Metabolism of cell wall polysaccharides from persimmon fruit. Pectin solubilization during fruit ripening occurs in apparent absence of polygalacturonase activity

Pectins from persimmon ( Diospyros kaki L.) fruit pericarp were sequentially extracted with 0. 05 M trans -1,2-diaminocyclohexane-N,N, N', N'-tetraacetic acid (CDTA), 0. 05 M Na₂CO₃ (1°C) and Na₂CO₃ (20°C) and the carbohydrate composition and metabolism during development determined. Young persimmon fruits contained a large proportion of pectins, 46% by dry weight, that decreased to 20% with ripening. This decrease occurred in the CDTA and Na₂CO₃ (1°C) fractions, mainly composed of uronic acids, and represents a net loss of uronic acids, arabinose and galactose. The amount of non-cellulosic neutral sugars was especially high in the Na₂CO₃ (20°C) fraction. The loss of pectins was also accompanied by a depolymerisation of the polysaccharides extracted in the three pectic fractions. However, none of these changes can be attributed to the action of polygalacturonase activity. Proteins were extracted from the pericarp tissue, but endopolygalacturonase (EC 3. 2. 1. 15) activity, determined as a decrease in viscosity of polygalacturonic acid, was not observed in the extract. Determination of exopolygalacturonase (EC 3. 2. 1. 67) activity by measuring the release of reducing groups from polygalacturonic acid was also negative. The results presented indicate that polygalacturonase is not responsible for the metabolism of pectins during persimmon fruit ripening.     

Implication of persimmon fruit hemicellulose metabolism in the softening process. Importance of xyloglucan endotransglycosylase

Hemicellulosic polysaccharides from persimmon fruit ( Diospyros kaki L.) pericarp were extracted from depectinated cell walls with 0.5, 1 and 4 M KOH at different stages of development: (I) maximal growth corresponding to the first sigmoidal growth phase; (II) cessation of growth corresponding to the lag between the first and the second sigmoidal phases; (III) maximal growth corresponding to the second sigmoidal phase; and (IV) cessation of growth when the fruit had reached its maximum size and the change in colour (green to red) had taken place. During fruit development the amount of total hemicelluloses per unit dry mass cell wall decreased twofold. Xyloglucan was present in the three hemicellulosic fractions, and also decreased with fruit age, although its amount relative to other hemicelluloses increased. The amount of xyloglucan was especially high in the hemicelluloses extracted with 4 M KOH, representing more than 50% at stages III and IV. The average molecular mass of xyloglucan increased from stage I through stage II (0.5 M hemicellulosic fraction) or through stage III (I and 4 M hemicellulosic fractions) and decreased after that. The xyloglucan endotransglycosylase (XET: EC 2.4.1.-) activity was measured as the incorporation of [³H]XXXGol (reduced xyloglucan heptasaccharide labelled at position 1 of the glucitol moiety) into partially purified persimmon fruit xyloglucan. XET specific activity increased greatly between stages I and II. The importance of this enzyme during fruit ripening is discussed.     

Climatic factors affecting the occurrence of cloudy stain on the fruit skin of ‘Triumph’ Japanese persimmon

Cloudy stain is a physiological disorder of the Japanese persimmon blemishing the skin and causing a great reduction of fruit quality, thus decreasing its commercial value. The stain starts at or just prior to the fruit colour-break stage, coinciding with the presence of cuticular cracks which increase cuticle water permeability and expose subepidermal cells to air and water, causing oxidation of polyphenol substances of large skin areas that turn almost black. The incidence of the disorder examined over four consecutive years in two growing areas of Spain revealed significant differences from year to year, between areas and even the tree face examined. The data consistently showed that cooler groves and fruit located on the northern face of the canopy were more prone to develop the stain. During the 4 years of the study, the average daily minimum temperature of a period of 15–25 days prior to the onset of staining followed the same order as the yearly staining rate, and accumulated hours below 13°C correlated significantly with the percentage of fruit starting the disorder. No relationship was found between the time-course of relative humidity (RH) and the onset of staining, but once the stain started the lower RH resulted in the lower percentage of affected fruit. Dew point, wind speed, rainfall and evapotranspiration did not correlate either with the onset of staining or with its incidence. Rind of cloudy-stained fruit had higher concentration of N, K, Mg and Fe and lower concentration of Ca than that of healthy fruit, but it is not the cause of the disorder. Accordingly, we conclude that cloudy stain of Japanese persimmon starts after a period of low temperature (≤ 13°C) and, afterwards, RH is responsible for its development and severity.     

低乙烯减压处理对柿果实乙烯生物合成的影响

在 1 0℃± 1℃的条件下 ,研究了低乙烯减压处理对火柿采后乙烯生物合成的影响。结果表明 ,低乙烯减压处理使火柿 ACC含量降低 ,EFE活性减弱 ,从而显著抑制了乙烯的产生 ,延迟了乙烯峰出现的时间。该处理能显著延缓火柿硬度的下降。     

Preferential transport activity of DkDTX5/MATE5 affects the formation of different astringency in persimmon

Proanthocyanidins(PAs) are specialized metabolites that infuence persimmon fruit quality.Normal astringent(A)-type and non-astringent(NA)-type mutants show significant variation in PA accumulation, but the infuencing mechanism remains unclear. In this study, among the six identified DTXs/MATEs proteins associated with PA accumulation, we observed that allelic variation and preferential transport by Dk DTX5/MATE5 induced variation in PA accumulation for A-type and NA-type fruit. The expression pa...     

Development of microsatellite markers in polyploid persimmon (Diospyros kaki Lf) from an enriched genomic library

The oriental persimmon (Diospyros kaki Lf) is believed to have originated in China with subsequent introduction into Japan and Korea in ancient times. The species was then brought to Europe, Brazil and the USA from Japan in the 19th century. Recent studies highlighted the poor state of identification of cultivars in these countries due to incorrect labelling and presence of synonyms among local varieties. Thus, molecular marker characterization of germplasm resources is of great value for genetic resource preservation and plant breeding of persimmon. Therefore, to identify accessions for further plant breeding and germplasm management, 37 microsatellite loci were developed from a CT/AG‐enriched persimmon genomic library.     

Control and yield loss modelling of circular leaf spot of persimmon caused by Mycosphaerella nawae

Symptoms of circular leaf spot of persimmon (CLSP), caused by Mycosphaerella nawae, consisted of necrotic spots on leaves, chlorosis and premature defoliation. Although CLSP is a foliar disease, early fruit maturation and abscission are frequently associated with the presence of lesions on leaves and defoliation, resulting in severe economic losses. Despite their importance for the design of efficient disease management programmes, quantitative relationships between CLSP incidence and yield loss are unknown. Therefore, fungicide efficacy trials were conducted during two consecutive years in Spain to induce different levels of disease severity, defoliation and yield loss. The effects of fungicide treatments on CLSP severity were analysed by ordinal logistic regression models. Relative yield loss values were regressed against the percentage of affected leaves or defoliated obtained at different evaluation dates. The disease had high negative impact and complete yield loss was observed in the absence of effective fungicide treatments. Preventive applications of pyraclostrobin, trifloxy‐strobin and mancozeb provided the best disease control and highest yields, up to 95.77 kg tree^?1. An exponential relationship of CLSP incidence and defoliation with yield loss was found. In general, model fit and predictive ability was superior when defoliation, rather than incidence, was used as explanatory variable. The impact of defoliation on yield loss was higher in earliest evaluation dates, suggesting that early leaf abscission may be the main factor contributing to premature fruit drop and subsequent yield loss. Substantial yield losses were observed even with relatively low levels of CLSP incidence and defoliation. Therefore, it was not possible to define a critical action threshold for CLSP management based on foliar symptoms.     

外源抗坏血酸处理对人工脱涩柿果的保鲜作用

该研究以广西中部地区的早熟品种‘牛心柿’(Diospyros kaki cv.Niuxin)的果实为材料,研究果实萼片施用5%抗坏血酸溶液处理对CO_2脱涩柿果采后贮藏期果实软化和细胞壁降解以及抗氧化体系等相关生理生化指标的影响。结果表明:抗坏血酸对人工脱涩后‘牛心柿’果实的保鲜效果明显,与空白和去离子水对照相比,外源抗坏血酸处理更好地延缓柿果实硬度的降低和总色差的升高。抗坏血酸处理后7~15 d,原果胶的下降速率和可溶性果胶的上升速率得以延缓,细胞壁降解酶PG和Cx活性均低于空白和对照。同时,外源抗坏血酸阻滞了整个贮藏期果实内源抗坏血酸的减少,并抑制了POD的活性和MDA的积累,果实自身抗氧化体系功能受到保护。此外,抗坏血酸处理还在一定程度上提高了果实还原糖的含量。因此,采后外源施用抗坏血酸不仅可以延长脱涩后鲜食柿果的货架期,而且还有助于提高果实的商品品质。     

广西柿种质资源果实性状多样性分析与模糊综合评价

为进一步揭示广西柿种质资源信息,运用多样性分析和模糊综合评价法对不同生态气候区的柿种质资源果实性状进行研究和评价。结果表明:不同柿种质的果实性状存在丰富的多样性。86份柿种质的模糊综合评价指数FCI值在1.849～3.947之间,平均2.835。根据FCI取值范围将所调查的柿种质分成5个等级,FCI值最高的第5级种质数量最少,占所调查种质的3.49%;FCI值最低的第1级种质数量最多,占所调查种质的48.84%。月柿、牛心柿、大方柿、京柿、鸡心柿、小方柿和火柿等柿品种及一些实生单株的模糊综合评价指数较高,它们的果实综合性状表现较优,建议作为优良品种和单株加以开发和利用。部分单个性状比较优良的实生单株虽不适合于直接推广种植,但可作为柿品种改良的资源加以利用。     

Hypoxia‐responsive ERFs involved in postdeastringency softening of persimmon fruit

Removal of astringency by endogenously formed acetaldehyde, achieved by postharvest anaerobic treatment, is of critical importance for many types of persimmon fruit. Although an anaerobic environment accelerates de‐astringency, it also has the deleterious effect of promoting excessive softening, reducing shelf life and marketability. Some hypoxia‐responsive ethylene response factors (ERFs) participate in anaerobic de‐astringency, but their role in accelerated softening was unclear. Undesirable rapid softening induced by high CO₂ (95%) was ameliorated by adding the ethylene inhibitor 1‐MCP (1 μL/L), resulting in reduced astringency while maintaining firmness, suggesting that CO₂‐induced softening involves ethylene signalling. Among the hypoxia‐responsive genes, expression of eight involved in fruit cell wall metabolism (Dkβ‐gal1/4, DkEGase1, DkPE1/2, DkPG1, DkXTH9/10) and three ethylene response factor genes (DkERF8/16/19) showed significant correlations with postdeastringency fruit softening. Dual‐luciferase assay indicated that DkERF8/16/19 could trans‐activate the DkXTH9 promoter and this interaction was abolished by a mutation introduced into the C‐repeat/dehydration‐responsive element of the DkXTH9 promoter, supporting the conclusion that these DkERFs bind directly to the DkXTH9 promoter and regulate this gene, which encodes an important cell wall metabolism enzyme. Some hypoxia‐responsive ERF genes are involved in deastringency and softening, and this linkage was uncoupled by 1‐MCP. Fruit of the Japanese cultivar ‘Tonewase’ provide a model for altered anaerobic response, as they lost astringency yet maintained firmness after CO₂ treatment without 1‐MCP and changes in cell wall enzymes and ERFs did not occur.     

基于网络药理学的柿果药理功能定位及作用机制

基于网络药理学,通过国内外文献检索获取柿果中的化合物,采用Swiss target prediction数据库对化合物进行潜在靶点垂钓以探讨柿果的药理功能定位及作用机制。以Cytoscape软件构建化合物-靶点网络,靶点-疾病名称-疾病分类网络,同时对靶点进行蛋白相互作用(PPI)网络构建,采用DAVID数据库对靶点进行通路富集分析。本研究共收集到柿果中16个化合物,可作用于68个靶点,这些靶点主要作用于心血管疾病、神经精神性疾病等。PPI网络图包含84个节点,226条边,其中degree值排前10的关键蛋白分别为ERS1、PGS2、MMP2、TIMP1、MMP9、MMP1、AR、SLC6A3、PRKCB、CYP19A1。上述靶点可调节氮素代谢、血清素能突触以及TRP通道炎症介质的调节等信号通路。本研究为阐明柿果的药理功能定位及其作用机制研究提供了可靠的依据。     

柿中具有△^12熊果烯骨架的五环三萜类化合物

柿（Diospyros kaki L.f.）的果实中分离出5个化合物,通过IR、MS、^1HNMR、^13CNMR鉴定,确定为：熊果辛酯（octyl yursolate）,△^5,6－3－（2′－甲在戊酰氧在）－熊果酸环己酯〔△^5,6－3－（2′－methyl pentanoyl）-ursolic acid cyclohexy ester〕,β－香树脂醇己醚（β－amyrin hexyl ether）,△^6,7－香树素（△^6,7－amyin）和熊果甲酯（methyl ursolate）。     

Immature Persimmon Suppresses Amyloid Beta (Aβ) Mediated Cognitive Dysfunction via Tau Pathology in ICR Mice

This study confirmed the ameliorating effect of immature persimmon (Diospyros kaki) ethanolic extract (IPEE) on neuronal cytotoxicity in amyloid beta (Aβ)_1–42-induced ICR mice. The administration of IPEE ameliorated the cognitive dysfunction in Aβ_1–42-induced mice by improving the spatial working memory, the short-term and long-term memory functions. IPEE protected the cerebral cholinergic system, such as the acetylcholine (ACh) level and acetylcholinesterase (AChE) activity, and antioxidant system, such as the superoxide dismutase (SOD), reduced glutathione (GSH) and malondialdehyde (MDA) contents. In addition, mitochondrial dysfunction against Aβ_1–42-induced toxicity was reduced by regulating the reactive oxygen species (ROS), mitochondrial membrane potential and ATP contents. In addition, IPEE regulated the expression levels of tau signaling, such as TNF-α, p-JNK, p-Akt, p-GSK3β, p-tau, p-NF-κB, BAX and caspase 3. Finally, gallic acid, ellagic acid and quercetin 3-O-(6″-acetyl-glucoside) were identified as the physiological compounds of IPEE using ultra-performance liquid chromatography ion mobility separation quadrupole time-of-flight/tandem mass spectrometry (UPLC IMS Q-TOF/MS²).     

基于叶绿体DNA ndhA和nrDNA ITS序列变异分析栽培柿及其近缘种亲缘关系

以柿属植物（Diospyros spp.）中与柿近缘的8种共30个基因型为试材,进行核糖体DNA(nrDNA)内转录间隔区（ITS）和叶绿体DNA ndhA序列变异分析,并通过软件计算两个序列及合并后的进化模型,依据进化模型采用ML法(maximum likelihood method)分析进化关系。为进一步弄清柿属植物种间亲缘关系和供试柿（Diospyros kaki Thunb.）种内分子差异提供了理论依据。结果表明：（1）ndhA序列长度变异范围在1 492~1 511,14个信息位点;ITS序列长度变异范围在660~761,56个信息位点。ITS、ndhA和ndhA+ITS（ndhA和ITS合并）最适碱基进化模型分别为（TrN+I+G）、（F81+I）和（GTR+I+G）。综合ITS和ndhA序列分析表明：柿与油柿和云南野毛柿亲缘关系最近,与美洲柿和乌柿最远。（2）21份柿品种材料的ITS长度均为730,包括4个变异位点,据此4个变异位点对供试柿种内21个品种进行聚类分析。研究认为,ndhA和ITS能较清楚解释了柿与其近缘种间的亲缘关系,并通过柿品种ITS的差异位点分析鉴别出栽培柿种内的差异。     

The bumblebee Bombus ardens ardens (Hymenoptera: Apidae) visits white clover in orchards before Oriental persimmon blooms

Flowers on the ground of orchards can provide substantial resources for wild pollinators of orchard trees. Few studies, however, have examined the relative importance of groundcover flowers to orchard pollination by analyzing pollen on the body surface of pollinators. Oriental persimmon trees bloom within the longer blooming period of white clover, which is occasionally found as a flowering plant on the ground of persimmon orchards in Japan. The present study compared the insect species assemblage collected on persimmon flowers with that on clover. Before persimmon bloomed, Bombus ardens ardens and Apis cerana japonica were the major visitors of clover flowers. Once persimmon bloomed, the former was the most abundant bee that visited persimmon flowers over the flowering period. Apis mellifera was captured only on clover flowers. We found numerous clover pollen grains on the body surface of bumblebees captured on persimmon flowers, but far fewer persimmon pollen grains on bees that visited clover. These findings show that B. ardens ardens utilized the clover flowers under the orchards before persimmon bloomed.     

涩柿生长,脱涩及返涩单宁细胞形态的变化

以甜柿为对照,观察了涩柿（牛心柿）、脱涩和返涩中单宁细胞的形态变化,结果表明：涩柿单宁细胞在生长期体积不断增大,数量不断增多,涩味加强,在９月初细胞停止生长;４种不同方法脱涩的柿,其单宁细胞的变化基本相同,都是体积缩小,最后变成疏散的小颗粒;返涩的柿,单宁从单宁细胞中溢出,呈网状结构分布于果肉组织中。     

柿果实ACC合成酶cDNA的克隆及其序列分析

根据其它植物ACC合成酶（1－aminocyclopropane-1-carboxylic acid synthase,ACS)氨基酸保守区,设计1组简并引物,用RT－PCR法,从柿（Diospyros kaki Thunb.)果实扩增出3个约1kb左右的cDNA片段,将其克隆至pGEM-T载体上,对这些重组克隆进行序列测定和氨基酸序列推导,DK－ACS1由1101个碱基组成,编码364个氨基酸;DK－ACS2是1086个碱基,编码359个氨基酸;DK－ACS3为1089个碱基,编码363个氨基酸。它们均具有其它植物ACS合成酶中存在的7个保守区和11个不变氨基酸残基,且在多肽水平上有较高的同源性。与番茄LE－ACS2的同源性DK－ACS1是60.5A%,DK-ACS2是70.7%,DK－ACS3为66．9％,与甜瓜CM－ACS1的同源性依次分别是60．4％,72．1％和64．4％。     

A role for jasmonates in climacteric fruit ripening

Jasmonates are a class of oxylipins that induce a wide variety of higher-plant responses. To determine if jasmonates play a role in the regulation of climacteric fruit ripening, the effects of exogenous jasmonates on ethylene biosynthesis and color, as well as the endogenous concentrations of jasmonates were determined during the onset of ripening of apple (Malus domestica Borkh. cv. Golden Delicious) and tomato (Lycopersicon esculentum Mill. cv. Cobra) fruit. Transient (12 h) treatment of pre-climacteric fruit discs with exogenous jasmonates at low concentration (1 or 10 μM) promoted ethylene biosynthesis and color change in a concentration-dependent fashion. Activities of both 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase and ACC synthase were stimulated by jasmonate treatments in this concentration range. The endogenous concentration of jasmonates increased transiently prior to the climacteric increase in ethylene biosynthesis during the onset of ripening of both apple and tomato fruit. The onset of tomato fruit ripening was also preceded by an increase in the percentage of the cis-isomer of jasmonic acid. Inhibition of ethylene action by diazocyclopentadiene negated the jasmonate-induced stimulation of ethylene biosynthesis, indicating jasmonates act at least in part via ethylene action. These results suggest jasmonates may play a role together with ethylene in regulating the early steps of climacteric fruit ripening. Received: 14 August 1997 / Accepted: 4 October 1997