Apple Fruit Skin Research Articles

Epoxidation catalysts

The major component of the cuticular hydroxyfatty acid polymer, cutin, of spinach leaf was identified as 18-hydroxy-cis-9,10-epoxystearic acid. A 3000g particulate fraction obtained from the homogenates of young spinach leaves catalyzed the epoxidation of 18-hydroxyoleic acid to 18-hydroxy-cis-9,10-epoxystearic acid. The epoxy acid was identified by thin-layer chromatography of the product of hydration of the oxirane function and subsequent reduction (octadecane-1,9,10,18-tetraol), on silica gel G and boric acid impregnated silica gel, and by radio gas-liquid chromatography of the periodate oxidation product of the tetraol derived from the epoxy acid. This enzymatic epoxidation required ATP, CoA, NADPH, and O2 for maximal activity. The pH optimum for epoxidation was 9.0, and the epoxidase activity was located mainly in the 3000g particulate fraction, which contained the cuticular membranes. The rate of epoxidation of the unsaturated acid was linear up to 2 h and up to a protein concentration of 1.5 mg/ml. The apparent Km and V of the enzyme was 7.5 × 10−5m and 180 nmol/1-h-mg protein, respectively. That the epoxidase has a stringent substrate specificity was indicated by the observation that 18-hydroxyelaidic acid, 18-acetoxyoleic acid, and oleic acid were not readily epoxidized by this enzyme preparation. Light reversible inhibition by CO suggested the involvement of cytochrome P-450 in the epoxidation. Chelators inhibited epoxidation, and this inhibition was reversed by FeSO4. The epoxidase was inhibited 30% by 0.5 mm NaCN, whereas the same concentration of NaN3 showed very little inhibition. The present epoxidase is unique in that it is associated with the 3000g particles which contain the cutin polymer and in that it requires activation of the carboxyl group distal to the epoxidation site. Enzyme preparations capable of epoxidizing 18-hydroxyoleic acid were also obtained from the skin of apple fruit and from the epidermis of Senecio odoris leaves. This is the first report on an enzyme involved in the biosynthesis of the C18 family of cutin monomer acids.

Developmental stages affect characteristics of wax apple fruit skin discs cultured with sucrose—with special reference to color

Wax apple ( Syzygium samarangense Merr. & Perry) fruit skin discs from different fruit development stages incubated with and without sucrose showed differential effects on diameter, weight, soluble solids (SSs) and skin color. Diameter, weight and chlorophyll content declined, while SSs and anthocyanin concentrations increased in a sucrose medium. Both SS and anthocyanin concentrations in the sucrose-added discs were higher than in the non-sucrose controls possibly at the expenses of diameter, weight, chlorophyll and soluble protein in disks taken from more mature fruits. Fruits from the rapid growth stage to red stage, i.e. from 4 to 8 weeks after anthesis, had greater anthocyanin induction in the presence of sucrose.

Germination and adhesion of fungal conidia on polycarbonate membranes and on apple fruit exposed to mycoactive acetate esters.

The adhesion and germination of conidia of nine fungal species were assessed on polycarbonate membranes or on the skin of apple fruit in sealed glass bottles injected or not injected with acetate esters. Adhesion was determined after dislodging conidia from surfaces using a sonication probe. Adhesion and germination of conidia of Aspergillus flavus, Aspergillus fumigatus, Aspergillus niger, Penicillium citrinum, Penicillium claviforme, or Trichoderma sp. on membranes after 48 h were not increased in a 1.84 microg mL(-1) headspace of butyl acetate (BA), ethyl acetate, hexyl acetate, 2-methylbutyl acetate, pentyl acetate, or propyl acetate. Adhesion and germination of Botrytis cinerea, Penicillium expansum, and Penicillium roquefortii conidia were stimulated by all esters. Only conidia of B. cinerea and P. expansum exhibited increased adhesion and germination on the skin of apple fruit in bottles exposed to 0.92 microg mL(-1) of BA. Only conidia of B. cinerea and P. expansum produced decay in inoculated puncture wounds on fruit. Freshly made puncture wounds or 24-h-old puncture wounds in fruit were more adhesive than the unpunctured skin of fruit to conidia of B. cinerea or P. expansum. Fresh wounds were more adhesive to both fungi than 24-h-old puncture wounds. The skin and wounds of fruit were as adhesive to B. cinerea conidia as they were to P. expansum conidia. A 4-h exposure to 1.43 microg mL(-1) of BA increased adhesion of B. cinerea and P. expansum conidia in 24-h-old wounds. Results suggest that acetate-ester stimulation most likely is not a rare phenomenon in the fungi. For nutrient-dependent decay pathogens of apple fruit, acetate esters may be an alternative chemical cue used to maintain adhesion of conidia to wound surfaces.

Accumulation of anthocyanins and quercetin glycosides in ‘Gala’ and ‘Royal Gala’ apple fruit skin with UV-B–Visible irradiation: modifying effects of fruit maturity, fruit side, and temperature

We measured the accumulation of the anthocyanins and quercetin glycosides, after irradiation with UV-B–Visible light, in the skin of apple fruit ( Malus domestica Borkh.), ‘Gala’ and ‘Royal Gala’, at two temperatures, 10 and 20°C for five harvests up to 2 weeks after the start of the commercial harvest. The response to irradiation was compared between sides of the fruit that were outer facing (exposed) and inner facing (shaded) when on the tree. The tree-shaded side of the fruit had a much greater potential to accumulate anthocyanins and quercetin glycosides than the tree-exposed side. This potential of the detached fruit to accumulate anthocyanin in the skin with UV-B–Visible radiation was decreased by fruit maturity at the last harvest. Lower temperature of 10°C prevented the accumulation of anthocyanins and quercetin glycosides compared with 20°C during irradiation with UV-B–Visible light. ‘Royal Gala’, the red sport of ‘Gala’, accumulated anthocyanins, but not quercetin glycosides, to higher concentrations than ‘Gala’. The results show that maturity, the fruit skin’s previous exposure to light and temperature are major modifying factors in the accumulation of anthocyanins by ‘Gala’ and its sport ‘Royal Gala’.

Light, Temperature, and Sucrose Affect Color, Diameter, and Soluble Solids of Disks of Wax Apple Fruit Skin

A study was conducted to assess the combined effects of light, temperature and sucrose on color, weight, diameter, and soluble solids of the skin of wax apple (Syzygium samarangense Merr. & Perry) fruits. Skin disks were cultured in a factorial arrangement of two light levels [dark or light (300 μmol·m-2·s-1)] as subplots and three sucrose concentrations (0%, 3%, or 6%) as sub-subplots within three temperature levels (20, 25, or 30 °C) as whole plot treatments. Weight, diameter, soluble solids concentration (SSC), and anthocyanin content were measured 2 weeks after incubation. Light increased SSC and anthocyanin, but reduced the increase in weight and diameter. Increasing the temperature limited increase in diameter and anthocyanin content. Weight, SSC, and anthocyanin contents increased in a linear fashion with concentration of sucrose in the culture solution. However, none of the three factors played a unique role in anthocyanin synthesis in wax apple. Among the 18 combinations, light/20°C/6% sucrose gave the highest SSC and anthocyanin content, while dark/20°C/6% sucrose produced the largest diameter.

A procedure for quantifying adhesion of conidia of <i>Botrytis cinerea </i>to the skin of apple fruit

A procedure for quantifying adhesion of conidia of <i>Botrytis cinerea </i>to the skin of apple fruit

Aglycone and glycoside specificity of apple skin flavonoid glycosyltransferase

O-Glycosyltransferase(s) extracted from apple (Malus pumila Mill) fruit skin showed activity towards a range of flavonols and anthocyanins. However, no glycosylating activity was shown towards a dihyroflavonol (dihydroquercetin), a flavanone (eriodictyol) or a flavone (luteolin). The enzyme preparation glycosylated those flavonoids normally present in apple skins (quercetin and cyanidin) and in addition several other related compounds (delphinidin, fisetin, isorhamnetin, kaempferol, myricetin and pelargonidin). This enzyme(s) specifically transferred the glycosyl moiety from sugar nucleotide donors to the 3-position of the flavonoid nucleus. Only flavonoid 3-glycosides occur naturally in apple skin. Activity with different sugar donors was in the order galactose>glucose>xylose, which reflected the ratios of cyanidin and quercetin glycosides found in apple fruit skin. There were slight differences in the relative UFGT activity with quercetin and the three different sugar donors between ‘Granny Smith’ and ‘Splendour’, and this was reflected by similar differences in the ratios of endogenous quercetin glycosides. ©1997 SCI

Developmental changes in the phenol concentrations of ‘Golden delicious’ apple fruits and leaves

The phenolic content of apple fruit skin and leaves was determined at the developmental stage of each organ. Phenolic levels decreased on a dry weight basis during the seasonal development of fruits and leaves with respect to their ontogenesis but the single compounds did not behave uniformly. A shift in flavanol pools from monomeric to oligomeric structures during fruit growth indicated the biosynthetic tendency towards the formation of procyanidins at the end of the growing period. Among the procyanidins identified epicatechin-(4β,6)-epicatechin-(4β,6)-epicatechin has not been reported previously from apple.

Biosynthesis of hydroxyfatty acid polymers: Enzymatic epoxidation of 18-hydroxyoleic acid to 18-hydroxy- cis-9,10-epoxystearic acid by a particulate preparation from spinach ( Spinacia oleracea)

The major component of the cuticular hydroxyfatty acid polymer, cutin, of spinach leaf was identified as 18-hydroxy- cis-9,10-epoxystearic acid. A 3000 g particulate fraction obtained from the homogenates of young spinach leaves catalyzed the epoxidation of 18-hydroxyoleic acid to 18-hydroxy- cis-9,10-epoxystearic acid. The epoxy acid was identified by thin-layer chromatography of the product of hydration of the oxirane function and subsequent reduction (octadecane-1,9,10,18-tetraol), on silica gel G and boric acid impregnated silica gel, and by radio gas-liquid chromatography of the periodate oxidation product of the tetraol derived from the epoxy acid. This enzymatic epoxidation required ATP, CoA, NADPH, and O 2 for maximal activity. The pH optimum for epoxidation was 9.0, and the epoxidase activity was located mainly in the 3000 g particulate fraction, which contained the cuticular membranes. The rate of epoxidation of the unsaturated acid was linear up to 2 h and up to a protein concentration of 1.5 mg/ml. The apparent K m and V of the enzyme was 7.5 × 10 −5 m and 180 nmol/1-h-mg protein, respectively. That the epoxidase has a stringent substrate specificity was indicated by the observation that 18-hydroxyelaidic acid, 18-acetoxyoleic acid, and oleic acid were not readily epoxidized by this enzyme preparation. Light reversible inhibition by CO suggested the involvement of cytochrome P-450 in the epoxidation. Chelators inhibited epoxidation, and this inhibition was reversed by FeSO 4. The epoxidase was inhibited 30% by 0.5 m m NaCN, whereas the same concentration of NaN 3 showed very little inhibition. The present epoxidase is unique in that it is associated with the 3000 g particles which contain the cutin polymer and in that it requires activation of the carboxyl group distal to the epoxidation site. Enzyme preparations capable of epoxidizing 18-hydroxyoleic acid were also obtained from the skin of apple fruit and from the epidermis of Senecio odoris leaves. This is the first report on an enzyme involved in the biosynthesis of the C 18 family of cutin monomer acids.

Biosynthesis of hydroxyfatty acid polymers: Enzymatic hydration of 18-hydroxy- cis-9,10-epoxystearic acid to threo-9,10,18-trihydroxystearic acid by a particulate preparation from apple ( Malus pumila)

A 3000 g participate fraction prepared from homogenates of the skin of young apple fruit catalyzed the hydration of 18-hydroxy- cis-9,10-epoxystearic acid to threo-9,10,18-trihydroxystearic acid, a key step in the biosynthesis of cutin, the hydroxyfatty acid biopolymer of the plant cuticle. Similar preparations from the internal tissue of apple fruit contained less than 6% of this epoxide hydrase activity. The enzymatic product was identified by thin-layer chromatography on silica gel G and on boric acid-silica gel G, as well as by radio gas-liquid chromatography of its periodate oxidation product. The epoxide hydrase required no cofactors, had a pH optimum of 6.5, and it was located mainly (>90%) in the 3000 g particle fraction which contained the cuticular membranes. The rate of hydration of the epoxy acid was linear up to 75 min and up to a protein concentration of 0.8 mg/ml. The apparent K m and V of the enzyme was 4 × 10 −4 m and 170 μmol/1-h-mg of protein, respectively. p-Chloromercuribenzoate strongly inhibited the enzyme (80% inhibition at 10 −4 m), while iodoacetamide and N-ethylmaleimide were less potent inhibitors. 1,1,1-Trichloropropylene-2,3-oxide at 5 × 10 −3 m gave 60% inhibition of hydration of 18-hydroxy-9,10-epoxystearic acid. Diisopropylfluorophosphate was mildly inhibitory, and the enzyme was insensitive to ethylenediaminetetraacetic acid. Methyl 18-hydroxy-9,10-epoxystearate, 18-acetoxy-9,10-epoxystearic acid, methyl 18-acetoxy-9,10-epoxystearate, 9,10-epoxystearic acid, and styrene oxide were not readily hydrated by this plant epoxide hydrase showing that it has a fairly stringent substrate specificity, unlike the catabolic epoxide hydrases of animals. Particulate preparations capable of hydrating 18-hydroxy-9,10-epoxystearic acid to the trihydroxy acid were also obtained from Senecio odoris leaf epidermis and Spinacia oleracea leaves. This is the first report on the enzyme which catalyzes this important step in the biosynthesis of the protective biopolymer, cutin.

Direct evidence for the involvement of epoxide intermediates in the biosynthesis of the C 18 family of cutin acids

Cutin, the structural component of plant cuticle, is a polymer of C 16 and C 18 hydroxy fatty acids. Previous results have suggested that oleic acid undergoes ω-hydroxylation, epoxidation of the double bond, and, finally, hydration of the epoxide to give rise to the three major components of the C 18 family of cutin acids. 18-Hydroxy [18- 3H]oleic acid and 18-hydroxy-9,10-epoxy[18- 3H]stfaric acid have been synthesized and, with these synthetic substrates, the conversion of 18-hydroxyoleic acid to 18-hydroxy-9,10-epoxystearic acid and the hydrolysis of 18-hydroxy-9,10-epoxystearic acid to 9,10,18-trihydroxystearic acid were directly demonstrated in apple fruit skin and in the leaves of apple and Senecio odoris. Trichloropropene oxide, an inhibitor of microsomal epoxide hydrases of animals, specifically inhibited the conversion of [1- 14C]oleic acid into 18-hydroxy-9,10-epoxystearic acid and 9,10,18-trihydroxystearic acid, while it had no effect on the conversion of [1- 14C]palmitic acid into hydroxylated palmitic acid, a process which does not involve epoxy acid intermediates. Therefore, it appears that this inhibitor affects epoxidation and or epoxide hydration steps involved in cutin biosynthesis.

Penetration of surface-applied45Ca into apple fruitt

SummaryThe uptake of 45Ca applied to the skin of apple fruit was examined in the laboratory with regard to the practice of spraying with calcium solutions to prevent bitter pit.The speed of penetration of 45Ca from a drop on the fruit surface was found to be much higher under conditions of 50–80% relative humidity of the air than at saturation. Evaporation of the drop apparently strongly influenced the absorption of Ca into the apple. Penetration was faster in cv. Cox’s Orange Pippin than in James Grieve. The Ca which entered the fruit remained either in the skin or immediately beneath it.When 45Ca was applied more than once, the subsequent treatments, especially those applied at high relative humidity, were less effective. It is concluded that for practical purposes it would be advisable to spray repeatedly when the relative humidity is low.