滇山茶狮子头及其芽变品种大玛瑙间花色差异分析

滇山茶是世界著名观赏花木,花色是其重要的观赏性状。滇山茶狮子头花色为深红色,而其芽变品种大玛瑙是滇山茶中唯一红白双色的名贵品种,极具观赏价值。以上述2个品种为研究材料,采用RHSCC比色卡比色法和色差仪测定2个品种滇山茶花蕾期和盛花期花瓣的花色表型,并基于转录组与代谢组分析挖掘呈色相关的关键代谢物及关键基因。花青素靶向代谢分析表明,在滇山茶2个品种中共鉴定出28种花青素代谢物,其中狮子头与大玛瑙红色区域花瓣间没有差异代谢物,狮子头与大玛瑙白色区域花瓣间的关键差异代谢物为矢车菊素-3-O-桑布双糖苷、原花青素B2、原花青素B3、阿福豆苷,大玛瑙花瓣的红白区域关键差异代谢物为矢车菊素-3-O-桑布双糖苷、原花青素B2、阿福豆苷。转录组KEGG分析结果表明,苯丙醇生物合成和类黄酮生物合成途径与大玛瑙红白双色花瓣的形成有关;植物激素信号转导和昼夜节律-植物途径与滇山茶花色芽变有关。转录代谢联合分析共筛选出与滇山茶呈色高度相关的差异表达基因共17条,包括4条CHS、3条HCT、2条F3′H、1条LAR、5条MYB和2条b HLH。本研究结果对进一步揭示花色芽变育种具有一定的参考意义。     

菊花不同花色品种中花青素苷代谢分析

应用高效液相色谱和多级质谱联用技术（HPLC-ESI-MSn）,分析菊花（Chrysanthemum×morifolium）白色、粉色、红色、紫色、红紫色和墨色6个色系共计82个品种中花青素苷合成过程的中间产物和最终产物,发现从白色、粉色、红色、紫色、红紫色到墨色花青素苷含量快速增加,分别为4.68、111.60、366.89、543.56、1220.36和2674.95μg·g-1,不同色系间花青素苷的含量差异显著（P〈0.01）,花青素苷含量越高花色越深;墨色菊花品种中总类黄酮含量显著高于其它花色品种（P〈0.01）,其它不同色系间总类黄酮含量差异不显著（P〉0.05）;随着菊花花色变深,从柚皮素分支到圣草酚的代谢流,以及从圣草酚分支到矢车菊素苷的代谢流比例增加。花青素苷成分分析发现：菊花中只含有矢车菊素苷类化合物。根据花青素苷代谢成分分析结果绘制了菊花中花青素苷代谢路径图,即在菊花类黄酮代谢途径中只存在矢车菊素苷代谢分支途径;菊花不同色系在柚皮素和圣草酚2个关键代谢分支点上向不同方向代谢流的分配比例不同,造成花青素苷产物含量不同,导致不同花色。以上研究结果为菊花花色改良的分子育种提供了理论依据。     

菊花不同花色品种中花青素苷代谢分析

应用高效液相色谱和多级质谱联用技术(HPLC-ESI-MSⁿ), 分析菊花(Chrysanthemum × morifolium)白色、粉色、红色、紫色、红紫色和墨色6个色系共计82个品种中花青素苷合成过程的中间产物和最终产物, 发现从白色、粉色、红色、紫色、红紫色到墨色花青素苷含量快速增加, 分别为4.68、111.60、366.89、543.56、1 220.36和2 674.95 μg·g^–1, 不同色系间花青素苷的含量差异显著(P<0.01), 花青素苷含量越高花色越深; 墨色菊花品种中总类黄酮含量显著高于其它花色品种(P<0.01), 其它不同色系间总类黄酮含量差异不显著(P>0.05); 随着菊花花色变深, 从柚皮素分支到圣草酚的代谢流, 以及从圣草酚分支到矢车菊素苷的代谢流比例增加。花青素苷成分分析发现: 菊花中只含有矢车菊素苷类化合物。根据花青素苷代谢成分分析结果绘制了菊花中花青素苷代谢路径图, 即在菊花类黄酮代谢途径中只存在矢车菊素苷代谢分支途径;菊花不同色系在柚皮素和圣草酚2个关键代谢分支点上向不同方向代谢流的分配比例不同, 造成花青素苷产物含量不同,导致不同花色。以上研究结果为菊花花色改良的分子育种提供了理论依据。     

基于LC-MS/MS分析马缨杜鹃花代谢物的变化

为分析马缨杜鹃(Rhododendron delavayi)花开花至凋谢过程中的代谢产物差异及其通路,该文采用LC-MS/MS技术对其花苞期、开裂期、传粉期、盛开期、衰老期和凋谢期的化学成分进行非靶向代谢组学分析。结果表明:(1)共鉴定到973种代谢物,主要包含黄酮类、有机酸、酚酸类、氨基酸及其衍生物、脂类、生物碱等。(2)主成分分析(PCA)表明样本间代谢物存在差异,结合正交偏最小二乘判别分析(OPLS-DA)、t检验的P值和单变量分析的差异倍数(fold-change)筛选差异代谢物(VIP>1,P<0.05,Fc>2或Fc<0.5),涉及591种,在马缨杜鹃花期进入衰老期和凋谢期后差异代谢物数量和表达量显著上升,其中花苞期至开裂期差异代谢物的表达主要呈现下调,而进入衰老期和凋谢期后差异代谢物的表达主要呈现上调。(3)KEGG注释到68条代谢通路,其中差异代谢物极显著富集(P < 0.01)通路3条,包括苯丙素类生物合成、植物激素的生物合成和类黄酮生物合成。(4)结合苯丙素类、黄酮类等有效成分生物合成通路共筛选到10种代谢物包括苯丙氨酸(L-phenylalanine)、反式肉桂酸(trans-cinnamic acid)、查耳酮(chalcone)、柚皮素(naringenin)、对香豆酰基莽草酸(p-coumaroyl shikimic acid)、阿魏酸(ferulic acid)、松柏醇(coniferyl alcohol)、芥子酸(sinapic acid)、紫丁香苷(syringin)、槲皮素(quercetin)。此外,有效成分的差异代谢物表明苯丙素类生物合成代谢活动随马缨杜鹃花的发育逐渐增强,而黄酮类化合物生物合成逐渐减弱,这些关键差异代谢物可能对马缨杜鹃花的发育有重要的调控作用。该研究为马缨杜鹃花开花至凋谢进程中的有效成分代谢途径活性物质的研究提供了代谢组学基础,为进一步研究马缨杜鹃花花期调控的分子机理提供参考。     

重金属复合污染对眼优角蚱代谢组的影响

【目的】对金属矿区和非金属矿区环境中生活的眼优角蚱Eucriotettix oculatus体内重金属含量及其代谢组进行分析,探究重金属复合污染对眼优角蚱体内重金属累积和代谢组的影响。【方法】使用ICP-MS法对眼优角蚱成虫体内重金属含量进行测定,同时采用基于 UPLC-MS/MS检测平台、自建数据库以及多元统计分析相结合的手段,对金属矿区和非金属矿区眼优角蚱成虫肠道中的代谢物进行差异分析,并利用 KEGG 数据库对差异代谢物进行注释以及通路富集分析。【结果】金属矿区眼优角蚱成虫体内9种重金属含量是非金属矿区的0.4~212.4倍。多元统计分析结果表明,金属矿区眼优角蚱成虫肠道中共有112种代谢物的含量发生了显著变化,主要为氨基酸类、脂肪酰类、有机酸类、核苷酸类、苯类等物质。KEGG注释及通路富集分析显示,可被注释到的显著差异代谢物共有49种,与代谢通路相关的显著差异代谢物有40种,富集最显著的通路是甲状腺激素合成通路、催产素信号通路、胆汁分泌通路、酪氨酸代谢通路。【结论】重金属复合污染环境中生存的眼优角蚱成虫体内有多类重金属累积,重金属可改变眼优角蚱肠道中的代谢物的组成,其中一些代谢物的改变可能是眼优角蚱适应重金属复合污染生境的策略。     

基于UPLC-MS/MS技术的野生及栽培韭菜籽的代谢组学研究

多星韭为贵州省赫章县喀斯特地貌区重要的野生植物资源之一,具有较高的开发利用价值。为分析野生多星韭(Allium wallichii )籽与栽培韭菜(A. tuberosum)籽代谢产物差异及其通路,该研究利用UPLC-MS/MS物质分离鉴定技术,对2种韭籽化学成分进行广泛靶向代谢组学分析。结果表明:(1)共检测到782种代谢产物。(2)主成分分析(PCA)显示样本间存在差异,正交偏最小二乘判别分析(OPLS-DA)共筛选出12类显著变化(P<0.05,VIP≥1)的差异代谢物,涉及492种,其中上调和下调幅度在前20的代谢物包括黄酮、甾体皂苷、黄酮醇、酚酸类、异黄酮、游离脂肪酸、三萜皂苷、生物碱、吲哚类生物碱、氨基酸及其衍生物等。(3)KEGG注释到84条差异代谢通路,其中差异代谢物显著富集(P<0.01)通路4条,此外还构建了未注释到的显著差异代谢物甾体皂苷的生物合成通路。该研究结果为韭籽有效成分代谢途径解析及药理活性物质研究提供了参考,也为赫章县野生多星韭的开发保护与多元化利用提供了新思路。     

基于非靶代谢组学多刺绿绒蒿不同器官代谢物差异分析

为了进一步探究传统藏药植物多刺绿绒蒿（Meconopsis horridula）中代谢物成分以及不同器官差异情况,采用UPLC-MS技术对多刺绿绒蒿的叶、根和花三个不同器官代谢物进行分析与鉴定。并利用主成分分析（PCA）、聚类热图分析、正交偏最小二乘-判别分析（OPLS-DA）和KEGG通路富集分析等方法进行不同器官差异代谢产物筛选与通路分析。结果显示,在ESI⁺和ESI^-模式下,共检测注释到947种代谢物,不同器官间差异代谢物进行分析,叶和根差异代谢物有301个,叶和花中差异代谢物有170个,根和花中差异代谢物有244个。通过聚类热图可以看出,大多数代谢物在根中含量较低;KEGG通路富集分析显示,差异代谢物大多富集在氨基酸代谢、花青素生物合成、黄酮类生物合成和生物碱合成等代谢途径。各器官优势黄酮类、萜类和生物碱类代谢物的分析为进一步探究多刺绿绒蒿的不同器官药用特征成分和开发利用提供一定的帮助。     

半枫荷调控RA模型的非靶向代谢组学研究

为探讨半枫荷干预类风湿性关节炎(rheumatoid arthritis, RA)模型大鼠血浆内容物代谢轮廓的变化和特征,该研究以半枫荷正丁醇提取物给药前后RA模型大鼠血浆为研究对象,借助超高效液相色谱联用四极杆飞行时间质谱(UPLC-QTOF/MS)技术进行非靶向代谢组学检测,并用SIMCA-P软件对代谢物测定结果进行多元变量统计分析,筛选差异代谢物并作通路富集分析。结果表明:(1)给药前后大鼠血浆代谢轮廓存在显著差异,与模型组相比,给药组在正负离子模式合并后筛选出321种差异代谢物,其中负离子模式鉴定到174种代谢物,正离子模式鉴定到192种代谢物。(2)鉴定到的所有代谢物根据其化学分类归属信息归为12种类型,有机酸及其衍生物和脂类及类脂分子这2类代谢物数量占比较高。(3)通路富集获得37个代谢通路且呈显著性差异(P<0.05),给药组中蛋白质的消化和吸收、肿瘤胆碱代谢通路和ABC转运蛋白通路出现较大扰动且富集到的差异代谢物数量最多,所有通路均显著上调(P<0.05)。这对阐明半枫荷调控RA症状的变化机制具有一定指导价值和理论意义。     

基于非靶代谢组学的糙皮侧耳子实体发育的关联物质及主要代谢通路分析

本研究采用代谢组学方法鉴定并分析了糙皮侧耳原基和刚分化的子实体中的小分子代谢物质,以期解析糙皮侧耳子实体形成和发育的潜在机制。糙皮侧耳原基和子实体中共鉴定出545种代谢物,包含酚酸、脂类、氨基酸及其衍生物、核苷酸及其衍生物、有机酸、生物碱、单宁、木脂素和香豆素以及其他代谢物共9类。主成分分析(PCA)和正交偏最小二乘判别分析(OPLS-DA)表明糙皮侧耳原基和子实体中的代谢物质具有显著差异。以VIP≥1、fold change≥2或≤0.5为条件,共筛选到253种差异代谢物。调控通路分析结果显示差异物质涉及76条代谢途径,推测该菌通过色氨酸代谢,苯丙氨酸、酪氨酸和色氨酸的生物合成,赖氨酸降解,D-精氨酸和D-鸟氨酸代谢,生物素代谢,莨菪烷、哌啶和吡啶生物碱的生物合成,酪氨酸代谢,嘌呤代谢,丙酸代谢和异喹啉生物碱的生物合成等主要代谢通路完成物质的深度转化及调控。本研究为糙皮侧耳子实体发育机制的研究提供了一定的理论依据。     

铁筷子花瓣中花青素苷成分及含量对其呈色的影响北大核心CSCD

该研究以7个品种铁筷子(Helleborus thibetanus Franch.)为试验材料,借助目视测色、RHSCC比色卡、色差仪进行花色表型的测定,采用高效液相色谱法-光电二极管阵列检测方法(HPLC-DAD)及高效液相色谱-电喷雾离子化-质谱联用技术(HPLC-ESI-MS)测定分析铁筷子花瓣中花青素苷成分及含量,以探究不同品种铁筷子的花色与花青素苷成分及含量之间的关系。结果显示:(1)紫色系品种花瓣的a*值最高b*值最低,黄色系品种花瓣的b*值最高a*值最低,不同品种的铁筷子花色越深L*值越低。(2)从5个有花青素苷积累的铁筷子品种中检测出11种花青素苷成分,分别为6种矢车菊素苷,4种飞燕草素苷,1种矮牵牛素苷;供试的铁筷子材料中红色系2个品种的花青素苷含量最高,紫色系品种次之;矢车菊素苷与飞燕草素苷为影响铁筷子花瓣呈色的主要色素物质。(3)不同种类的花青素和修饰基团的差异,导致铁筷子花瓣呈现不同的色彩,含有多种酰基化修饰的飞燕草素苷使铁筷子花色蓝移进而使花色加深。(4)相关分析表明,铁筷子花瓣的L*值与a*值呈显著负相关关系,与b*值呈显著的正相关关系;L*值与总花青素苷含量呈显著负相关关系,且随着花青素苷含量的累积a*值增加,花色红移。研究表明,花青素苷的成分及含量是导致铁筷子花瓣呈现不同颜色的主要原因,矢车菊素苷和飞燕草素苷的互作以及酰基化的修饰使铁筷子呈现不同程度的紫色,花青素苷的不同累积量影响了花瓣颜色的明暗变化,从而使铁筷子花瓣颜色丰富。     

Genetic engineering of the anthocyanin biosynthetic pathway with flavonoid-3′,5′-hydroxylase: specific switching of the pathway in petunia

Flavonoid-3',5'-hydroxylase (F3'5'H) is the key enzyme in the synthesis of 3',5'-hydroxylated anthocyanins, which are generally required for the expression of blue or purple flower color. It has been predicted that the introduction of this enzyme into a plant species that lacks it would enable the production of blue or purple flowers by altering the anthocyanin composition. We present here the results of the genetic engineering of petunia flower color, pigmentation patterns and anthocyanin composition with sense or antisense constructs of the F3'5'H gene under the control of the CaMV 35S promoter. When sense constructs were introduced into pink flower varieties that are deficient in the enzyme, transgenic plants showed flower color changes from pink to magenta along with changes in anthocyanin composition. Some transgenic plants showed novel pigmentation patterns, e.g. a star-shaped pattern. When sense constructs were introduced into blue flower petunia varieties, the flower color of the transgenic plants changed from deep blue to pale blue or even pale pink. Pigment composition analysis of the transgenic plants suggested that the F3'5'H transgene not only created or inhibited the biosynthetic pathway to 3',5'-hydroxylated anthocyanins but switched the pathway to 3',5'-hydroxylated or 3'-hydroxylated anthocyanins.     

Characterisation of flower colouration in 30 Rhododendron species via anthocyanin and flavonol identification and quantitative traits

• Floral colour is a key reproductive character, often associated with environmental adaptation, and subject to human intervention. A large number of Rhododendron species differ widely in flower colour, providing a good model for flower colouration. The chromatic features and anthocyanin compositions of 30 species from seven subgenera were systematically analysed.
• The Royal Horticultural Society Colour Chart and CIE L*a*b* system were employed to describe and investigate flower colours. The UPLC‐PDA/ESI‐MSⁿ system was used to identify and quantify anthocyanins in petal extracts.
• The flower colours of 30 Rhododendron species were categorised into four groups – red, purplish pink, purple and white. Seven anthocyanins were identified and quantified in petals: delphinidin, cyanidin and malvidin 3‐O‐arabinoside‐5‐O‐glucosides, cyanidin 3,5‐di‐O‐glucoside, 3‐O‐galactoside and 3‐O‐arabinoside, and delphinidin 3‐O‐glucoside. The red‐flowered species mainly contained cyanidin monoglycosides and had much higher total anthocyanin content than purplish pink‐ and purple‐flowered species. Purplish pink‐ and purple‐flowered species had similar anthocyanin types and content. The chromatic differences were significant among groups, except the purplish pink and purple groups. Statistical analysis showed that Cy3Gal and Cy3Arb are characteristic for red‐flowered species, and Mv3Arb5G and Dp3Arb5G play important roles in purple colouration; their contents were major components that greatly affected the chromatic parameters. In total, 21 flavonol derivates were identified. However, total flavonol content and co‐pigmentation index showed no significant difference or correlation among/with colour groups, suggesting that flavonols might not play a major role in colouration.
• These results enhance our knowledge of the biochemical basis of flower colouration in Rhododendron species, and provide a foundation for genetic variation studies and aid in breeding cultivars with novel flower colours.

     

RELATION OF FLOWER COLOR TO OPTICAL-DENSITY SPECTRA OF INTACT TISSUE AND OF ANTHOCYANIN EXTRACTS

Data from measurements of optical density of intact tissue and of anthocyanins in extracts resolved on cellulose thin layer plates were compared with visual evaluations of color quality and intensity in poinsettia, rose, and snapdragon. Visual evaluation was in good agreement with both instrumental and chemical determinations. However, the number or kinds of anthocyanins present could not be predicted from the visual evaluation or from the spectra of the fresh tissue. Data from the resolved extracts did not provide a basis for predicting the optical-density spectrum or the color of the intact tissue. In addition to the genetic factors which have been shown to control (1) the type of anthocyanin, (2) the amount of anthocyanin, and (3) the distribution of anthocyanins within the flower, we suggest another group of genes which apparently affect color through control of structural modification of individual anthocyanins in the living cell through shift in pH, metal chelation, and/or copigmentation. Such genes are apparently responsible for the modification of red color within the Wh Wh genotype of poinsettias containing both pelargonidin and cyanidin glycosides and for a very similar pink color in a snapdragon and a rose, each containing a single anthocyanin, a pelargonidin glycoside, and a cyanidin glycoside, respectively.     

Temporal and spatial regulation of anthocyanin biosynthesis provide diverse flower colour intensities and patterning in <Emphasis Type="Italic">Cymbidium</Emphasis> orchid

Main conclusion

This study confirmed pigment profiles in different colour groups, isolated key anthocyanin biosynthetic genes and established a basis to examine the regulation of colour patterning in flowers of Cymbidium orchid. Cymbidium orchid (Cymbidium hybrida) has a range of flower colours, often classified into four colour groups; pink, white, yellow and green. In this study, the biochemical and molecular basis for the different colour types was investigated, and genes involved in flavonoid/anthocyanin synthesis were identified and characterised. Pigment analysis across selected cultivars confirmed cyanidin 3-O-rutinoside and peonidin 3-O-rutinoside as the major anthocyanins detected; the flavonols quercetin and kaempferol rutinoside and robinoside were also present in petal tissue. β-carotene was the major carotenoid in the yellow cultivars, whilst pheophytins were the major chlorophyll pigments in the green cultivars. Anthocyanin pigments were important across all eight cultivars because anthocyanin accumulated in the flower labellum, even if not in the other petals/sepals. Genes encoding the flavonoid biosynthetic pathway enzymes chalcone synthase, flavonol synthase, flavonoid 3′ hydroxylase (F3′H), dihydroflavonol 4-reductase (DFR) and anthocyanidin synthase (ANS) were isolated from petal tissue of a Cymbidium cultivar. Expression of these flavonoid genes was monitored across flower bud development in each cultivar, confirming that DFR and ANS were only expressed in tissues where anthocyanin accumulated. Phylogenetic analysis suggested a cytochrome P450 sequence as that of the Cymbidium F3′H, consistent with the accumulation of di-hydroxylated anthocyanins and flavonols in flower tissue. A separate polyketide synthase, identified as a bibenzyl synthase, was isolated from petal tissue but was not associated with pigment accumulation. Our analyses show the diversity in flower colour of Cymbidium orchid derives not from different individual pigments but from subtle variations in concentration and pattern of pigment accumulation.

     

Cloning of FaPAL6 gene from strawberry fruit and characterization of its expression and enzymatic activity in two cultivars with different anthocyanin accumulation

The accumulation of anthocyanin pigments is one of the most important traits that turn strawberry fruit attractive to consumers. During ripening, strawberry fruit color development is associated to anthocyanin synthesis through the phenylpropanoid pathway. Phenylalanine ammonia-lyase (PAL) is a key enzyme in this pathway, having a determining role in strawberry fruit quality. In this work, we studied the level of anthocyanins during fruit ripening of two cultivars that differ in color development (Camarosa and Toyonoka). Toyonoka showed a lower anthocyanin accumulation that was limited to external fruit tissue, while Camarosa accumulated higher amount of anthocyanins in both internal and external sections. In addition, we cloned a full-length gene (FaPAL6) and analyzed its expression in different strawberry plant tissues. The expression of this gene is fruit specific, and increases during fruit ripening in both cultivars along with anthocyanin accumulation. The mRNA level of FaPAL6 was higher in Camarosa. PAL enzyme activity increased at similar rates in both cultivars at early ripening stages, but at the end of ripening PAL activity diminished in Toyonoka while it rose markedly in Camarosa. PAL activity was higher in internal fruit tissue, showing no correlation with anthocyanin level of the same section in both cultivars. The higher FaPAL6 expression and activity detected in Camarosa could be associated to the enhanced anthocyanin accumulation found in this cultivar.