Enzymatic browning is one of the most important reactions that occur in fruits and vegetables, usually resulting in negative effects on color, taste, flavor, and nutritional value. The reaction is a consequence of phenolic compounds' oxidation by polyphenol oxidase (PPO), which triggers the generation of dark pigments. This is particularly relevant for apples, which are rich in polyphenols and highly susceptible to enzymatic browning. The objective of the present work was to quantify enzymatic browning and PPO activity and identify and quantify target polyphenols in apple [Malus ×sylvestris (L.) Mill. var. domestica (Borkh.) Mansf.] pulp in the cultivars (cvs.) Aori27, Elstar, Fuji, and Mellow at three fruit developmental stages (FDS). The enzymatic browning was quantified by tristimulus colorimetry; PPO activity was quantified by an enzyme–substrate spectrophotometric assay; phenolic compounds were determined and quantified by reverse-phase high-performance liquid chromatography–ultraviolet/visible–mass spectrometry. Enzymatic browning showed significant difference among cvs. and FDSs and interaction between both factors. PPO activity showed significant difference among cultivars and FDSs. A significant difference was evidenced for polyphenol content among cultivars and FDSs with interaction between both factors. Chlorogenic acid was the major phenolic compound in ‘Aori27’ and ‘Mellow’. In ‘Fuji’, chlorogenic acid and (–)-epicatechin were the major phenolics and in ‘Elstar’ (–)-epicatechin and procyanidin B2 were the major phenolics at different FDSs. The enzymatic browning showed high correlation to polyphenol content in all cultivars and high correlation was observed between browning and PPO activity in ‘Aori27’ and ‘Elstar’. The magnitude of the correlation between browning and polyphenols and PPO activity is genotype-specific. At the commercial harvest, ‘Fuji’ showed the highest polyphenol content and ‘Aori27’ showed the lowest level for enzymatic browning. Chemical names used: 3-(3,4-dihydroxycinnamoyl) quinic acid (chlorogenic acid), (–)-cis-3,3′,4′,5,7-pentahydroxyflavane (epicatechin), and cis,cis″-4,8″-Bi(3,3′,4′,5,7-pentahydroxyflavane) (procyanidin B2).

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Expression of MdACS3a , one of the ripening-related ACC synthase genes, plays a pivotal role in initiating the burst of ethylene production by MdACS1 in apple fruit. Although previous studies have demonstrated the presence of MdACS3a -null alleles through deficiency of transcription activity or loss of enzyme activity due to amino acid substitution, which may affect the storage properties of certain fruit cultivars, an overall picture of these null alleles in cultivars is still lacking. The present study investigated the distribution of null allelic genes in 103 cultivars and 172 breeding selections by using a simple sequence repeat (SSR) marker linked to them. The results indicated that both allelic genes were widely distributed throughout the examined cultivars and selections, some occurring as the null genotype, either homozygously or heterozygously, with each null allele. The implications of MdACS3a distribution results and the influence of its null allelotypes in fruit characters are discussed.

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