Flowers Of Cynara Cardunculus Research Articles

Exploring Cynara cardunculus L. by-products potential: Antioxidant and antimicrobial properties

Cynara cardunculus L. (cardoon), a perennial crop indigenous to the Mediterranean region, has gained recognition for its remarkable resilience to diverse weather conditions and its multifaceted applications across various industries, which includes the use of the flower as a vegetable rennet to produce some cheeses, as a source of biomass for energy, or its seed oil for human consumption, biodiesel, and animal feed. In some applications (e.g. biomass or seed production), when crop is harvested at the end of the growth cycle, the leaves remain as the main by-products, along with the flowers. In the context of a circular economy, the aim of this work was to undergone studies to determinate their biological properties (antioxidant and antimicrobial). Methanolic and ethanolic extracts of C. cardunculus L. (globe artichoke var. scolymus (L.) Fiori) and cultivated cardoon (var. altilis DC.)) leaves and flowers were characterised in terms of their polyphenol profile (total phenolic content (TPC), total flavonoid compounds (TFC), and ultra-high performance liquid chromatography coupled with time-of-flight mass spectrometry (UHPLC-ToF-MS)), antioxidant capacity (free radical DPPH inhibition system, β-carotene bleaching assay), and antimicrobial capacity (minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), antifungal). In addition, the cultivated cardoon leaves extracts were assessed before and after they were dried in an oven with forced air circulation to evaluate if this treatment affected their bioactive profile. Chlorogenic acid, apigenin, and luteolin were the most quantified of a total of sixteen compounds identified by UHPLC-ToF-MS. Cultivated cardoon dry leaf extract presented the best antioxidant capacity for both methanolic (EC50 = 0.8 mg/mL, antioxidant activity coefficient (AAC) = 279.67) and ethanolic (EC50 = 2.1 mg/mL, AAC = 448.06) extracts, compared to the cardoon flower extracts and the globe artichoke leaf extracts. Dried cultivated cardoon leaf extracts presented higher antioxidant capacity than fresh cultivated cardoon leaf extracts, but a greater number of polyphenolic compounds were identified in fresh cultivated cardoon leaf extract. The Gram-positive bacteria were more sensitive to the activity of both ethanolic and methanolic extracts than the Gram-negative and cultivated cardoon dry leaf ethanolic extract presented lower MIC and MBC values (125–2000 µg/mL) for most of the tested microorganisms, thus showing higher antimicrobial activity. As for the cultivated cardoon leaf extracts, the dried leaf extracts exhibited better antimicrobial activity, with lower MIC values, than the fresh leaf extracts. The extracts only demonstrated a slight inhibition against the fungi Aspergillus fumigatus. In conclusion, studies performed indicate that dried leaves maintain their biological activities compared to fresh leaves, and that flowers present significant biological activity which suggests the great potential of the by-products of this crop as a source of active compounds in different industrial applications (e.g. food industry).

Onopordum platylepis (Murb.) Murb. as a novel source of thistle rennet: First application to the manufacture of traditional Italian raw ewe’s milk cheese

In this study, for the very first time, aqueous extracts obtained from flowers of spontaneously grown or cultivated Onopordum platylepis (Murb.) Murb. thistles were used as coagulating agents for the pilot-scale manufacture of Caciofiore, a traditional Italian raw ewe’s milk cheese. Cheese prototypes were compared to control cheeses curdled with a commercial thistle rennet obtained from flowers of Cynara cardunculus L. After 45 days of ripening under controlled conditions, both the experimental and control cheese prototypes were analyzed for: cheese yield, physico-chemical (pH, titratable acidity, aw, proximate composition), morpho-textural (color and texture), and microbiological parameters (viable cell counts and species composition assessed by Illumina sequencing), as well as volatile profile by SPME-GC–MS. Slight variations in titratable acidity, color, and texture were observed among samples. Based on the results overall collected, neither the yield nor the proximate composition was apparently affected by the type of thistle coagulant. However, the experimental cheese prototypes curdled with extracts from flowers of both spontaneous or cultivated thistles showed 10 % higher values of water-soluble nitrogen compared to the control prototypes. On the other hand, these latter showed slightly higher loads of presumptive lactococci, thermophilic cocci, coliforms, and eumycetes, but lower counts of Escherichia coli. No statistically significant differences were revealed by the metataxonomic analysis of the bacterial and fungal biota. Though most volatile organic compounds (VOCs) were consistent among the prototypes, significant variability was observed in the abundance of some key aroma compounds, such as butanoic, hexanoic, and octanoic acids, ethanol, propan-2-ol, isobutyl acetate, 2-methyl butanoic acid, and 3-methyl butanal. However, further investigations are required to attribute these differences to either the type of coagulant or the metabolic activity of the microorganisms occurring in the analyzed cheese samples. The results overall collected support the potential exploitation of O. platylepis as a novel source of thistle coagulant to produce ewe’s milk cheeses.

A new methodology for a rapid and high-throughput comparison of molecular profiles and biological activity of phytoextracts.

To robustly discover and explore phytocompounds, it is necessary to evaluate the interrelationships between the plant species, plant tissue, and the extraction process on the extract composition and to predict its cytotoxicity. The present work evaluated how Fourier Transform InfraRed spectroscopy can acquire the molecular profile of aqueous and ethanol-based extracts obtained from leaves, seeds, and flowers of Cynara Cardunculus, and ethanol-based extracts from Matricaria chamomilla flowers, as well the impact of these extracts on the viability of mammalian cells. The extract molecular profile enabled to predict the extraction yield, and how the plant species, plant tissue, and extraction process affected the extract's relative composition. The molecular profile obtained from the culture media of cells exposed to extracts enabled to capture its impact on cells metabolism, at a higher sensitivity than the conventional assay used to determine the cell viability. Furthermore, it was possible to detect specific impacts on the cell's metabolism according to plant species, plant tissue, and extraction process. Since spectra were acquired on small volumes of samples (25 µL), after a simple dehydration step, and based on a plate with 96 wells, the method can be applied in a rapid, simple, high-throughput, and economic mode, consequently promoting the discovery of phytocompounds.

Technological characteristics of tofu from soybean coagulated with cardoon flower extract.

The objective of the present study was to develop tofu with soybean-water soluble extract coagulated with cardoon flower (F1) and magnesium chloride (MgCl2, F2). The produced tofu was characterized in terms of physical, chemical, microbiological, and sensory properties during 14days of storage. The yield of F1 was higher (p < 0.05) (195g/100 soybean seeds) than F2 (162g/100 soybean seeds). F1 presented higher moisture, protein, acidity, syneresis, and lipids when compared with F2, and a reduction of these contents during the storage. F1 presented lower hardness, stickiness, springiness, and cohesiveness compared with F2. The acceptability of F1 showed a score of 6.00 and F2 of 4.68, and the purchase intention was 3.22 for F1 and 2.23 for F2. This study recommended the use of cardoon flower at 35% level as it has great potential as a coagulant for the elaboration of tofu with higher yield, and acceptability and reasonable purchasing intention.

Queijo Serra da Estrela PDO Cheese: Investigation into Its Morpho-Textural Traits, Microbiota, and Volatilome.

Queijo Serra da Estrela is a PDO Portuguese cheese produced through coagulation of raw ewe’s milk using cardoon (Cynara cardunculus L.) flowers. The present research was aimed at depicting an up-to-date and comprehensive overview of the microbiota of Queijo Serra da Estrela cheese. To this end, viable counting and metataxonomic analysis were carried out on cheeses sampled from four Portuguese artisan producers. Physico-chemical and morpho-textural analyses were also performed, together with the analysis of volatile organic compounds (VOCs). Finally, non-starter lactic acid bacteria (NSLAB) isolated from the cheeses were characterized for their enzymatic activities using a semi-quantitative method. According to the metataxonomic analysis, Lactococcus lactis and Lactococcus piscium were the species occurring at the highest relative abundance. The isolates collected from the cheeses were assigned to Enterococcus durans, Enterococcus faecalis, Enterococcus faecium, Enterococcus lactis, Levilactobacillus brevis, Latilactobacillus graminis, Leuconostoc mesenteroides, and the Lacticaseibacillus casei group. The enzymatic characterization of these cultures highlighted esterase, aminopeptidase, acid phosphatase, beta-galactosidase, alpha-glucosidase, and beta-glucosidase among the major enzymatic activities. Fungal populations were dominated by Debaryomyces hansenii and Kurtzmaniella zeylanoides; however, species rarely found in cheese (e.g., Candida boidinii, Vishniacozyma victoriae, and Starmerella) were also detected. The volatile compounds characterizing the analyzed cheeses were carboxylic acids and esters, followed by carbonyl compounds and alcohols.

Properties of tofu coagulated with Cardoon flower (Cynara Cardunculus L.)

Tofu was elaborated using water-soluble extract of soybean grains from the cultivar BRS 267 with cardoon flower (C. Cardunculus L.), as coagulant as an alternative to the traditional coagulant magnesium chloride. The tofu were evaluated in relation to physical, chemical, microbiological and sensory properties during 14 days. Was obsersed for the tofu coagulated with cardoon flower that the moisture, pH, total mineral, protein, and syneresis decreased with the storage, while Kunitz Trypsin Inhibitor, ureatic activity and phytic acid did not change. It was also obtained a positive effect for color (L*, a*) parameters, phytic acid, yield, syneresis, and acceptance. The textural properties, showed a tendency to increase of hardness and stickiness, while springiness and cohesiveness decreased during storage. Structurally, presented irregular-shaped, small pores or cavities. The sensory analysis, showed has great acceptance and purchase intent. Then, the cardoon flower can be used as vegetal coagulant improving the quality properties of tofu, in relation to magnesium chloride.

Front Cover: Synthesis of 2,2’‐Bipyridines through Catalytic C−C Bond Formations from C−H Bonds (Eur. J. Org. Chem. 39/2021)

The Front Cover shows – drawn in a cartoon-style illustration – the different strategies to synthesize 2,2′-bipyridines involving a C−H bond functionalization step. On the left side, the three approaches are inlaying in green lines. The catalysts (transition metal) are drawn in a bubble to illustrate the challenge of maintaining their catalytic activity while 2,2’-bipyridines can coordinate and affect their properties. On the right side, the 2,2′-bipyridine is designed as an allegory featuring a pink Cardoon flower for ligands and a bee for metals, showing their mutual bonding. Anne-Sophie Pochet is thanked for taking the original photography on which this cover is built. More information can be found in the Minireview by J.-F. Soulé et al.

Tobacco BY2 cells expressing recombinant cardosin B as an alternative for production of active milk clotting enzymes

Cynara cardunculus L. or cardoon is a plant that is used as a source of milk clotting enzymes during traditional cheese manufacturing. This clotting activity is due to aspartic proteases (APs) found in the cardoon flower, named cyprosins and cardosins. APs from cardoon flowers display a great degree of heterogeneity, resulting in variable milk clotting activities and directly influencing the final product. Producing these APs using alternative platforms such as bacteria or yeast has proven challenging, which is hampering their implementation on an industrial scale. We have developed tobacco BY2 cell lines as an alternative plant-based platform for the production of cardosin B. These cultures successfully produced active cardosin B and a purification pipeline was developed to obtain isolated cardosin B. The enzyme displayed proteolytic activity towards milk caseins and milk clotting activity under standard cheese manufacturing conditions. We also identified an unprocessed form of cardosin B and further investigated its activation process. The use of protease-specific inhibitors suggested a possible role for a cysteine protease in cardosin B processing. Mass spectrometry analysis identified three cysteine proteases containing a granulin-domain as candidates for cardosin B processing. These findings suggest an interaction between these two groups of proteases and contribute to an understanding of the mechanisms behind the regulation and processing of plant APs. This work also paves the way for the use of tobacco BY2 cells as an alternative production system for active cardosins and represents an important advancement towards the industrial production of cardoon APs.

Biochemical diversity of cardoon flowers (Cynara cardunculus L.): Predicting PDO Mediterranean cheese textures

Cardoon flower extract is a vegetable coagulant used in the manufacture of some highly appreciated Mediterranean Protected Designation of Origin (PDO) sheep cheeses, due to their typical flavor and smooth texture. Flowers from 6 cardoon plants (1M–6M), selected on the basis of the total and specific A0, A and B cardosin concentrations were characterized using chromatographic and electrophoretic analysis. Extracts of these flowers were used to investigate the relationship between the biochemical profile and the proteolytic activity on caseins, coagulation properties and cheese texture. Milk-clotting activity and coagulation were measured using an Optigraph. Cheese making trials were done using the traditional protocol of Serra da Estrela PDO cheese manufacture. A multivariate exploratory PCA analysis showed that plants 4M, 5M and 6M with the highest total concentration of cardosins, the prevalence of cardosin A, proteolytic action with Ƙ-casein, and the lowest time of coagulation yielded cheeses with a firmer paste. Plants 1M, 2M and 3M, without the presence of cardosin A, were negatively influenced by the proteolytic action with β-casein, starting flocculation and gel consistency. Plant 3M showed a slow micellar aggregation rate and low gel firmness during milk coagulation, which was reflected in the smooth texture of the cheese. The specific characterization of natural biochemical diversity of cardoon flowers and their influence on cheese texture may help producers to choose the appropriated cardoon flowers to obtain a desired texture.

Evaluation of gas holes in "Queijo de Nisa" PDO cheese using computer vision.

"Queijo de Nisa" is a traditional Portuguese cheese, granted with PDO label, produced with raw ewe's milk in which the aqueous extract of cardoon flower Cynara cardunculus L. is the only coagulant allowed. As in similar cheeses with no use of starter cultures or pasteurisation, the quality and food safety are depending on prevention, high hygienic standards and a proper manufacturing process. This study investigated the use of computer vision as novel method for the evaluation of gas holes in Queijo de Nisa in three different ripening dates (0, 15 and 35days). A total of 48 samples were produced using cardoon flower from three different origins (C1, C2 and C3) and a commercial vegetable coagulant (C4). The results presented a high correlation between image-dependent attributes and physical-chemical properties during ripening time, especially within the first 15days of ripening time, where major structural changes were observed inside the Queijo de Nisa cheese. Principal component analysis presented a strong correlation (p < 0.05) between image parameters and the physical-chemical evolution until 15days. From 15 to 35days, the evolution of cheeses was mainly depending on structural parameters, like G'1Hz and hardness. No influence was observed due to the geographical origin of cardoon flower.

Influence of Cardoon Flower (Cynara cardunculus L.) and Flock Lactation Stage in PDO Serra da Estrela Cheese.

Serra da Estrela (SE) cheese is one of the most appreciated Portuguese cheeses, being produced only from raw ewe’s milk, cardoon flower and salt. Cardoon takes part in two important processes in cheese production—coagulation and proteolysis—contributing to its unique features. Furthermore, milk chemical characteristics change during the milking season, being another factor that account for the high variability of cheese attributes. Therefore, the purpose of this work is to study the influence of cardoon flower (commercial, 6 M and 3 M) and flock lactation stage (November 2018, February and April 2019) in the final characteristics of SE cheese. The parameters analysed were moisture, protein, fat and salt contents, texture and colour. Results showed that flock lactation stage has the highest influence in all the studied characteristics, corresponding the early stages of lactation to the most protein-rich and low-fat cheeses. Cardoon flower affects mainly fat and rind colour. This study allows us to conclude that seasonal changes in ewe’s milk have a considerable impact in cheese attributes, and that although cardoon type had a more restrained effect, when used with expertise it may help adjust cheese sensory characteristics in order to obtain a final product that matches consumer acceptability requirements.

Algerian cardoon flowers express a large spectrum of coagulant enzymes with potential applications in cheesemaking

Proteases from flowers of Algerian cultivated and wild cardoons were purified, characterised and compared with those purified from flowers of a Portuguese variety. Three cardosins (A0, A and B) were obtained from each variety. All of them were dimeric and comprised heavy and light chain. Cardosins from Algerian varieties presented higher molecular masses and were less glycosylated than their Portuguese counterparts. Milk coagulation and curd yield parameters revealed a large difference between cardosins A0, A and B and among the same cardosin from different origins. The enzymatic specificity of cardosins, studied against β-chain of oxidised insulin and κ-casein, showed no prevalent effect of varieties. However, compared with cardosins A0 and A, cardosin B was more proteolytic and led to more complex digestion profiles. The present study reports the first description of the diversity of cardosins in Algerian cardoon flowers and sustains their potential use as milk coagulants for cheesemaking.

Cardosin A endocytosis mediated by integrin leads to lysosome leakage and apoptosis of epithelial cells.

Cardosin A is an aspartic protease present in large amount in the pistils of cardoon flowers. This protease is known to contain an -Arg-Gly-Asp- (RGD) motif located on the molecular surface. In this study, we found that isolated recombinant cardosin A attached to human epithelial cells A549, mediated by the binding of its RGD motif to cell surface integrins. The cell bound cardosin A was internalized to endosomes and lysosomes and triggered the permeability of lysosomal membrane leading to apoptosis of the epithelial cells. These events are identical to those observed for three RGD-containing aspartic proteases, Saps 4-6, secreted by Candida albicans. Such a process, which has been called the Trojan Horse mechanism, is believed to benefit the invasion of C. albican into the epithelium of the host. The location of the RGD motifs of cardosin A and Saps 4-6 are on the opposite ends of the homologous three-dimensional structures, suggesting that the Trojan Horse mechanism is insensitive to the RGD position. Current finding also suggests that cardosin A may have a defensive function against the ingestion of cardoon flowers by human, insects, and other herbivores.

Characterization of Cynara cardunculus L. flower from Alentejo as a coagulant agent for cheesemaking

The cardoon (Cynara cardunculus L.) is a mandatory vegetable coagulant for certain Protected Designation of Origin Portuguese cheeses. It grows wild in Portugal and is used without any type of control regarding flower picking or extract preparation, representing some uncertainty in cheese manufacture. The variability in technological properties, in the context of traditional cheese manufacture, of cardoon flower ecotypes from the Alentejo region was evaluated, including milk clotting and proteolytic activities, coagulation properties and potential cheesemaking yield of flower extracts. Multivariate statistics highlighted the variability of flower properties for cheesemaking, but allowed the aggregation of the ecotypes into five groups under the major influence of milk clotting activity and effect on gel firmness and micellar aggregation rate, followed by proteolytic activity. These differences may have an impact on cheese properties and therefore can allow the selection of cardoon flower for the manufacture of different types of cheese.

Enzymatic extract from flowers of Algerian spontaneous Cynara cardunculus: Milk‐clotting properties and use in the manufacture of a Camembert‐type cheese

The aim of this study was to assess the coagulant properties of crude extract from cardoon flowers (CECF) as a replacement for the imported coagulants used in the local Algerian cheese factories. Optimal coagulation activity of the extract was observed at pH 5 and 60 °C with a CaCl2 concentration of 0.02 M. The electrophoretic pattern revealed that the crude extract contains mainly cardosins A and B. The study of interactions involved in the formation of milk gel coagulated by CECF showed that hydrogen forces, hydrophobic interactions and calcium bridges contributed highly to the formation of milk gel. Camembert‐type cheese obtained by CECF was judged by trained panellists and was found to be in agreement with standard specifications. The present study highlights the possibility of using Algerian CECF as a promising plant coagulant in cheesemaking for the dairy industry.

Selected Cardoon (Cynara cardunculus L.) Genotypes Suitable for PDO Cheeses in Mediterranean Regions.

Cardoon flower extract is a traditional and exclusive rennet used for some PDO cheeses in several Mediterranean regions, due to its extremely high concentration in cardosins. In this preliminary study, six individual cardoon genotypes (1M-6M) were selected because they revealed a wide and consistent diversity of total and specific cardosin concentrations in flowers. During three growing seasons, the stability of 12 biochemical characteristics of flower extracts and 26 plant morphological descriptors was confirmed. Surprisingly, the cardosin profiles of each genotype, based on four main groups A0, A1, A and B, were stable during the annual flower harvesting period and over all threeyears using ion-exchange chromatography and native-PAGE electrophoresis. This knowledge will allow an improvement in the quality and standardization of cardosin profiles from cardoon flowers used for cheese production and other innovative applications. The results obtained are promising for the development of a plant breeding program based on biochemical and morphological characteristics in order to obtain the most adapted plant architecture for combined purposes related to specific cardosins composition, flower and plant biomass production, and ease of harvesting.

Influence of the ripening stage and the lyophilization of wild cardoon flowers on their chemical composition, enzymatic activities of extracts and technological properties of cheese curds

The selection of an appropriate Cynara cardunculus flowers batch is a preliminary step to produce rennet with better clotting properties. For this reason, we proposed to study the influence of the ripening stage and the lyophilization of cardoon flowers on their chemical composition, enzymatic activities of extracts, and technological properties of cheese curds. Results of flowers composition have shown that lyophilized flowers harvested at the middle of ripening stage (A) could be employed to produce mainly proteins or milk-clotting proteases. To confirm this, enzymatic activities of extracts and technological properties of curds were assessed. The experimental findings revealed that flowers lyophilization seems to be an efficient way to produce rennet with better clotting properties, leading to higher yield, moisture, and texture parameters of curd. These findings allowed us to select lyophilized flowers (A) for further cheese making process.

Morphological evaluation of cardoon (Cynara cardunculus L.): assessing biodiversity for applications based on tradition, innovation and sustainability

Selected cardoon cultivars established in the “Serra da Estrela” region were analysed to evaluate morphological characteristics related principally to the production of cardoon flowers, which are a compulsory ingredient for particular Mediterranean PDO cheese regions. The biodiversity of twelve cardoon cultivars installed in an experimental field were evaluated over three growing seasons using thirty-four morphological descriptors. Statistically significant differences were found between cultivars for twenty-four morphological characteristics which indicate a wide genetic diversity. The relationship among the cultivars and characteristics was analysed using principal component analysis. A three dimensional template was found to be very significant and explained 71% of the total variation. The first component is dominated positively by plant height, diameter of stalk, inflorescence characteristics and flower production, while the second component is positively dominated by leaf characteristics. Cultivars A26, D32 and D33, present a plant architecture simultaneously well adapted for flower production, ease of harvesting and plant biomass. These characterizations and understandings can be useful for a plant breeding programme to develop cultivars for innovative potential applications besides flowers, and also for application by other cheese producers in Mediterranean regions.

Pappi of cardoon (Cynara cardunculus L.): The use of wetting during the harvesting aimed at recovering for the biorefinery

The pappi of cardoon (Cynara cardunculus L.) flowers form a crown of plumose filaments which aid the wind dispersal of the seeds connected to them. The high cellulose content of this fraction can be a valuable raw material for fiber or pulp products. During threshing, the pappi are discharged together with the other biomass, but as they are liable to blow away, they fly away during discharging and from the windrow. As the biomass is baled within 3–4h after harvesting, the objective of the study was to test a wetting system applied to a combine and the effect of some adjuvant in limiting pappi dispersal caused by the wind. The system included a water tank (1m3), an electric pump, a flow regulator, and three pairs of nozzles to spray the solution onto the threshed material discharged from the rear of the combine. Four adjuvants in two concentrations were used to extend the retention time. The percentage of retention was evaluated on an artificial surface (plastic panels) and on the crop residues left in the windrows behind the combine. The retention time analysis was made one (T1) and four hours (T4) after harvesting. On the plastic panels, the pappi treated just with water showed a low level of sticking because the covered area had decreased by 92.9% at T1 compared to the time of harvesting (T0). The amount of pappi remaining on the windrow at T4 proved the efficacy of one of the adjuvants (AL-02) in limiting dispersal (52.8kgha−1), bearing in mind that without wetting (untreated), the amount remaining was seven times less (7.51kgha−1). Therefore, wetting appears a feasible option, compared to no treatment, to keep a certain amount of pappi on the soil. In order to reduce the dispersal and to increase the recovery of the pappi, the combine (equipped with the wetting system) should be rapidly followed by a baler, before the wetting effect disappears.

Effect of extraction pH on techno-functional properties of crude extracts from wild cardoon (Cynara cardunculus L.) flowers.

In the present paper, techno-functional properties of rennet derived from C. cardunculus, prepared at various pH (from 3 to 6), were investigated. It was found that the extract prepared at pH 3 had the best milk-clotting properties (MCA/PA ratio). It presented also the highest enzyme content, determined by densitometry analysis on 1D and 2D gels electrophoresis. Formation of milk gels produced by the extracts and chymosin was monitored using dynamic rheology and turbiscan. It can be assumed that by lowering rennet pH, milk pH decreases, causing a significant increase of curd firmness and both elastic (G') and viscous (G″) moduli. Results obtained by turbiscan showed similar ΔBS values of gels produced, after 2h, by chymosin and the crude extract at pH 3. In conclusion, C. cardunculus extract prepared at low pH has the potential to be employed as an efficient milk-clotting agent in the production of dairy products.