshed mutants of Navel and Valencia sweet oranges (Citrus sinensis).

Kirkwood Navel and Ruby Valencia are two spontaneous bud mutations of the respective parental lines of sweet orange (Citrus sinensis) Palmer Navel and Olinda Valencia, showing an atypical red pigmentation of the pulp. These red-fleshed varieties are commercially available and highly attractive for consumers but their carotenoid metabolism and the basis of the mutation have not been investigated. The red colour of Kirkwood and Ruby pulp was observed from the very early stages of fruit development until full maturity and associated with an altered carotenoid profiling. The red-fleshed varieties accumulated from 6- up to 1000-times more total carotenoids compared to the standard oranges. Specifically, the pulp of Kirkwood and Ruby accumulated large amounts of phytoene and phytofluene, and moderate contents of lycopene. Moreover, the red-fleshed oranges contained other unusual carotenes as δ-carotene, and lower concentrations of downstream products such as ß,ß-xanthophylls, abscisic acid (ABA) and ABA-glucosyl ester. This peculiar profile was associated with chromoplasts with lycopene crystalloid structures and round vesicles likely containing colourless carotenes. The flavedo and leaves of Kirkwood and Ruby showed minor changes in carotenoids, mainly limited to higher levels of phytoene. The carotenoid composition in Kirkwood and Ruby fruits was not explained by differences in the transcriptional profile of 26 genes related to carotenoid metabolism, covering the main steps of biosynthesis, catabolism and other processes related to carotenoid accumulation. Moreover, sequence analysis of the lycopene cyclase genes revealed no alterations in those of the red-fleshed oranges compared to the genes of the standard varieties. A striking event observed in Kirkwood and Ruby trees was the reddish coloration of the inner side of the bark tissue, with larger amounts of phytoene, accumulation of lycopene and lower ABA content. These observation lead to the conclusion that the mutation is not only manifested in fruit, affecting other carotenogenic tissues of the mutant plants, but with different consequences in the carotenoid profile. Overall, the carotenoid composition in the red-fleshed mutants suggests a partial blockage of the lycopene ß-cyclization in the carotenoid pathway, rendering a high accumulation of carotenes upstream lycopene and a reduced flow to downstream xanthophylls and ABA.

Tunable degrees of neurodegeneration in rats based on microsphere-induced models of chronic glaucoma.

This study compares four different animal models of chronic glaucoma against normal aging over 6 months. Chronic glaucoma was induced in 138 Long-Evans rats and compared against 43 aged-matched healthy rats. Twenty-five rats received episcleral vein sclerosis injections (EPIm cohort) while the rest were injected in the eye anterior chamber with a suspension of biodegradable microspheres: 25 rats received non-loaded microspheres (N-L Ms cohort), 45 rats received microspheres loaded with dexamethasone (MsDexa cohort), and 43 rats received microspheres co-loaded with dexamethasone and fibronectin (MsDexaFibro cohort). Intraocular pressure, neuroretinal function, structure and vitreous interface were evaluated. Each model caused different trends in intraocular pressure, produced specific retinal damage and vitreous signals. The steepest and strongest increase in intraocular pressure was seen in the EPIm cohort and microspheres models were more progressive. The EPIm cohort presented the highest vitreous intensity and percentage loss in the ganglion cell layer, the MsDexa cohort presented the greatest loss in the retinal nerve fiber layer, and the MsDexaFibro cohort presented the greatest loss in total retinal thickness. Function decreased differently among cohorts. Using biodegradable microspheres models it is possible to generate tuned neurodegeneration. These results support the multifactorial nature of glaucoma based on several noxa.

Bioactive Compounds, Nutritional Quality and Antioxidant Capacity of the Red-Fleshed Kirkwood Navel and Ruby Valencia Oranges.

Kirkwood Navel and Ruby Valencia are two spontaneous bud-mutations of the ordinary Washington Navel and Valencia late oranges characterized by the red coloration of their flesh. The purpose of this study was to analyze the physiological features, internal fruit quality, contents of relevant bioactive compounds and antioxidant capacity in the pulps of the red-fleshed fruits compared with the ordinary oranges during late development and maturation. In general, the content of sugars, organic acids, vitamin C, tocopherols, total phenolics and flavonoids, the hydrophilic antioxidant capacity and their changes during maturation were similar in the red-fleshed oranges and in the corresponding blond oranges. However, the mature Ruby fruits contained lower concentrations of sugars, malic and succinic acid and higher levels of citric acid than the ordinary Valencia. The major difference between the pulps of the Kirkwood and Ruby oranges and those of the ordinary oranges was the higher lipophilic antioxidant capacity and SOAC (singlet oxygen absorption capacity) of the former. Together, the high and unique content and composition of carotenoids in Kirkwood and Ruby may contribute to an enhanced antioxidant capacity without any detrimental effects on other fruit-quality attributes, making these varieties good sources of phytochemicals for the fresh-fruit and juice-processing citrus industries.

Ascorbic Acid Content and Transcriptional Profiling of Genes Involved in Its Metabolism during Development of Petals, Leaves, and Fruits of Orange (Citrus sinensis cv. Valencia Late).

Citrus fruit is one of the most important contributors to the ascorbic acid (AsA) intake in humans. Here, we report a comparative analysis of AsA content and transcriptional changes of genes related to its metabolism during development of petals, leaves and fruits of Valencia Late oranges (Citrus sinensis). Petals of close flowers and at anthesis contained the highest concentration of AsA. In fruits, AsA content in the flavedo reached a maximum at color break, whereas the pulp accumulated lower levels and experienced minor fluctuations during development. AsA levels in leaves were similar to those in the flavedo at breaker stage. The transcriptional profiling of AsA biosynthetic, degradation, and recycling genes revealed a complex and specific interplay of the different pathways for each tissue. The D-galacturonic acid pathway appeared to be relevant in petals, whereas in leaves the L-galactose pathway (GGP and GME) also contributed to AsA accumulation. In the flavedo, AsA content was positively correlated with the expression of GGP of the L-galactose pathway and negatively with DHAR1 gene of the recycling pathway. In the pulp, AsA appeared to be mainly controlled by the coordination among the D-galacturonic acid pathway and the MIOX and GalDH genes. Analysis of the promoters of AsA metabolism genes revealed a number of cis-acting elements related to developmental signals, but their functionalities remain to be investigated.

Analysis of Parainflammation in Chronic Glaucoma Using Vitreous-OCT Imaging.

Glaucoma causes blindness due to the progressive death of retinal ganglion cells. The immune response chronically and subclinically mediates a homeostatic role. In current clinical practice, it is impossible to analyse neuroinflammation non-invasively. However, analysis of vitreous images using optical coherence tomography detects the immune response as hyperreflective opacities. This study monitors vitreous parainflammation in two animal models of glaucoma, comparing both healthy controls and sexes over six months. Computational analysis characterizes in vivo the hyperreflective opacities, identified histologically as hyalocyte-like Iba-1+ (microglial marker) cells. Glaucomatous eyes showed greater intensity and number of vitreous opacities as well as dynamic fluctuations in the percentage of activated cells (50-250 microns2) vs. non-activated cells (10-50 microns2), isolated cells (10 microns2) and complexes (>250 microns2). Smaller opacities (isolated cells) showed the highest mean intensity (intracellular machinery), were the most rounded at earlier stages (recruitment) and showed the greatest change in orientation (motility). Study of vitreous parainflammation could be a biomarker of glaucoma onset and progression.

Regulation of Tocopherol Biosynthesis During Fruit Maturation of Different Citrus Species.

Tocopherols are plant-derived isoprenoids with vitamin E activity, which are involved in diverse physiological processes in plants. Although their biosynthesis has been extensively investigated in model plants, their synthesis in important fruit crops as Citrus has scarcely been studied. Therefore, the aim of this work was to initiate a physiological and molecular characterization of tocopherol synthesis and accumulation in Citrus fruits during maturation. For that purpose, we selected fruit of the four main commercial species: grapefruit (Citrus paradisi), lemon (Citrus limon), sweet orange (Citrus sinensis), and mandarin (Citrus clementina), and analyzed tocopherol content and the expression profile of 14 genes involved in tocopherol synthesis during fruit maturation in both the flavedo and pulp. The selected genes covered the pathways supplying the tocopherol precursors homogentisate (HGA) (TAT1 and HPPD) and phytyl pyrophosphate (PPP) (VTE5, VTE6, DXS1 and 2, GGPPS1 and 6, and GGDR) and the tocopherol-core pathway (VTE2, VTE3a, VTE3b, VTE1, and VTE4). Tocopherols accumulated mainly as α- and γ-tocopherol, and α-tocopherol was the predominant form in both tissues. Moreover, differences were detected between tissues, among maturation stages and genotypes. Contents were higher in the flavedo than in the pulp during maturation, and while they increased in the flavedo they decreased or were maintained in the pulp. Among genotypes, mature fruit of lemon accumulated the highest tocopherol content in both the flavedo and the pulp, whereas mandarin fruit accumulated the lowest concentrations, and grapefruit and orange had intermediate levels. Higher concentrations in the flavedo were associated with a higher expression of all the genes evaluated, and different genes are suitable candidates to explain the temporal changes in each tissue: (1) in the flavedo, the increase in tocopherols was concomitant with the up-regulation of TAT1 and VTE4, involved in the supply of HGA and the shift of γ- into α-tocopherol, respectively; and (2) in the pulp, changes paralleled the expression of VTE6, DXS2, and GGDR, which regulate PPP availability. Also, certain genes (i.e., VTE6, DXS2, and GGDR) were co-regulated and shared a similar pattern during maturation in both tissues, suggesting they are developmentally modulated.

Chronic Glaucoma Using Biodegradable Microspheres to Induce Intraocular Pressure Elevation. Six-Month Follow-Up.

BACKGROUND: To compare two prolonged animal models of glaucoma over 24 weeks of follow-up. A novel pre-trabecular model of chronic glaucoma was achieved by injection of biodegradable poly lactic-co-glycolic acid (PLGA) microspheres (10-20 µm) (Ms20/10) into the ocular anterior chamber to progressively increase ocular hypertension (OHT). METHODS: Rat right eyes were injected to induce OHT: 50% received a suspension of Ms20/10 in the anterior chamber at 0, 2, 4, 8, 12, 16 and 20 weeks, and the other 50% received a sclerosing episcleral vein injection biweekly (EPIm). Ophthalmological clinical signs, intraocular pressure (IOP), neuroretinal functionality measured by electroretinography (ERG), and structural analysis of the retina, retinal nerve fiber layer (RNFL) and ganglion cell layer (GCL) protocols using optical coherence tomography (OCT) and histological exams were performed. RESULTS: Both models showed progressive neuroretinal degeneration (p < 0.05), and contralateral eye affectation. The Ms20/10 model showed a more progressive increase in IOP and better preservation of ocular surface. Although no statistical differences were found between models, the EPIm showed a tendency to produce thicker retinal and thinner GCL thicknesses, slower latency and smaller amplitude as measured using ERG, and more aggressive disturbances in retinal histology. In both models, while the GCL showed the greatest percentage loss of thickness, the RNFL showed the greatest and earliest rate of thickness loss. CONCLUSIONS: The intracameral model with biodegradable microspheres resulted more like the conditions observed in humans. It was obtained by a less-aggressive mechanism, which allows for adequate study of the pathology over longer periods.

Particle film technology modulates xanthophyll cycle and photochemical dynamics of grapevines grown in the Douro Valley.

Field-grown grapevines are often exposed to multiple environmental stresses, which challenges wine-growers to develop sustainable measures to sustain vine growth, yield, and quality. Under field conditions this task is demanding, due to differences in the magnitudes of stresses and associated plant responses. In this study we explored the hypothesis that kaolin-particle film application improves grapevine photoprotection through the regulation of xanthophyll cycle genes, limiting the thermal dissipation of excess energy under harsh environmental conditions. Hence, we selected two grapevine varieties, Touriga-Nacional (TN) and Touriga-Franca (TF), grown in the Douro Demarcated Region, and evaluated changes in light dissipation mechanisms, xanthophyll cycle components, and the expression of xanthophyll cycle genes during the 2017 summer season. The results showed that, from veraison to ripening, kaolin triggered the up-regulation of violaxanthin de-epoxidase (VvVDE1) and zeaxanthin epoxidase (VvZEP1) genes, indicating optimised regulation of the xanthophyll cycle. Kaolin treatment also decreased chlorophyll (Chla, Chlb, Chl(a+b)) and carotenoid (Car) accumulation under increasing summer stress conditions in both varieties and lowered the non-photochemical quenching (NPQ) of grapevines on ripening, suggesting a long-term response to summer stress. In addition, kaolin-treated grapevines showed increased Chla/Chlb and lower Chl(a+b)/Car ratios, displaying some features of high light adapted leaves. Overall, this study suggests that kaolin application enabled grapevines to benefit from fluctuating periods of summer stress by managing chlorophyll and carotenoid content and limiting down-regulation of both photochemistry and photoinhibition processes. Under Mediterranean field conditions, kaolin application can be considered an efficient method of minimising summer stress impact on grapevines.

Novel Use of PLGA Microspheres to Create an Animal Model of Glaucoma with Progressive Neuroretinal Degeneration.

Progressive degeneration of neuroretinal tissue with maintained elevated intraocular pressure (IOP) to simulate chronic glaucoma was produced by intracameral injections of poly (lactic-co-glycolic) acid (PLGA) microspheres (Ms) in rat eyes. The right eye of 39 rats received different sizes of PLGA-Ms (2 µL suspension; 10% w/v): 14 with 38-20 µm Ms (Ms38/20 model) and 25 with 20-10 µm particles (Ms20/10 model). This novel glaucoma animal model was compared to the episcleral vein sclerosis (EPI) model (25 eyes). Injections were performed at baseline, two, four and six weeks. Clinical signs, IOP, retina and optic nerve thicknesses (using in vivo optical coherence tomography; OCT), and histological studies were performed. An IOP increment was observed in all three groups, however, the values obtained from the PLGA-Ms injection resulted lower with a better preservation of the ocular surface. In fact, the injection of Ms20/10 created a gentler, more progressive, and more sustained increase in IOP. This IOP alteration was correlated with a significant decrease in most OCT parameters and in histological ganglion-cell count for the three conditions throughout the eight-week follow-up. In all cases, progressive degeneration of the retina, retinal ganglion cells and optic nerve, simulating chronic glaucoma, was detected by OCT and corroborated by histological study. Results showed an alternative glaucoma model to the well-known episcleral vein model, which was simpler to perform, more reproducible and easier to monitor in vivo.

Monitoring New Long-Lasting Intravitreal Formulation for Glaucoma with Vitreous Images Using Optical Coherence Tomography.

Intravitreal injection is the gold standard therapeutic option for posterior segment pathologies, and long-lasting release is necessary to avoid reinjections. There is no effective intravitreal treatment for glaucoma or other optic neuropathies in daily practice, nor is there a non-invasive method to monitor drug levels in the vitreous. Here we show that a glaucoma treatment combining a hypotensive and neuroprotective intravitreal formulation (IF) of brimonidine-Laponite (BRI/LAP) can be monitored non-invasively using vitreoretinal interface imaging captured with optical coherence tomography (OCT) over 24 weeks of follow-up. Qualitative and quantitative characterisation was achieved by analysing the changes in vitreous (VIT) signal intensity, expressed as a ratio of retinal pigment epithelium (RPE) intensity. Vitreous hyperreflective aggregates mixed in the vitreous and tended to settle on the retinal surface. Relative intensity and aggregate size progressively decreased over 24 weeks in treated rat eyes as the BRI/LAP IF degraded. VIT/RPE relative intensity and total aggregate area correlated with brimonidine levels measured in the eye. The OCT-derived VIT/RPE relative intensity may be a useful and objective marker for non-invasive monitoring of BRI/LAP IF.

Effect of fruit shading and cold storage on tocopherol biosynthesis and its involvement in the susceptibility of Star Ruby grapefruit to chilling injury.

The aim of this study was to investigate the role of tocopherols in the susceptibility of Star Ruby grapefruit to postharvest chilling injury (CI). Fruit exposed to normal sunlight (NC, non-covered) and deprived of light (C, covered) in the last stages of development were used. Tocopherol contents increased in the flavedo of both NC and C fruit during development, concomitantly with the up-regulation of TAT1 and most genes of the tocopherol-core pathway. Fruit shading reduced total contents by repressing γ-tocopherol accumulation, associated to a down-regulation of GGDR and VTE1 and, to a lesser extent, of VTE2, VTE3a and VTE4. During cold storage, total and α-tocopherol contents increased in NC and C fruit, and no direct relationship between tocopherol accumulation and CI tolerance was found. Cold stress up-regulated most genes involved in the synthesis of tocopherol precursors and down-regulated those of the tocopherol-core pathway, but changes seemed to be cold-mediated and not related to CI development.

Effect of age and sex on neurodevelopment and neurodegeneration in the healthy eye: Longitudinal functional and structural study in the Long-Evans rat.

The processes involved in neurodevelopment and aging have not yet been fully discovered. This is especially challenging in premorbid or borderline situations of neurodegenerative diseases such as Alzheimer's or glaucoma. The retina, as part of the central nervous system, can be considered the easiest and most accessible neural structure that can be analyzed using non-invasive methods. Animal studies of neuroretinal tissue in situations of health and under controlled conditions allow the earliest sex- and aging-induced changes to be analyzed so as to differentiate them from the first signs occurring in manifested disease. This study evaluates differences by age and sex based on intraocular pressure (IOP) and neuroretinal function and structure in healthy young and adult rats before decline due to senescence. For this purpose, eighty-five healthy Long-Evans rats (31 males and 54 females) were analyzed in this 6-month longitudinal study running from childhood to adulthood. IOP was measured by tonometer (Tonolab; Tiolat Oy Helsinki, Finland), neuroretinal function was recorded by flash scotopic and light-adapted photopic negative response electroretinography (ERG) (Roland consult® RETIanimal ERG, Germany) at 4, 16 and 28 weeks of age; and structure was evaluated by in vivo optical coherence tomography (OCT) (Spectralis, Heidelberg® Engineering, Germany). Analyzing both sexes together, IOP was below 20 mmHg throughout the study; retina (R), retinal nerve fiber layer (RNFL) and ganglion cell layer (GCL) thicknesses measured by OCT decreased over time; an increase in ERG signal was recorded at week 16; and no differences were found between right and left eyes. However, analyzing differences by sex revealed that males had higher IOP (even reaching ocular hypertension [>20 mmHg] by the end of the study [7 months of age]), exhibited greater neuroretinal thickness but higher structural percentage loss, and had worse dark- and light-adapted function as measured by ERG than females. This study concludes that age and sex influenced neurodevelopment and neurodegeneration. Different structural and functional degenerative patterns were observed by sex; these occurred earlier and more intensely in males than in age-matched females.

Dexamethasone delivery to the ocular posterior segment by sustained-release Laponite formulation.

This paper presents a novel nanoformulation for sustained-release delivery of dexamethasone (DEX) to the ocular posterior segment using a Laponite (LAP) carrier-DEX/LAP 1:10 w w-1 formulation; 10 mg ml-1. In vivo ocular feasibility and pharmacokinetics after intravitreal (IV) and suprachoroidal (SC) administration in rabbit eyes are compared against IV administration of a DEX solution (1 mg ml-1). Thirty rabbit eyes were injected with the DEX/LAP formulation (15 suprachoroid/15 intravitreous). Ophthalmological signs were monitored at day 1 and at weeks 1-4-12-24 post-administration. Three eyes per sample time point were used to quantify DEX concentration using high-performance liquid chromatography-mass spectrometry. The ocular tissues' pharmacokinetic parameters (lens, vitreous humour, choroid-retina unit and sclera) were studied. DEX/LAP was well tolerated under both administration methods. Peak intraocular DEX levels from the DEX/LAP were detected in the vitreous humour after both deliveries soon after administration. The vitreous area under the curve was significantly greater after both DEX/LAP deliveries (IV: 205 968.47; SC: 11 442.22 ng g-1 d-1) than after IV administration of the DEX solution (317.17 ng g-1 d-1). Intravitreal DEX/LAP delivery extended higher vitreous DEX levels up to week 24 (466.32 ± 311.15 ng g-1). With SC delivery, DEX levels were detectable in the choroid-retina unit (12.04 ± 20.85 ng g-1) and sclera (25.46 ± 44.09 ng g-1) up to week 24. This study demonstrated the intraocular feasibility of both SC and IV administration of the DEX/LAP formulation. The LAP increased the intraocular retention time of DEX when compared with conventional solutions. DEX/LAP could be considered a biocompatible and useful sustained-release formulation for treating posterior-pole eye diseases.

Visualization of Carotenoid-Storage Structures in Fruits by Transmission Electron Microscopy.

Plastids are cell organelles that, beside other functions, have the capability to store carotenoids in specialized structures, which may vary among the different plant species, tissues or according to the carotenoid complement. Fruits are an important source of carotenoids, and during ripening, chloroplasts differentiate into chromoplasts that are able to accumulate large amounts of carotenoids, rendering then the characteristic fruit coloration. Whereas lycopene or ß-carotene may accumulate as crystal in the chromoplasts of some fruit, other xanthophyll-accumulating fruits differentiate plastoglobuli as a preferred system to enhance carotenoids stability and storage. Visualization of plastid ultrastructure and their transformation during ripening or in fruit of contrasting coloration are fundamental objectives within carotenoids research in fruits. Therefore, in this chapter, we describe a protocol for the visualization and analysis of plastid ultrastructure by transmission electron microscopy (TEM), specially designed and adapted to fruit tissues.

Evaluation of Carotenoids Protection Against Oxidative Stress in the Animal Model Caenorhabditis elegans.

The nematode Caenorhabditis elegans is a versatile and powerful model organism for animal experimental research and, despite being an invertebrate, displays remarkably similar molecular bases and conserved cellular pathways to those of humans. Oxidative stress is an etiological factor that influences numerous diseases, degenerative processes and aging. C. elegans has revealed as an opportune and feasible organism to investigate the antioxidant effects of different bioactives or complex food matrices, and a number of protocols have been developed by using different oxidative stressors. Carotenoids are recognized as quenchers and scavengers of reactive oxygen species, and many of their related health benefits attributed in the diet are tightly linked to their antioxidant properties. In this chapter, we report a simple and rapid assay to evaluate the protection capacity of pure carotenoids or complex carotenoid extracts against oxidative stress in the model system C. elegans. The protocol describes a representative feeding experiment by adding carotenoids to the nematode growth medium and after an incubation period, the C. elegans populations fed with carotenoids are exposed to an acute oxidative stress by using H2O2 as oxidative agent. The protection against oxidative stress is evaluated as the survival rate of the nematodes fed with the carotenoid prior to receiving oxidative treatment compared with the survival rate of control nematode population. In order to confirm the carotenoid intake by the nematodes during the feeding experiment a bioassimilation experiment is also reported.

Empirical Mode Decomposition-Based Filter Applied to Multifocal Electroretinograms in Multiple Sclerosis Diagnosis.

As multiple sclerosis (MS) usually affects the visual pathway, visual electrophysiological tests can be used to diagnose it. The objective of this paper is to research methods for processing multifocal electroretinogram (mfERG) recordings to improve the capacity to diagnose MS. MfERG recordings from 15 early-stage MS patients without a history of optic neuritis and from 6 control subjects were examined. A normative database was built from the control subject signals. The mfERG recordings were filtered using empirical mode decomposition (EMD). The correlation with the signals in a normative database was used as the classification feature. Using EMD-based filtering and performance correlation, the mean area under the curve (AUC) value was 0.90. The greatest discriminant capacity was obtained in ring 4 and in the inferior nasal quadrant (AUC values of 0.96 and 0.94, respectively). Our results suggest that the combination of filtering mfERG recordings using EMD and calculating the correlation with a normative database would make mfERG waveform analysis applicable to assessment of multiple sclerosis in early-stage patients.

Computer-Aided Diagnosis of Multiple Sclerosis Using a Support Vector Machine and Optical Coherence Tomography Features.

The purpose of this paper is to evaluate the feasibility of diagnosing multiple sclerosis (MS) using optical coherence tomography (OCT) data and a support vector machine (SVM) as an automatic classifier. Forty-eight MS patients without symptoms of optic neuritis and forty-eight healthy control subjects were selected. Swept-source optical coherence tomography (SS-OCT) was performed using a DRI (deep-range imaging) Triton OCT device (Topcon Corp., Tokyo, Japan). Mean values (right and left eye) for macular thickness (retinal and choroidal layers) and peripapillary area (retinal nerve fibre layer, retinal, ganglion cell layer-GCL, and choroidal layers) were compared between both groups. Based on the analysis of the area under the receiver operator characteristic curve (AUC), the 3 variables with the greatest discriminant capacity were selected to form the feature vector. A SVM was used as an automatic classifier, obtaining the confusion matrix using leave-one-out cross-validation. Classification performance was assessed with Matthew's correlation coefficient (MCC) and the AUCCLASSIFIER. The most discriminant variables were found to be the total GCL++ thickness (between inner limiting membrane to inner nuclear layer boundaries), evaluated in the peripapillary area and macular retina thickness in the nasal quadrant of the outer and inner rings. Using the SVM classifier, we obtained the following values: MCC = 0.81, sensitivity = 0.89, specificity = 0.92, accuracy = 0.91, and AUCCLASSIFIER = 0.97. Our findings suggest that it is possible to classify control subjects and MS patients without previous optic neuritis by applying machine-learning techniques to study the structural neurodegeneration in the retina.

Genuine Carotenoid Profiles in Sweet Orange [Citrus sinensis (L.) Osbeck cv. Navel] Peel and Pulp at Different Maturity Stages.

The carotenogenesis in the endocarp and flavedo of Navel oranges over four consecutive maturity stages was assessed by high-performance liquid chromatography-diode array detection-atmospheric pressure chemical ionization-multistage mass spectrometry. After optimization of the extraction method, 77 carotenoids, including 26 monoesters and 33 diesters of violaxanthin, ß-citraurin, and antheraxanthin, were characterized. Whereas chloroplast-specific pigments, such as (all-E)-lutein and (all-E)-ß-carotene, predominated in the flavedo of green-ripe fruit, a highly complex pattern of xanthophyll esters was found in the mature oranges. Total carotenoid contents of flavedo were approximately 9-fold higher [12 605 µg/100 g of fresh weight (FW)] than those in the endocarp (1354 µg/100 g of FW) at the fully mature stage. The mature endocarp abundantly contained violaxanthin mono- and diesters, in addition to diverse antheraxanthin esters, which were exclusively detected in this fruit fraction. Likewise, ß-citraurin esters were found to be unique flavedo constituents of mature fruit. Therefore, they may support the detection of fraudulent use of peel fractions during orange juice production.