Malus Pumila Cv Research Articles

Anti-Inflammatory, Antioxidant, and Genoprotective Effects of Callus Cultures Obtained from the Pulp of Malus pumila cv Miller (Annurca Campana Apple).

Apples are rich in phytochemicals useful for human health. However, environmental factors can greatly affect the accumulation of these compounds. To face this problem, the callus culture technique was used to obtain large quantities of phytochemicals. Specifically, two callus cultures were obtained from ripe Annurca apple pulp (Malus pumila cv Miller) and cultivated under different light conditions: darkness and an 18-h photoperiod. The hydro-alcoholic extracts from the calli underwent analysis using GC-MS, GC-FID, and HPLC-DAD-ESI-MSn to determine the qualitative and quantitative content of phenolic and triterpenic acids. The study revealed the predominant presence of triterpenic compounds in both calli. Furthermore, we investigated their radical scavenging and antioxidant activities through DPPH, ABTS, ORAC assays, and lipoxygenase inhibition activity. Genoprotection was evaluated via nicking assay, and the anti-inflammatory effect was investigated via Griess assay on LPS-injured murine macrophages. All the analyses performed were compared with peel and pulp hydroalcoholic extracts. The results showed that both calli primarily show anti-inflammatory activity and moderate antioxidant effect and can protect DNA against oxidative stimuli. This data encouraged further research aimed at utilizing callus as a bioreactor to produce secondary metabolites for use in preventive and therapeutic applications to combat acute or chronic age-associated diseases.

Cover Cropping Increases Soil Fungal-Bacterial Community Diversity and Network Complexity in Apple Orchards on the Loess Plateau, China

Soil microorganisms play a key role in soil fertility. Exploring the microbial community composition and diversity in response to cover crops is important for improving soil fertility in orchards. This study investigated how cover cropping can improve soil fertility by altering microbial community composition and the interrelations among soil microorganisms. Soil physicochemical and biological indicators and microbial community composition were evaluated after a 6-years application of cover cropping in an apple (Malus pumila cv. Fuji) orchard located on the Loess Plateau, China. Three treatments were applied: Trifolium repens treatment (TR), Lolium perenne treatment (LP), clear tillage treatment (CT). The soil of the cover crop treatments had a more complex microbial co-occurrence network than that of the clear tillage treatment; in particular, the correlations among fungi were significantly increased. Cover cropping increased the levels of soil organic matter (SOM), microbial biomass carbon (MBC), and total nitrogen (TN) in orchard soil. Changes in the fungal community were more related to microbial biomass nitrogen (MBN), TN, and carbon-nitrogen ratio (C:N). This indicates that cover cropping not only increases the diversity of soil microorganisms but also increases the interrelations between microbial groups, thereby improving the soil fertility of apple orchards on the Loess Plateau.

Construction of full-length infectious cDNA clones of Apple stem grooving virus using Gibson Assembly method

Apple stem grooving virus (ASGV) belongs to the genus Capillovirus within the family Betaflexiviridae. In this work, we described the construction of full-length infectious cDNA clones of ASGV isolate jilin-shaguo (JL-SG) using the Gibson Assembly approach (New England BioLabs). The isolate was previously detected in a Chinese pear-leaf crab apple (Malus asiatica Nakai.) in Baicheng, Jilin province, China. Two full-length cDNA clones of ASGV JL-SG were obtained, and they are identical to each other in sequence. The full-length cDNA clone was infectious on Chenopodium quinoa, Nicotiana glutinosa, and N. occidentalis 37B via agroinfiltration. Through sap inoculation, the infection was additionally spread to C. amaranticolor. N. benthamiana could not be infected, neither through agroinfiltration nor sap inoculation. In infected herbaceous plants, typical ASGV particles with morphology of flexuous filaments were observed by transmission electron microscope (TEM). Moreover, seeds of infected N. glutinosa and N. occidentalis 37B were collected and germinated, the seedlings were ASGV-free in RT-PCR test, suggesting ASGV JL-SG is not seed-transmissible in the tested Nicotiana species. In addition, the cDNA clone was agroinfiltrated into seedlings of Malus pumila cv. Fuji. The infection was symptomless, and can be spread to C. quinoa via sap inoculation, causing typical symptoms. ASGV JL-SG was also detected by RT-PCR in the infected Fuji plants, however, no virion was observed by TEM.

미성숙 사과의 최종당화산물 생성 저해물질의 동정

미성숙 사과의 최종당화산물 생성 저해물질의 동정

미성숙 사과의 항산화 및 tyrosinase 저해 활성 평가

In this study, the antioxidant and tyrosinase inhibitory activities of aqueous ethanolic extract from the immature fruits of Malus pumila cv. Fuji were evaluated. The antioxidant capacities of the extract was investigated employing radical scavenging assays using DPPH and ABTS+ radicals. The skin-whitening effect of M. pumila cv. Fuji extract was tested using mushroom tyrosinase assay. In addition, the total phenolic content was measured by a spectrophotometric analysis. All tested samples showed a dose-dependent radical scavenging and tyrosinase inhibitory activities. Among the tested samples, the ethyl acetate (EtOAc)-soluble portion from the immature fruits of M. pumila cv. Fuji was showed the significant DPPH and ABTS+ radicals scavenging activities. Also, the tyrosinase inhibitory activity of the ethyl acetate (EtOAc)-soluble portion from immature apples was higher than other solvent-soluble portion. These results suggest that unripe fruits of M. pumila cv. Fuji could be considered as a new valuable source of natural antioxidant and skin-whitening agents. Systematic investigation of immature fruits of Malus pumila cv. Fuji will be performed for the further development of its biological properties.

Oral administration of apple condensed tannins delays rheumatoid arthritis development in mice via downregulation of T helper 17 (Th17) cell responses.

Apples are known to contain high concentrations of phenolic compounds such as condensed tannins. Consumption of condensed tannins has been reported to reduce the risk of many types of chronic diseases including allergies. However, their therapeutic effectiveness and potential in treating autoimmune disease remain controversial. Here, the effect of oral administration of apple condensed tannins (ACT) prepared from apples (Malus pumila cv. Fuji) on bovine type II collagen (CII)-induced arthritis in DBA1/J mice, a well-established murine model of human rheumatoid arthritis (RA), was evaluated. As compared to the control (without ACT administration) group, RA development was delayed and a significant reduction in the RA clinical score was observed in the ACT-administered group. Using cultured splenocytes isolated from CII-immunized mice, ACT-administration was shown to decrease the CII-induced increases in IL-17 expression and production in vitro. We propose that downregulation of T helper (Th) 17 cells is responsible for the ACT-induced RA suppression.

Effects of unripe apple polyphenols on the expression of matrix metalloproteinase-1 and type-1 procollagen in ultraviolet irradiated human skin fibroblasts

Bioassay-guided isolation of an aqueous ethanolic extract from the unripe fruits of Malus pumila cv. Fuji using matrix metalloproteinase-1 (MMP-1) inhibition and type-1 procollagen inducing assays led to isolation of eight phenolic compounds, reynoutrin (1), quercitrin (2), isoquercitrin (3), phloridzin (4), phloretin-2′-O-xyloglucoside (5), 5-O-p-courmaroylquinic acid (6), chlorogenic acid methyl ester (7), and chlorogenic acid (8). The structures were established on the basis of nuclear magnetic resonance and mass spectroscopic data interpretations. All isolates were evaluated for their inhibitory effects on MMP-1, and compounds 1, 2, and 6 exhibited potent inhibitory effects on MMP-1 with IC50 values of 1.3±0.1, 1.4±0.1, and 3.2±0.6 μM, respectively. When evaluated for their effects on type I procollagen synthesis on human fibroblast cells, compounds 1 and 3 were respectively found to display potent activity for induction of type-1 procollagen by 78.5 and 78.6% at the tested concentration of 12.5 M.

Isolation and structure elucidation of tetrameric procyanidins from unripe apples (Malus pumila cv. Fuji) by NMR spectroscopy

Procyanidins are plant secondary metabolites widely consumed and known to have various physiological functions, but their bioavailability and mechanism of action are still unclear especially for larger oligomers. One of the reasons is scarce information about the detailed structure of oligomeric procyanidins. As for apple, structures of procyanidin components larger than trimers are scarcely known. In this study, 11 tetrameric procyanidins including two known compounds were isolated from unripe apples (Malus pumila cv. Fuji) and identified by NMR spectroscopic analysis and phloroglucinol degradation. As a result, the detailed structural diversity of tetrameric procyanidins in apple was established.

Critical responses of photosynthetic efficiency of goldspur apple tree to soil water variation in semiarid loess hilly area

Goldspur apple (Malus pumila cv. Goldspur) is one of the main fruit trees planted in semiarid loess hilly areas. The photosynthetic efficiency in leaves of eight-year-old trees were studied under different soil water conditions with a Li-6400 portable photosynthesis system and a Li-Cor1600 portable steady state porometer in order to explore the effects of soil water stress on photosynthesis and the suitable soil water content (SWC) for water-saving irrigation of apple orchards. The results showed that the leaf net photosynthetic rate (P N), transpiration rate (E), water-use efficiency (WUE), stomatal conductance (g s), intercellular CO2 concentration (C i), and stomatal limiting value (L s) displayed different threshold responses to soil water variation. When SWC was within a range of about 60%-86% of field capacity (FC), P N and E were maintained in a relative steady state. At an elevated level but below 60% of FC, both P N and E decreased evidently with decreasing soil moisture. The SWC needed to support WUE in a relatively steady state and at a high level was in the range of about 50%-71% of FC. When SWC was less than 48% of FC, g s and L s declined with decreasing soil moisture, while C i increased rapidly. Based on the analysis of the stomatal limitation of photosynthesis using two criteria (C i and L s) suggested by Farquhar and Sharkey, it was implied that the predominant cause of restricting P N had changed from stomatal limitation to nonstomatal one under severe water stress. In terms of water-saving irrigation for enhancing water-use efficiency, it was concluded that in semiarid loess hilly areas, the suitable range of SWC for water-saving irrigation in goldspur apple orchards is in the range of about 50%-71% of FC, and the most severe degree of soil water stress tolerated for photosynthesis is about 48% of FC.

Responses of Gas Exchange Parameters of Goldspur Apple Tree to Soil Water Variation

Goldspur Apple (Malus pumila cv. Goldspur) is one of the main fruit trees planted on the Loess Plateau in a semi-arid region. Gas exchange parameters in leaves of ten-year-old trees were studied under different soil water conditions with a LI-6200 portable photosynthesis system and a LI-1600 portable steady state porometer, in order to explore the effects of soil water stress on the photosynthesis and the suitable soil water content (SWC) for water-saving irrigation of the apple orchard.

Effect of Cryopreservation on Apple Genetic Resources at Morphological, Chromosomal, and Molecular Levels

Shoot tips of three apple genotypes, namely, Malus pumila cv. M26, Gala, and Hokkaido No. 9, were successfully cryopreserved using a modified encapsulation-dehydration method. As a result, in addition to a high survival rate and regeneration rate, the capacity of shoots regenerated from cryopreserved samples to root was enhanced. Eight M26 single-bud sibling lines were used to assess genetic stability. Although cytological examination revealed a ploidy difference in the noncryopreserved control, the ploidy constitution remained relatively stable during the period of cryopreservation. Amplified fragment length polymorphism (AFLP) assay was performed to detect DNA-level variation. No change in DNA fragment pattern and number was observed between the control and the cryopreserved samples. In addition, methylation-sensitive amplified polymorphism (MSAP) assay was carried out to investigate the DNA methylation status during the period of cryopreservation. It was found that cryopreservation induced a decrease in DNA methylation level.

Identification of Catechin Oligomers from Apple (Malus pumila cv. Fuji) in Matrix‐assisted Laser Desorption/Ionization Time‐of‐flight Mass Spectrometry and Fast‐atom Bombardment Mass Spectrometry

Identification of Catechin Oligomers from Apple (Malus pumila cv. Fuji) in Matrix‐assisted Laser Desorption/Ionization Time‐of‐flight Mass Spectrometry and Fast‐atom Bombardment Mass Spectrometry

Identification of Catechin Oligomers from Apple (Malus pumila cv. Fuji) in Matrix-assisted Laser Desorption/Ionization Time-of-flight Mass Spectrometry and Fast-atom Bombardment Mass Spectrometry

Molecular size information for polymerized catechin larger than the decamer in unripe apple was obtained by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and by fast-atom bombardment mass spectrometry. Matrix-assisted laser desorption/ionization time of flight mass spectrometry provided evidence for the pentadecamer using trans-3-indoleacrylic acid as the matrix in the presence of silver ion. Even in the absence of silver ion, the dodecamer and undecamer were observed in the positive- and negative-ion modes, respectively. Fast-atom bombardment mass spectrometry also afforded evidence for the undecamer in both positive- and negative-ion modes.

Integrated Molecular and Biological Assays for Rapid Detection of Apple Scar Skin Viroid in Pear

A viroid in oriental pears (Pyrus hybrids) was associated with dappling and cracking fruit symptoms in graft-inoculated Malus pumila cv. Lord Lambourne. The viroid hybridized with a cRNA probe to apple scar skin viroid (ASSVd). Detection of the viroid by hybridization assays of tissue blots gave erratic results when used with some pear cultivars. Two methods for improving detection of the pear viroid were compared. In the first, pear bark chips were graft-inoculated into a viroid host, which was then tested by tissue blot hybridization. Digoxigenin-labeled cRNA probe and an antidigoxigenin antibody conjugate were used with a chemiluminescent substrate for detection of viroid in the tissue blots. Greenhouse-grown Pyrus communis cv. Nouveau Poiteau and Malus pumila cv. Virginia Crab amplified ASSVd to levels that were detectable in tissue blots of shoots and leaf petioles, even though the leaves and shoots were symptomless. In the second method, pears were indexed on the ASSVd indicator plants Stark's Earliest and Sugar Crab apple grown in growth chambers under a 24-h photoperiod. ASSVd-infected apple and the infected pears induced similar symptoms of leaf epinasty in the indicators in 5 to 8 weeks. Although infection in pear was rapidly and reliably detected by both methods, tissue blot hybridization was faster. Inoculation of an amplification host was useful when numerous cultivars or genetically diverse plants were tested and genetically similar positive and negative control plants were unavailable.

Acclimatization and subsequent gas exchange, water relations, survival and growth of microcultured apple plantlets after transplanting them in soil

Previous findings indicated that leaf conductance and net photosynthesis were positively correlated with water status (relative water content) of tissue‐cultured shoots and plantlets, and that acclimatization was associated with a reduction in leaf conductance and transpiration. In the present study, the effect of acclimatization on plant water status after plants were removed from culture and transplanted in soil, as well as the relation of water status with survival and growth after acclimatization were determined. Tissue‐cultured plantlets (Malus pumila cv. Greensleaves) were acclimatized by exposure to a 4‐day gradual reduction in humidity before they were transplanted. Once plants were transplanted, acclimatized plants maintained a higher relative water content than nonacclimatized plants, probably because of a lower initial stomatal conductance and less transpiration. High plant water status, as indicated by high relative water content, appeared to be an important factor for both plant survival and growth after the plants were transplanted. Higher water status was associated with higher growth rates and net assimilation rates. The differences in gas exchange, water relations and growth characteristics between plantlets in vitro and at 4 weeks after they were transplanted were also studied. Compared to in vitro plants, transplanted plants had a higher water status and higher leaf conductance and net photosynthesis. There was also an increase in the leaf area ratio (leaf area to plant biomass ratio) after they were transplanted which might have contributed to the higher net assimilation rate in transplanted compared to in vitro plants.

Immunochemical and immunohistochemical study of apple chlorogenic acid oxidase

Mouse antibody was raised against chlorogenic acid oxidase from apple (Malus pumila cv. Fuji). This antiserum was used for immunochemical characterization of the enzyme of apple and other plants and then for the immunocytolocalization of the enzyme in apple fruit to clarify the relationship between the distribution of the enzyme and browning. Western blot analysis revealed that the antibody recognized only the purified enzyme from apple fruit, with a molecular weight of 65 000. All of the apple cultivars showed the same band as the purified enzyme of Fuji. When the cross-reactivity of polyphenol oxidase between apple and other plants was examined, the antiserum only recognized the enzyme of pear. The browning of the apple fruit sections was predominantly observed around the core

Evaluation of Cutinase Activity of Various Industrial Lipases

Eight industrial lipases were tested for cutinase activity on apple (Malus pumila cv. Golden Delicious) cutin and compared with cutinase from the fungus Fusarium solani f. sp pisi. The analysis of released cutin monomers was performed by normal‐phase h.p.l.c with an evaporative light‐scattering detector. Calibration curves of cutin monomers were established for quantification of cutinolytic activity of the industrial lipases. The pH optimum for cutinase activity of the lipase preparations was approx. 8.0. A high concentration of detergent was necessary for expression of cutinase activity. The most active lipase was found to be Amano PS‐800, which was capable of hydrolysing 16% of apple cutin.

Purification and some properties of chlorogenic acid oxidase from apple (Malus pumila).

Chlorogenic acid oxidase was extensively purified to homogeneity from apple flesh (Malus pumila cv. Fuji). The enzyme was purified 470-fold, with a total yield close to 70% from the plastid fraction by ammonium sulfate precipitation, gel filtration and ion-exchange chromatography. The molecular weight was determined to be 65,000 by both SDS-PAGE and gel filtration chromatography. The optimum pH for the enzyme activity was around 4.0, and the enzyme was stable in the range of pH 6-8. The pI obtained by isoelectrofocusing was 5.4, and the N-terminal amino acid sequence was N-Asp-Pro-Leu-Ala-Pro-Pro-. The reaction rate of the purified enzyme was much larger for chlorogenic acid than for other o-diphenols such as (+)-catechin, (-)-epicatechin and 4-methylcatechol, and the enzyme lacked both cresolase activity and p-diphenol oxidase activity. The Km value for the enzyme was found to be 122 microM toward chlorogenic acid. The purified enzyme had far less thermal stability than the enzyme of the plastid fraction. Diethyl-dithiocarbamate, sodium azide, o-phenanthroline and sodium fluoride markedly inhibited the enzyme activity.

Endogenous gibberellin- and cytokinin-like substances in cultured shoot tissues of apple, Malus pumila cv. Jonathan, in relation to adventitious root formation

The frequencies of adventitious root formation in vitro of isolated shoots from bud cultures of apple (Malus pumila cv. Jonathan) after 1, 7 and 31 subcultures (weeks 5, 29 and 109 after the initial culture) were 5, 78 and 95% respectively. Endogenous gibberellin-like substances (GA) were extracted, chromatographed on SiO2 partition columns, and assayed on dwarf rice (Oryza sativa cv. Tan-ginbozu). The levels of GA in shoots from the 1st, 7th and 31st subcultures were 40, 19 and 14 ng GA3 eq./g dry weight of tissue, respectively, a trend which suggests an inverse relationship between endogenous GA level and rooting ability. This is consistent with the fact that applied GA3 inhibits rooting in apple and many other species. The major peak of GA activity eluted coincidentally with GA1/GA3/GA19. Endogenous cytokinin-like substances (CK) were chromatographed on paper and assayed with soybean hypocotyl sections. In contrast to the decrease in GA activity, CK activity increased 1.5–2.7 fold in the later subcultures (cytokinin activity per shoot, however, declined).