Fruit Pigmentation Research Articles

Elevated Gene Expression inChalcone SynthaseEnzyme Suggests an Increased Production of Flavonoids in Skin and Synchronized Red Cell Cultures of North American Native Grape Berries

Anthocyanins are antioxidants and are among the natural products synthesized via the flavonoid biosynthesis pathway. Anthocyanins have been recommended for dietary intake in the prevention of cardiovascular diseases, cancer, and age-related conditions such as Alzheimer's disease or dementia. With an increasingly aging population in many parts of the world, strategies for the commercial production of in vitro synchronized red cell cultures as natural antioxidants will be a significant contribution to human medicine. Red pigmented fruits such as grapes (Vitis sp.) are a major source of bioavailable anthocyanins and other polyphenols. Since the level of antioxidants varies among cultivars, this study is the first one that phytochemically and genetically characterizes native grape cultivars of North America to determine the optimal cultivar and berry cells for the production of anthocyanins as antioxidants. Using real-time PCR and bioinformatics approaches, we tested for the transcript expression of the chalcone synthase (CHS) gene, an enzyme involved in the flavonoid and anthocyanin biosynthesis pathway, in different parts of physiologically mature grape berries and in vitro synchronized red cells. A low level of expression was recorded in berry flesh, compared with an elevated expression in berry skins and in vitro synchronized red cells, suggesting increased production of flavonoids in skin and cell cultures. This preliminary study demonstrates the potential of functional genomics in natural products research as well as in systematic studies of North American native grapes, specifically in muscadine (Vitis rotundifolia).

Expression of Anthocyanin Biosynthetic Genes during Fruit Development in ‘Fengxiang’ Strawberry

Anthocyanins are the main pigments in flowers and fruits. In most cases, anthocyanin accumulation in fruit is highly controlled by the developmental level. In this study, the cDNA fragments of three genes, chalcone synthase (CHS), dihydroflavonol 4-reductase (DFR), and anthocyanidin synthase (ANS), which are involved in the flavonoid pathway, were isolated from total RNA of strawberry ripe fruit by using polymerase chain reaction technique and labeled as probes to determine the expression of anthocyanin biosynthetic genes. Northern analysis showed that a correlation between anthocyanin accumulation and expression of the flavonoid pathway genes during the ripening of strawberry fruits. At the early stages of fruit development, the mRNA levels encoding CHS, DFR, ANS were high probably responsible for the accumulation of condensed tannins, but the levels decreased dramatically when fruits turned white from green. During the stage of pigment accumulation, their mRNA levels increased strongly to be involved anthocyanin biosynthesis. Difference of CHS in mRNA abundance was correlated with differential accumulation of anthocyanins throughout the process of fruit development. Therefore, CHS could be a key structure gene involved in anthocyanin synthesis. Furthermore, the co-ordination of expression of anthocyanin biosynthetic genes implied a common regulatory mechanism controlling the expression of structural genes in the flavonoid pathway.

Salinity-induced changes in key anti-oxidant enzyme activities and in the levels of some anti-oxidants, osmo-protectants, inorganic ions, and chlorophyll pigments in okra fruit (Abelmoschus esculentus L.)

SummaryTo study the influence of NaCl stress on various biochemical components in okra (Abelmoschus esculentus L.) fruit, two differential cultivars, ‘Nirali’ and ‘Posa Sawni’, were grown under non-saline and saline conditions until fruit development. Salt treatment (50 mM or 100 mM NaCl) significantly decreased fresh fruit yields (by 47.5 – 67.7%), soluble protein contents (by 62.3 – 138.4%), K+ ion levels (by 17.7 – 29.0%), and peroxidase (POD) activity (by 17.7 – 44.6%) compared to fruit from non-saline treated okra plants. NaCl increased Na+ ion levels (by 47.9 – 118.8%), glycine betaine contents (GB; by 34.3 – 115.2%), H2O2 contents (by 4.14 – 64.4%), and ascorbic acid levels (by 73.0 –109.0%) and superoxide dismutase (SOD) activity in the fruit of both okra cultivars. ‘Nirali’ had higher fresh fruit yields, K+ ion, GB, H2O2, and soluble protein contents, and higher SOD and POD activities, while ‘Posa Sawni’ fruit had higher ascorbic acid and total phenolic compounds contents and catalase (CAT) activity. Overall, the higher fresh yield of ‘Nirali’ could be attributed to enhanced levels of K+ ions, GB, soluble proteins, and SOD and POD activities. Fruit collected from all salt-stressed okra plants (‘Nirali’ and ‘Posa Sawni’) contained higher amounts of some key anti-oxidants and osmo-protectants which are considered beneficial for human health.

Expression of Anthocyanin Biosynthetic Genes during Fruit Development in ‘Fengxiang’ Strawberry

Anthocyanins are the main pigments in flowers and fruits. In most cases, anthocyanin accumulation in fruit is highly controlled by the developmental level. In this study, the cDNA fragments of three genes, chalcone synthase (CHS), dihydroflavonol 4-reductase (DFR), and anthocyanidin synthase (ANS), which are involved in the flavonoid pathway, were isolated from total RNA of strawberry ripe fruit by using polymerase chain reaction technique and labeled as probes to determine the expression of anthocyanin biosynthetic genes. Northern analysis showed that a correlation between anthocyanin accumulation and expression of the flavonoid pathway genes during the ripening of strawberry fruits. At the early stages of fruit development, the mRNA levels encoding CHS, DFR, ANS were high probably responsible for the accumulation of condensed tannins, but the levels decreased dramatically when fruits turned white from green. During the stage of pigment accumulation, their mRNA levels increased strongly to be involved anthocyanin biosynthesis. Difference of CHS in mRNA abundance was correlated with differential accumulation of anthocyanins throughout the process of fruit development. Therefore, CHS could be a key structure gene involved in anthocyanin synthesis. Furthermore, the co-ordination of expression of anthocyanin biosynthetic genes implied a common regulatory mechanism controlling the expression of structural genes in the flavonoid pathway.

White‐fruited Duchesnea indica (Rosaceae) is impaired in ANS gene expression

Duchesnea indica is a wild strawberry-like species that has red fruits. In a recent survey in the highlands of Tucumán (Argentina), a plant of D. indica with white fruits was discovered. The aim of this study was to investigate whether the white-fruited character was due to a phenotypic or genotypic change. The stability and heritability of the character and the expression of genes involved in anthocyanins synthesis were studied and compared with red-fruited genotypes. This study contributes to understanding the molecular basis of some factors involved in fruit pigmentation, a horticulturally and taxonomically important trait. Stability and heritability of the white-fruited character were evaluated in plants obtained by asexual propagation or by sexual crosses between the white- and red-fruited genotypes. Asexual multiplications were carried out by stolon rooting and sexual multiplications by germination of achenes obtained from crosses. The expression level of the genes involved in the synthesis and regulation of the anthocyanins pathway (CHS, F3H, DFR, ANS, and MYB10) were evaluated by RT-PCR using specific primers. Plants with the white-fruited character always yielded white-fruited progeny when propagated asexually, whereas in sexually propagated plants fruit color depended on the mother. Red-fruited mothers yielded red-fruited progeny, and white-fruited mothers yielded fruits ranging from dark pink to white. Molecular analysis suggested that the white-fruited character was due to the low expression of the ANS gene. Results obtained indicate that the white-fruited character was stable. Mother progenitors exert a strong influence on the expression of the white-fruited character. The white-fruited phenotype is due to the impairment or downregulation of the ANS gene.

Genetic diversity of red-fleshed apples (Malus)

Anthocyanins are flavonoid pigments imparting red, blue, or purple pigmentation to fruits, flowers and foliage. These compounds are powerful antioxidants in vitro, and are widely believed to contribute to human health. The fruit of the domestic apple (Malus x domestica) is a popular and important source of nutrients, and is considered one of the top ‘functional foods’—those foods that have inherent health-promoting benefits beyond basic nutritional value. The pigmentation of typical red apple fruits results from accumulation of anthocyanin in the skin. However, numerous genotypes of Malus are known that synthesize anthocyanin in additional fruit tissues including the core and cortex (flesh). Red-fleshed apple genotypes are an attractive starting point for development of novel varieties for consumption and nutraceutical use through traditional breeding and biotechnology. However, cultivar development is limited by lack of characterization of the diversity of genetic backgrounds showing this trait. We identified and cataloged red-fleshed apple genotypes from four Malus diversity collections representing over 3,000 accessions including domestic cultivars, wild species, and named hybrids. We found a striking range of flesh color intensity and pattern among accessions, including those carrying the MYB10R6 allele conferring ectopic expression of a key transcriptional regulator of anthocyanin biosynthesis. Although MYB10R6 was strongly associated with red-fleshed fruit among genotypes, this allele was neither sufficient nor required for this trait in all genotypes. Nearly all red-fleshed accessions tested could be traced back to ‘Niedzwetzkyana’, a presumed natural form of M. sieversii native to central Asia.

Identification and Cancer Therapeutic Properties of Microfloral Anthocyanin Metabolites

Colon cancer is very common, but those who consume anthocyanins, common red pigments in wine, grapes and other fruits, appear to have lower rates of colon cancer. We investigated the fate of grape anthocyanins in the colon by using a pig model of the lower intestine and observed the rapid conversion to three simple phenols. These were studied, along with metabolites reported by others, for their effect on cell proliferation and apoptosis, and we observed promising results; cell proliferation was suppressed and apoptosis enhanced. Thus, these metabolites are promising anti-cancer agents and the effect of anthocyanins on colon cancer appears to be mediated by these bacterial metabolites.

Influence of ellagic acid on prostate cancer cell proliferation: A caspase–dependent pathway

Objective To evaluate the effect of allagic acid treatment on the cell viability of human prostate cancer cells. Methods Ellagic acid (10-100 mol/L) treatment (48 h) of human prostate carcinoma PC3 cells was found to result in a dose-dependent inhibition of cell growth and apoptosis of PC3 cells as assessed by MTT assay, western blotting, flow cytometry and confocal microscopy. Results We observed that ellagic acid treatment of PC3 cells resulted in a dose dependent inhibition of cell growth/cell viability. This ellagic acid caused cell growth inhibition was found to be accompanied by induction of apoptosis, as assessed by the cleavage of poly (ADP-ribose) polymerase (PARP) and morphological changes. Further, induction of apoptosis accompanied a decrease in the levels of antiapoptotic protein Bcl-2 and increase in proapoptotic protein Bax, thus shifting the Bax: Bcl-2 ratio in favor of apoptosis. Ellagic acid treatment of PC3 cells was also found to result in significant activation of caspases, as shown by the dose dependent decrease in the protein expression of procaspase−3, −6, −8 and −9. This ellagic acid-mediated induction of apoptosis was significantly (80%-90%) inhibited by the caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp (OMe)-fluoromethylketone (Z-VAD-FMK). Thus these data suggested an essential role of caspases in ellagic acid-mediated apoptosis of PC3 cells. Conclusions It is tempting to suggest that consumption of tropical pigmented fruits and vegetables could be an effective strategy to combat prostate cancer.

Pigments of guava paluma cultivar stored under environmental conditions

This study evaluated the levels of pigments in guavas such as chlorophyll, total carotenoids, total anthocyanins and yellow flavonoids. In the study, guava fruits (Paluma cultivar) were used, and they were stored under environmental conditions for nine days. The evaluation of guava fruit pigment levels was carried out every two days. Findings showed significant differences regarding levels of chlorophyll and carotenoid (p<0.05) during the storage period. Anthocyanin contents varied from 0.24 ± 0.01 to 0.37 ± 0.02 mg 100 g-1 guava pulp, showing no significant difference until the fifth day of storage. Values of yellow flavonoids were different from the seventh day of storage on. Levels of pigments evaluated in guava fruits (P. cultivar) changed during the storage period under environmental conditions, thus showing the influence of the continuous metabolic activity in the fruits during the post-harvest period. Key words: Psidium guajava L., chlorophyll, carotenoids, anthocyanins, flavonoides, storage.

Why fruits go to the dark side

The colours of fleshy fruits are usually attributed to attract seed dispersers to the plant. A cursory look at the gaudy colours of fleshy fruits on offer in a local fruit stall gives the impression that plants use primarily bright colours to attract fruit consumer. This impression is misleading; many small fruits ‘go to the dark side’ and become dark purple or black when ripe. Intermingled in foliage, these colours, which are produced by anthocyanins, can be fairly inconspicuous and are thus not easily reconciled with a signalling function to attract seed dispersers. In this review I therefore discuss complementary hypotheses on the function and evolution of fruit colouration. First, I focus on the evidence that fruit colours indeed function as signals to attract seed dispersers. I then show that anthocyanins, the most prevalent fruit pigments, are important dietary antioxidants that can be selected by blackcaps ( Sylvia atricapilla) which are important avian seed dispersers of many European plants. Moreover, the consumption of anthocyanins increases the likelihood that blackcaps mount an immune response during immune challenges. As a next step, I review evidence that anthocyanins accumulate in fruit skin in response to abiotic factors, in particular high illumination coupled with low temperature favour the increase of anthocyanins. Finally, I show that anthocyanins can also be selected for by fruit antagonists, consumers that do not disperse seeds. In particular, high contents of anthocyanins strongly reduce fungal growth in fruit tissue. Taken together, there are various selective pressures which likely influence fruit colour evolution. Currently, the relative importance of each of these selective agents is unknown. There is consequently a need to develop a more encompassing framework on fruit colour evolution.

Abstract 5422: Delphinidin dietary anthocyanidin modulates EGFR/HER-2 signaling pathway in breast cancer cells

Abstract Breast cancer is the second leading cause of cancer deaths in women in the United States. The epidermal growth factor receptor (EGFR) and HER-2 are expressed in many breast cancer patients and are associated with poor prognosis. Over-expression of EGFR and HER-2 lead to resistance against tamoxifen, a selective estrogen receptor (ER) modulator. In addition, the efficacy of trastuzumab or lapatinib for the treatment of ER-negative breast cancer as a monotherapy or in combination with chemotherapy is limited in more advanced stages of HER-2 positive breast cancers by development of therapeutic resistance. Considerable attention has been devoted towards the development of dual inhibitors of EGFR and HER-2 for breast cancer treatment. Several of these inhibitors such as lapatinib have been tested preclinically and clinically, but their use is limited because of their unacceptable cytotoxic effects on normal cells or cancer cells acquire resistance to such treatment. Therefore, identification of a natural, nontoxic agent(s) capable of inhibiting both EGFR and HER-2 is of utmost importance. Delphinidin, an anthocyanidin, present in many pigmented fruits and vegetables possesses anti-oxidant and anti-proliferative properties. We showed that delphinidin inhibits constitutive and EGF-induced kinase activities of EGFR (Afaq et. al., Int. J Cancer 2008; 123: 1508-1515) in breast cancer cell lines. Recently, we found that delphinidin treatment also inhibits protein expression of HER-2 in breast cancer cells. The goal of the present study was to determine the antiproliferative and proapoptotic potentials of delphinidin in ER-positive and ER-negative breast cancer cell lines that overexpress EGFR/HER2. We first determined whether blockade of EGFR/HER-2 signaling pathways by delphinidin exhibits growth inhibitory effect. Delphinidin (5-60 μM; 72 hrs) treatment of ER-positive (BT-474 and MCF-7) and ER-negative (MDA-MB-231 and MDA-MB-468) breast cancer cells resulted in a dose-dependent decrease in cell viability but had only minimal effects on normal mammary epithelial cell line, 184A1. The signaling pathways induced by activated EGFR/HER-2 include the PI3K/AKT and MAPK, both of which play a role in the mitogenic and cell survival responses. Therefore, we analyzed the expression of these proteins and found that delphinidin treatment of ER-positive and ER-negative breast cancer cells inhibited the activation of PI3K and phosphorylation of AKT and MAPKs. Treatment of BT-474 and MDA-MB-231 cells with delphindin (5-60 μM; 72 hrs) resulted in (i) cleavage of PARP protein, (ii) activation of caspase-3 and-9, (iii) downregulation of cyclin D1 and Bcl-2, and (iv) increased expression of p27 and Bax. Based on these observations, we suggest that delphinidin, alone or as an adjuvant to current therapies, could be useful for the management of both ER-positive and ER-negative breast cancers. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5422. doi:10.1158/1538-7445.AM2011-5422

Extraction and determination of polyphenols and betalain pigments in the Moroccan Prickly pear fruits (Opuntia ficus indica)

Polyphenols and betalain pigments respond property’s antioxidants benefic for the human health (Will Strike et al., 2002; Lecerf, 1999). The item present shows procured results for strengths in polyphenols and in betalains to the time in fruits and in their juice for the period 2008–2009. Our searches show that rate in polyphenols is more increased in the entire fruit than in only the juice. We have equally remarked that the red prickly pear contains amounts of polyphenols more raised than those of the yellow variety.This study has permitted us equally to value betalain pigments extracted from fruity juice. These are the betalains present in the epidermis and the pulp of the prickly pear confers on it its color varying from yellow to purple. Results show that yellow and red prickly pears contain imported strengths in betalains. Our work shows that the red prickly pear contains betaxanthin pigments in excess of the indicaxanthin that permits to valorize human’s potential spring of genuine colorings. Betalains and polyphenols are antioxidants that contribute to nutritional prickly pears’ quality and to their products of transformation.

뽕나무 오디를 이용한 cyanidin-3-glucoside 함유 천연식용색소 개발

뽕나무 오디함유 천연색소 C3G의 식용색소화 및 안정생산을 위한 최적 전처리 조건은 0.1 % citric acid-70% EtOH로서 오디로부터 제조한 순수 오디색소 분말의 수율은 8%이었다. 실제 농가에서 오디 색소 추출시 추출 시간 및 제조 비용을 대폭 절감할 수 있는 추출 방법을 개발하였다. 오디색소 추출시 품종의 선택이 중요하므로 C3G 함량이 높은 품종을 선택하도록 해야 할 것이다. 오디 색소 분말은 알코올, 물 및 밀가루 등 식품 재료나 가공방법에 상관없이 적색소의 특성이 유지되므로 다양한 식품에 적용하여 고부가가치를 향상시킬 수 있을 것으로 판단된다. Study on extraction and color characteristics of mulberry fruit pigment(C3G; cyanidin-3-glucoside) was performed to increase utilization as new source of natural food colorant. C3G was extracted with 0.1 % citric acid-70% EtOH. Then it was evaporated with large scale evaporation system. After adding dextrin to C3G concentration materials, we made pigment powder with freezing dryer.

Carotenoids and their isomers: color pigments in fruits and vegetables.

Fruits and vegetables are colorful pigment-containing food sources. Owing to their nutritional benefits and phytochemicals, they are considered as ‘functional food ingredients’. Carotenoids are some of the most vital colored phytochemicals, occurring as all-trans and cis-isomers, and accounting for the brilliant colors of a variety of fruits and vegetables. Carotenoids extensively studied in this regard include β-carotene, lycopene, lutein and zeaxanthin. Coloration of fruits and vegetables depends on their growth maturity, concentration of carotenoid isomers, and food processing methods. This article focuses more on several carotenoids and their isomers present in different fruits and vegetables along with their concentrations. Carotenoids and their geometric isomers also play an important role in protecting cells from oxidation and cellular damages.

Pigment Metabolism of ‘Sikitita’ Olive (Olea europaea L.): A New Cultivar Obtained by Cross-Breeding

The new olive cultivar 'Sikitita' was obtained from a cross between the 'Picual' and 'Arbequina' varieties. 'Sikitita' was selected for its features, making it particularly suited to high-density olive hedgerow orchards. From the standpoint of chloroplast pigment metabolism, the fruits of the 'Picual' and 'Arbequina' varieties have significant differences. It is therefore extremely interesting to analyze the descendants of both cultivars. With regard to chlorophyll catabolism, 'Sikitita' has proven to be a cultivar with low pigmentation and low levels of chlorophyllase activity. This is contrary to the findings obtained to date, where varieties with low pigmentation are a consequence of high chlorophyllase activity ('Arbequina') and highly pigmented fruits are due to low chlorophyllase activity ('Picual'). 'Arbequina' was, until recently, the only cultivar described that had developed a carotenogenic process, despite its anthocyanic ripening. However, from its father ('Arbequina'), the 'Sikitita' cultivar has inherited the pool of enzymes necessary to esterify xanthophylls at the chromoplast level. This makes 'Sikitita' a very interesting cultivar, with potential chemotaxonomic differences (such as esterified xanthophylls in the olive oils), and demonstrates the interest in genetic improvement programs for olive cultivars with different organoleptic characteristics.

Study of 'Redhaven' peach and its white-fleshed mutant suggests a key role of CCD4 carotenoid dioxygenase in carotenoid and norisoprenoid volatile metabolism.

BackgroundCarotenoids are plant metabolites which are not only essential in photosynthesis but also important quality factors in determining the pigmentation and aroma of flowers and fruits. To investigate the regulation of carotenoid metabolism, as related to norisoprenoids and other volatile compounds in peach (Prunus persica L. Batsch.), and the role of carotenoid dioxygenases in determining differences in flesh color phenotype and volatile composition, the expression patterns of relevant carotenoid genes and metabolites were studied during fruit development along with volatile compound content. Two contrasted cultivars, the yellow-fleshed 'Redhaven' (RH) and its white-fleshed mutant 'Redhaven Bianca' (RHB) were examined.ResultsThe two genotypes displayed marked differences in the accumulation of carotenoid pigments in mesocarp tissues. Lower carotenoid levels and higher levels of norisoprenoid volatiles were observed in RHB, which might be explained by differential activity of carotenoid cleavage dioxygenase (CCD) enzymes. In fact, the ccd4 transcript levels were dramatically higher at late ripening stages in RHB with respect to RH. The two genotypes also showed differences in the expression patterns of several carotenoid and isoprenoid transcripts, compatible with a feed-back regulation of these transcripts. Abamine SG - an inhibitor of CCD enzymes - decreased the levels of both isoprenoid and non-isoprenoid volatiles in RHB fruits, indicating a complex regulation of volatile production.ConclusionsDifferential expression of ccd4 is likely to be the major determinant in the accumulation of carotenoids and carotenoid-derived volatiles in peach fruit flesh. More in general, dioxygenases appear to be key factors controlling volatile composition in peach fruit, since abamine SG-treated 'Redhaven Bianca' fruits had strongly reduced levels of norisoprenoids and other volatile classes. Comparative functional studies of peach carotenoid cleavage enzymes are required to fully elucidate their role in peach fruit pigmentation and aroma.

Study of the influence of berry-blanching on syneresis in blueberry purées

A wider range of fruit-based products, preserving high nutritional and sensory attributes, aid towards achieving a healthy diet. Pigmented fruit, specially blueberries, are a natural source of phenolic phytochemicals which could be exploited through proper technologies, limiting by-product loss and detrimental impact of processing on fruit bioactive compounds. The influence of the mild thermal pretreatment of steam blanching on the syneresis of frozen blueberry purées was studied. Syneresis kinetic parameters, colour and phenolic profile of the serum from syneresis were analyzed, highlighting the relationship between quality data and pigment localization in the berry by a correlative microscopy study. Blanching induced a marked anthocyanin diffusion from the vacuoles of pigmented epidermal cells down to the core of berries. Monomeric anthocyanin pigments (MAP) decreased in blanched (BL) berries (−17.6%), but no further decrease was observed in BL purées, while puréeing of not blanched (NB) berries caused a significant loss in MAP (−28.0%) and total phenolic compounds (TPC) (−35.8%) and an increase in percent polymeric color. As a result BL purées had higher MAP and TPC compared to NB ones. A different syneresis behaviour was evidenced between BL and NB purées and a model fitting data was proposed; the retention of syneresis serum was higher in BL purée during the first 180min after thawing, then it was higher in NB ones till 340min, and thereafter was similar. BL serum samples showed higher TPC (+69%), MAP (+215%), lightness (L*) and red colour component (a*) compared to NB ones. The introduction of a berry-blanching step improves physical and chemical stability of ready-to-eat frozen blueberry purées. Correlative microscopy provides a useful integrated approach to fulfil quality requirements.

Apple skin patterning is associated with differential expression of MYB10

BackgroundSome apple (Malus × domestica Borkh.) varieties have attractive striping patterns, a quality attribute that is important for determining apple fruit market acceptance. Most apple cultivars (e.g. 'Royal Gala') produce fruit with a defined fruit pigment pattern, but in the case of 'Honeycrisp' apple, trees can produce fruits of two different kinds: striped and blushed. The causes of this phenomenon are unknown.ResultsHere we show that striped areas of 'Honeycrisp' and 'Royal Gala' are due to sectorial increases in anthocyanin concentration. Transcript levels of the major biosynthetic genes and MYB10, a transcription factor that upregulates apple anthocyanin production, correlated with increased anthocyanin concentration in stripes. However, nucleotide changes in the promoter and coding sequence of MYB10 do not correlate with skin pattern in 'Honeycrisp' and other cultivars differing in peel pigmentation patterns. A survey of methylation levels throughout the coding region of MYB10 and a 2.5 Kb region 5' of the ATG translation start site indicated that an area 900 bp long, starting 1400 bp upstream of the translation start site, is highly methylated. Cytosine methylation was present in all three contexts, with higher methylation levels observed for CHH and CHG (where H is A, C or T) than for CG. Comparisons of methylation levels of the MYB10 promoter in 'Honeycrisp' red and green stripes indicated that they correlate with peel phenotypes, with an enrichment of methylation observed in green stripes.ConclusionsDifferences in anthocyanin levels between red and green stripes can be explained by differential transcript accumulation of MYB10. Different levels of MYB10 transcript in red versus green stripes are inversely associated with methylation levels in the promoter region. Although observed methylation differences are modest, trends are consistent across years and differences are statistically significant. Methylation may be associated with the presence of a TRIM retrotransposon within the promoter region, but the presence of the TRIM element alone cannot explain the phenotypic variability observed in 'Honeycrisp'. We suggest that methylation in the MYB10 promoter is more variable in 'Honeycrisp' than in 'Royal Gala', leading to more variable color patterns in the peel of this cultivar.

NON-INVASIVE SENSING OF FRUIT DEVELOPMENT IN BANANA AND PAPAYA BY MEANS OF A SPECTROSCOPIC APPROACH

In climacteric tropical fruit such as banana and papaya characteristic pigment changes occur during fruit development in the supply chain. A handheld spectrophotometer has been developed to undertake fruit pigment readings in-situ. Using fruit spectra obtained in optical geometry for non-invasive remittance analysis in-situ as well as diffuse reflectance readings with a laboratory spectrophotometer, spectral variations were analyzed in fruits at different maturity stages as well as after chilling injury treatment. Whole spectra calibration models and two-wavelength indices were evaluated and implemented in the embedded software of the handheld device. In banana and papaya, fruit maturity development and the appearance of chilling injury (CI) symptoms were monitored. Fruit maturity was estimated by means of chlorophyll changes analysed in the red and near infrared wavelength ranges of the fruit spectra. The occurrence of CI was monitored using autocorrelation analysis of the time series.

Non-destructive analysis of anthocyanins in cherries by means of Lambert–Beer and multivariate regression based on spectroscopy and scatter correction using time-resolved analysis

In high-value sweet cherry ( Prunus avium), the red coloration – determined by the anthocyanins content – is correlated with the fruit ripeness stage and market value. Non-destructive spectroscopy has been introduced in practice and may be utilized as a tool to assess the fruit pigments in the supply chain processes. From the fruit spectrum in the visible (Vis) wavelength range, the pigment contents are analyzed separately at their specific absorbance wavelengths. A drawback of the method is the need for re-calibration due to varying optical properties of the fruit tissue. In order to correct for the scattering differences, most often the spectral intensity in the visible spectrum is normalized by wavelengths in the near infrared (NIR) range, or pre-processing methods are applied in multivariate calibrations. In the present study, the influence of the fruit scattering properties on the Vis/NIR fruit spectrum were corrected by the effective pathlength in the fruit tissue obtained from time-resolved readings of the distribution of time-of-flight (DTOF). Pigment analysis was carried out according to Lambert–Beer law, considering fruit spectral intensities, effective pathlength, and refractive index. Results were compared to commonly applied linear color and multivariate partial least squares (PLS) regression analysis. The approaches were validated on fruits at different ripeness stages, providing variation in the scattering coefficient and refractive index exceeding the calibration sample set. In the validation, the measuring uncertainty of non-destructively analyzing fruits with Vis/NIR spectra by means of PLS or Lambert–Beer in comparison with combined application of Vis/NIR spectroscopy and DTOF measurements showed a dramatic bias reduction as well as enhanced coefficients of determination when using both, the spectral intensities and apparent information on the scattering influence by means of DTOF readings. Corrections for the refractive index did not render improved results.