Comparison of Phytochemical Composition and Biological Activities of Rubus ulmifolius Extracts Originating from Four Regions of Tunisia

In the current study, the phenolic composition, antioxidant and antimicrobial activities of extracts from Rubus ulmifolius Schott leaves harvested in four localities (Sejnen, Tabarka, Faija and Ain drahem) in Tunisia were investigated for the first time. Great differences were found for the chemical composition, total phenol contents and biological activities among the evaluated extracts. HPLC analysis of methanolic extracts showed that the dominant compounds were kaempferol 3‐O‐rutinoside and naringenine. In addition, significant correlations were observed between antioxidant activities and phenolic contents. In fact, leaves collected from Sejnen presented higher total phenol content (53.32 mg GAE/g DW) and antioxidant activities (IC₅₀ = 39.40 mg/l) than the others samples. All extracts showed significant antimicrobial activity against six used bacteria with the inhibition zones diameters and minimal inhibitory concentration values were in the range of 8 – 16 mm and 6.25 – 25 mg/ml, respectively. The highest antimicrobial activities were recorded in Sejnen extract against Gram‐positive bacteria.     

Profiling of Compositions of Essential Oils and Volatiles of Salvia limbata Using Traditional and Advanced Techniques and Evaluation for Biological Activities of Their Extracts

In the present work, the essential oils and volatiles from flowers, leaves, and stems of Salvia limbata obtained using microwave‐assisted hydrodistillation, solvent‐free microwave extraction, headspace‐assisted analysis, and headspace‐solid phase microextraction have been characterized for the first time. The results have been also compared with those from traditional separation techniques involving hydrodistillation and steam distillation. Regardless of some common compounds in all of the profiles, some dissimilarities were noted due to the use of different extracting approaches. Taking into account the chemical categories, sesquiterpene hydrocarbons were found as the most represented group of natural compounds contributing to the chemical profiles. It was also noted that the methanol extracts obtained from the flowers of Sal. limbata showed a desirable antioxidant activity, comparable to the standard antioxidant butylated hydroxytoluene. Furthermore, using the disc diffusion and broth microdilution methods, all the tested bacteria demonstrated weak to moderate and moderate to strong sensibilities to the MeOH extracts obtained from different plant parts of Sal. limbata.     

Pollen aroma fingerprint of two sunflower (Helianthus annuus L.) genotypes characterized by different pollen colors

Samples of fresh pollen grains, collected from capitula in full bloom from two genotypes of sunflower (Helianthus annuus L.) and characterized by a different color, i.e., white‐cream (WC) and orange (O), were analyzed by the HS‐SPME (headspace solid phase microextraction)/GC/MS technique. This study defined for the first time the fingerprint of the sunflower pollen, separated from the disc flowers, to define its contribution to the inflorescence aroma. In the GC/MS fingerprints of the WC and O genotypes, 61 and 62 volatile compounds were identified, respectively. Monoterpene hydrocarbons (34% in O vs. 28% in WC) and sesquiterpene hydrocarbons (37% in O vs. 31% in WC) were ubiquitous in all samples analyzed and represented the main chemical classes. α‐Pinene (21% in O vs. 20% in WC) and sabinene (11% in O vs. 6% in WC) were the dominant volatiles, but also a full range of aliphatic hydrocarbons and their oxygenated derivatives gave a decisive contribution to the aroma composition (10% in O vs. 12% in WC). In addition, dendrolasin (3% in O vs. 4% in WC) and some minor constituents such as (E)‐hex‐2‐en‐1‐ol (0.4% in O vs. 0.1% in WC) were pointed out not only for their contribution to the pollen scent, but also for their well‐known role in the plant ecological relationships. Having evaluated two pollen morphs with different carotenoid‐based colors, the study sought to highlight also the presence of some volatile precursors or derivatives of these pigments in the aroma. However, the pollen aroma of the two selected genotypes made a specific chemical contribution to the sunflower inflorescence scent without any influence on carotenoid derivatives.     

NMR,HS‐SPME‐GC/MS,and HPLC/MSn Analyses of Phytoconstituents and Aroma Profile of Rosmarinus eriocalyx

In this work, a comprehensive study on the chemical constituents of the aerial parts of Rosmarinus eriocalyx (Lamiaceae), an aromatic shrub traditionally consumed as a food and herbal remedy in Algeria, is presented. The aroma profile was analysed by headspace solid phase microextraction (HS‐SPME) coupled with gas chromatography‐mass spectrometry (GC/MS), whereas the crude extract constituents were analyzed by ¹H‐NMR and by high performance liquid chromatography coupled with mass spectrometry (HPLC/MSⁿ). Thirty‐nine volatile compounds, most of them being monoterpenes, have been identified, with camphor, camphene, and α‐pinene as the most abundant constituents. ¹H‐NMR analysis revealed the presence of phenolic compounds and betulinic acid while HPLC/MSⁿ allowed the identification of glycosilated and aglyconic flavonoids as well as phenylpropanoid derivatives. Some of these constituents, namely as betulinic acid, rosmanol, and cirsimaritin were reported for the first time in R. eriocalyx.     

Variation in Phenolic Content,Profile, and Antioxidant Activity of Seeds among Different Paeonia ostii Cultivated Populations in China

The purpose of this study was to analyze the phenolic profiles of seeds from fifteen Paeonia ostii cultivated populations in China and identify their relationship with antioxidant activities and associated environmental factors. Thirteen individual phenolic compounds were quantitatively determined by HPLC, and (+)‐catechin was the most abundant phenolic compound in the seeds. Correlation analysis showed that phenolics were the most effective antioxidant compound class by evaluating DPPH, ABTS, and hydroxyl radical scavenging activities as well as ferric reducing antioxidant power. Latitude and annual rainfall had significant effects on the contents of many phenolic compounds, and elevation was only significantly correlated with gallic acid content. Within fifteen P. ostii cultivated populations, the seeds of Tongling population exhibited the highest phenolic contents and strongest antioxidant activities. These results suggest that Tongling population has a relatively high utilization value and a potential for sources of natural antioxidants.     

Marker Gene Overexpression in Flowers Treated with Resistance Inducers does not Correlate with Protection Against Flower‐Infecting Fungi in Tomato and Blueberry

Flowers can serve as infection courts for specialized and unspecialized plant pathogens, but little is known about the ability of floral tissues to undergo induced resistance (IR) responses against these pathogens. We studied the expression of IR marker genes in tomato and blueberry flowers treated with the inducers methyl jasmonate (MeJA), benzothiadiazole‐S‐methyl ester (BTH) and 2,6‐dichloroisonicotinic acid (INA). In tomato, spray application of MeJA and BTH (but not INA) to entire plants (leaves, stems and flowers) resulted in a significant (P < 0.05) overexpression of Pin2 (5.2‐fold) and PR‐4 (5.6‐fold) in pistil tissues, respectively. A statistically similar expression was obtained in pistils when flowers were protected from direct spray, indicating a systemic response. In blueberry, where information about IR marker genes is limited, PR‐3 and PR‐4 orthologs were first identified and characterized using in silico and wet‐laboratory techniques. In subsequent induction experiments, INA and BTH induced overexpression of PR‐4 in blueberry pistils by 3.2‐ and 1.8‐fold, respectively, when entire plants were treated. In blueberry flowers protected from spray applications, all chemicals applied to vegetative tissues led to significant overexpression of PR‐4 (MeJA: 1.4‐fold, BTH: 2.9‐fold and INA: 1.6‐fold), with BTH also inducing PR‐3 (1.7‐fold). The effect of these responses in protecting flowers was studied by inoculating treated tomato flowers with the necrotroph Botrytis cinerea and blueberry flowers with the hemi‐biotroph Monilinia vaccinii‐corymbosi. In both pathosystems, no significant disease suppression associated with resistance inducer application was observed under the conditions studied. Thus, although IR marker genes were shown to be inducible in floral tissue, the magnitude of this response was insufficient to suppress pathogen ingress.     

不同产地丁香挥发性成分分析

采用顶空固相微萃取-气相色谱-质谱联用技术对产自国内外4个地区的丁香(Eugenia caryophyllata Thunb.)挥发性成分进行了检测,并以峰面积归一化法计算了各组分的相对含量。结果显示,从4个产地13批次丁香中共鉴定出挥发性成分72种(匹配度均高于75),其中,主要成分含量排名前两位的均为丁香酚、(-)-α-芹子烯,排名第三位的挥发性成分分别为:顺式-α-没药烯(马达加斯加塔马塔夫,平均为3.75%)、罗勒烯(中国广东,平均为4.21%)、亚麻三烯(中国广西,平均为3.74%)、丁香烯(印度尼西亚爪哇岛,平均为3.60%)。表明同一产地的丁香挥发性成分具有一定的相似性,不同产地丁香挥发性成分也有一定的差异,这对丁香产地的鉴别具有重要意义。主成分分析和聚类分析的结果既可以将4个产地的丁香很好地区分开来,又能反映出它们之间的亲缘关系。本研究采用HS-SPME-GC-MS方法检测不同产地丁香挥发性成分并结合主成分分析(PCA)及聚类分析法,能有效区分4个产地的丁香,该方法可作为丁香产地的鉴别方法,也为进一步比较不同产地丁香挥发性成分的差异及质量控制提供了新思路。     

5个番石榴品种果实食用品质和香气特征分析

为比较番石榴(Psidium guajava)不同品种果实的外观、营养价值和香气特征,采用国家标准方法,对5个番石榴品种(‘红宝石’、‘粉红蜜’、‘西瓜’、‘水蜜’、本地种)果实的外观和营养成分进行测定,采用顶空气质联用(HS-GC-MS)技术对5个品种果实的香气成分进行测定。结果表明,不同品种果实外观及营养成分差异明显。‘水蜜’的果形指数最低,果实扁圆形,种籽最少;大多数糖类物质(总糖、还原糖、蔗糖等)、糖酸比、总酚含量最高;果糖、VC和总黄酮含量位居第2;总酸、脂肪、粗纤维、灰分含量最低。主成分分析表明,‘水蜜’作为鲜食水果的食用品质最高。己醛和石竹烯是红肉型品种的特征风味物质,3-羟基-2-丁酮是白肉型品种的特征香气成分。     

Volatile Compounds of Viola odorata Absolutes: Identification of Odorant Active Markers to Distinguish Plants Originating from France and Egypt

Absolutes isolated from Viola odorata leaves, valuable materials for the flavor and fragrance industry, were studied. Violets are mainly cultivated in France and Egypt and extracted locally. The absolutes of the two origins showed different olfactory profiles both in top and heart notes, as evidenced by sensory analysis. The aims of this study were i) to characterize the volatile compounds, ii) to determine the odorant‐active ones, and iii) to identify some markers of the plant origin. Two complementary analytical methods were used for these purposes, i.e., headspace solid‐phase microextraction (HS‐SPME) using different fiber coatings followed by GC/MS analysis and gas chromatography – olfactometry/mass spectrometry (GC‐O/MS) applied to violet leaf extracts. From a total of 70 identified compounds, 61 have never been reported so far for this species, 17 compounds were characterized by both techniques (with seven among them known from the literature), 23 compounds were solely identified by HS‐SPME GC/MS (among them only two being already mentioned as components of violet absolutes in the literature), and, finally, 30 compounds were only identified by GC‐O/MS. According to the HS‐SPME GC/MS analyses, ethyl hexanoate and (2E,6Z)‐nona‐2,6‐dienol were specific volatile compounds of the sample with French origin, while (E,E)‐hepta‐2,4‐dienal, hexanoic acid, limonene, tridecane, and eugenol were specific of the samples with Egyptian origin. Additional compounds that were not detected by HS‐SPME GC/MS analysis were revealed by GC‐O analyses, some of them being markers of origin. Pent‐1‐en‐3‐ol, 3‐methylbut‐2‐enal, 2‐methoxy‐3‐(1‐methylethyl)pyrazine, 4‐ethylbenzaldehyde, β‐phenethyl formate, and 2‐methoxy‐3‐(2‐methylpropyl)pyrazine revealed to be odorant markers of the French sample, whereas cis‐rose oxide, trans‐rose oxide, and 3,5,5‐trimethylcyclohex‐2‐enone were odorant markers of the Egyptian samples.     

Eudesmanolides and Other Constituents from the Flowers of Wedelia trilobata

Two eudesmane sesquiterpene lactones, wedetrilides B ( 1 ) and C ( 2 ), along with five known analogues ( 3 – 8 ), an ent‐kaurane diterpenoid ( 9 ), a steroid ( 10 ), as well as cinnamic acid derivatives ( 11 – 13 ), were isolated from the flowers of Wedelia trilobata. Their structures were elucidated on the basis of extensive spectroscopic analyses and by comparison of their NMR data with those of related compounds. Furthermore, the structures of 1 and 3 – 5 were confirmed by X‐ray single‐crystal diffraction analyses. Compounds 4 and 5 exhibited weak cytotoxic activities against the MCF‐7, HeLa, and A549 cell lines. Compounds 3 – 5 were also evaluated for their inhibitory effects against HIV lytic replication.     

Differential Organ Distribution,Pathogenicity and Benomyl Sensitivity of Colletotrichum spp. from Blackberry Plants in Northern Colombia

Blackberry anthracnose, caused by Colletotrichum spp., is an important disease of cultivated blackberry in the world. In Colombia, it is the number one limiting factor for commercial production. This study was conducted to determine the species of Colletotrichum infecting blackberry plants as well as the organ distribution, pathogenicity and response to benomyl of the isolated strains. Sixty isolates from stems (n = 20), thorns (n = 20) and inflorescences (n = 20) were identified as Colletotrichum acutatum and Colletotrichum gloeosporioides by a species‐specific polymerase chain reaction (PCR). Both Colletotrichum species were found in the same plant but on different organs. Colletotrichum gloeosporioides species predominated in thorn lesions (n = 16) and C. acutatum in stems (n = 15) and inflorescence (n = 15). Pathogenicity assays on detached blackberry organs demonstrated differences between the two species with an average period of lesion development of 8.7 days for C. gloeosporioides and 10.3 days for C. acutatum. Wound inoculated organs had 90% disease development compared to 17.5% in non‐wounded. All C. acutatum isolates (n = 34) were benomyl tolerant, whereas C. gloeosporioides isolates (n = 26) were 30.7% sensitive and 69.2% moderately tolerant. Phylogenetic analysis with ITS sequences of a subset of 18 strains showed that strains classified as C. gloeosporioides had 100% identity to Colletotrichum kahawae, which belongs to the C. gloeosporioides species complex, whereas C. acutatum strains clustered into two different groups, with high similarity to the A2 and the A4 molecular groups. These data demonstrate for the first time the differential distribution of both species complexes in blackberry plant organs and further clarifies the taxonomy of the strains.     

嘉宝果不同发育期花果叶的挥发性成分分析

为了解嘉宝果(Myrciaria cauliflora)的挥发性成分,利用顶空-气相色谱/质谱联用技术对其不同发育期的花、果、叶的挥发性成分进行测量。结果表明,萜烯类是花、果、叶的主要挥发性成分,随开花进程呈增加趋势,随果实和叶片成熟进程而下降,单萜类是其中的优势成分,主要有α-蒎烯、β-蒎烯、D-柠檬烯、β-罗勒烯等;花苞期、初花期和嫩叶以β-蒎烯含量最高,盛花期和老叶以α-蒎烯最高,末花期和果实中均以D-柠檬烯含量最高,嫩叶中α-蒎烯和β-蒎烯的总含量高达62.07%。酯类在花期中以初花期含量最高(16.92%),在果实中以完熟期含量最高(14.81%),老叶中酯类含量(4.35%)显著高于嫩叶(0.26%)。因此,α-蒎烯和β-蒎烯是花、果、叶共有主香成分,柳酸甲酯和苯乙醇是花苞期和初花期特征主香成分,毕澄茄烯是完熟果特征风味物质,β-石竹烯是嫩叶特征主香成分,桉叶油醇和β-胡椒烯是老叶特征主香成分。     

Phloroglucinol Derivatives from the Fruits of Eucalyptus globulus and Their Cytotoxic Activities

A new phloroglucinol derivative, named eucalyptin B ( 1 ), along with five related known compounds ( 2 – 6 ), was isolated from the fruits of Eucalyptus globulus. Their structures were elucidated by means of 1D‐ and 2D‐NMR spectroscopy, with the absolute configuration of 1 determined by electronic circular dichroism (ECD) calculations. All isolated compounds ( 1 – 6 ) were evaluated for their cytotoxic activities against lung (A549), breast (4T1), and skin (B16F10) cancer cell lines. On the basis of cell viability assay, the cytotoxic activity of eucalyptin B ( 1 ) was further confirmed by apoptosis assay. Additionally, after treatment with eucalyptin B ( 1 ), the apoptosis factor proteins (Bcl2 and Bax) and caspase‐3 levels in A549 cells were also determined by Western‐blot analysis. By cytotoxic assay, eucalyptin B ( 1 ) exhibited potent cytotoxicity against A549 cells with an IC₅₀ value of 1.51 μm and induced concentration dependent apoptosis of up to 49%. Additionally, eucalyptin B ( 1 ) inhibited 5‐fold and increased 10‐folds in the level of Bcl2 and Bax, respectively. Furthermore, the 11‐fold increase in the level of caspase‐3 confirmed eucalyptin B ( 1 ) activated caspase dependent apoptosis pathway. In conclusion, the isolated compound eucalyptin B ( 1 ) has promising cytotoxic activity in tumor cells, especially in A549.     

Differences in the Detection of BrdU/EdU Incorporation Assays Alter the Calculation for G1, S,and G2 Phases of the Cell Cycle in Trypanosomatids

Trypanosomatids are the etiologic agents of various infectious diseases in humans. They diverged early during eukaryotic evolution and have attracted attention as peculiar models for evolutionary and comparative studies. Here, we show a meticulous study comparing the incorporation and detection of the thymidine analogs BrdU and EdU in Leishmania amazonensis, Trypanosoma brucei, and Trypanosoma cruzi to monitor their DNA replication. We used BrdU‐ and EdU‐incorporated parasites with the respective standard detection approaches: indirect immunofluorescence to detect BrdU after standard denaturation (2 M HCl) and “click” chemistry to detect EdU. We found a discrepancy between these two thymidine analogs due to the poor detection of BrdU, which is reflected on the estimative of the duration of the cell cycle phases G1, S, and G2. To solve this discrepancy, we increase the exposure of incorporated BrdU using different concentrations of HCl. Using a new value for HCl concentration, we re‐estimated the phases G1, S, G2 + M, and cytokinesis durations, confirming the values found by this approach using EdU. In conclusion, we suggest that the studies using BrdU with standard detection approach, not only in trypanosomatids but also in others cell types, should be reviewed to ensure an accurate estimation of DNA replication monitoring.     

The Volatile Profiles of a Rare Apple (Malus domestica Borkh.) Honey: Shikimic Acid‐Pathway Derivatives,Terpenes, and Others

The volatile profiles of rare Malus domestica Borkh . honey were investigated for the first time. Two representative samples from Poland (sample I) and Spain (sample II) were selected by pollen analysis (44–45% of Malus spp. pollen) and investigated by GC/FID/MS after headspace solid‐phase microextraction (HS‐SPME) and ultrasonic solvent extraction (USE). The apple honey is characterized by high percentage of shikimic acid‐pathway derivatives, as well as terpenes, norisoprenoids, and some other compounds such as coumaran and methyl 1H‐indole‐3‐acetate. The main compounds of the honey headspace were (sample I; sample II): benzaldehyde (9.4%; 32.1%), benzyl alcohol (0.3%; 14.4%), hotrienol (26.0%, 6.2%), and lilac aldehyde isomers (26.3%; 1.7%), but only Spanish sample contained car‐2‐en‐4‐one (10.2%). CH₂Cl₂ and pentane/Et₂O 1 : 2 (v/v) were used for USE. The most relevant compounds identified in the extracts were: benzaldehyde (0.9–3.9%), benzoic acid (2.0–11.2%), terpendiol I (0.3–7.4%), coumaran (0.0–2.8%), 2‐phenylacetic acid (2.0–26.4%), methyl syringate (3.9–13.1%), vomifoliol (5.0–31.8%), and methyl 1H‐indole‐3‐acetate (1.9–10.2%). Apple honey contained also benzyl alcohol, 2‐phenylethanol, (E)‐cinnamaldehyde, (E)‐cinnamyl alcohol, eugenol, vanillin, and linalool that have been found previously in apple flowers, thus disclosing similarity of both volatile profiles.     

Phytochemical Profile and Biological Activities of Helianthemum canum l. baumg. from Turkey

In the current study, antioxidant, antibacterial activities, and the phenolic compositions of extracts from Helianthemum canum L. Baumg . (Apiaceae) aerial parts were investigated for the first time. The H . canum was extracted with 70% methanol (HCM eOH ) and water (HCW ). Both extracts were determined by total phenolic contents (3 mg/ml), flavonoids (1.5 mg/ml), flavonols (1.5 mg/ml), qualitative–quantitative compositions, iron (II ) chelation activities (0.1 – 5 mg/ml), free radical scavenging activities (DPPH ^?: 0.01 – 0.6 mg/ml and ABTS ^+?: 0.125 – 0.5 mg/ml) and the effect upon inhibition of β ‐carotene/linoleic acid co‐oxidation (1 mg/ml). The peroxidation level was also determined using the thiobarbituric acid method (0.01 – 1.5 mg/ml). The results of the activity tests given as IC ₅₀ values were estimated from non‐linear algorithm and compared with standards. Antibacterial activities of extracts and standards were evaluated against Gram‐ negative and ‐positive ten standard strains using disc diffusion and broth microdilution methods. The MIC results (312.5 – 2500 μg/ml) against tested microorganisms varied from 625 to 2500 μg/ml. In HPLC analysis, 3,5‐dihydroxybenzoic acid was found as the main substance in both extracts. These results showed that HCM eOH was richer in phenolic compounds (284.13 ± 0.30 mg GAE /g extract) from HCW (244.55 ± 0.35 mg GAE /g extract)_. In conclusion, H . canum extracts showed in vitro antibacterial and antioxidant activities.