Callus Cultures Research Articles

Induction and Characteristics of Callus Cultures of the Medicinal Plant Tussilago farfara L.

Tussilago farfara L. is a traditional medicinal plant valued for its potentially health-promoting metabolites. Its herbal raw material has been recognized and used since ancient times and continues to be widely used in traditional medicine. Introducing this plant species to in vitro cultivation is a challenging task, but once the protocol is developed, such cultures can provide an abundant and inexhaustible source of plant material. In this study, we report the successful induction and growth of vigorous T. farfara callus in vitro. Callus induction was achieved on MS solid media with the combination of indole-3-acetic acid (3 mg/L) and benzyl aminopurine (2 mg/L) in darkness, whereas it appeared inefficient under light conditions and in suspension culture. We present a detailed description of callus growth kinetics, morphological analysis, photosynthetic activity, and biochemical parameters (including protein content and photosynthetic pigments) supported by histological studies. Furthermore, we observed the potential for organogenesis and somatic embryogenesis. This method for the in vitro propagation of T. farfara, along with callus culture maintenance, offers a wide range of applications in pharmacy for the production of valuable metabolites. Moreover, it could benefit the environment by reducing the depletion of natural populations of this species and may serve as an alternative strategy for species conservation in light of global warming.

Silver Nanoparticles (AgNPs) Act as Nanoelicitors in Melissa officinalis to Enhance the Production of Some Important Phenolic Compounds and Essential Oils

ABSTRACTNanoparticles (NPs) are well‐known biostimulants in plant biotechnology, utilised to enhance the physical properties of plants and exhibit positive effects on them. The important key role is the most suitable type, effective dose and size of NP to be used in plant tissue culture systems. In this study, various concentrations of silver nanoparticles (AgNPs; 0, 25, 50, 75 and 100 μg L−1) were tested as elicitors in callus culture with the aim of enhancing secondary metabolite production in lemon balm (Melissa officinalis L.). According to the results obtained, callus formation rates have shown an increase in all applications compared to the control group. The highest callus formation, weight and diameter were observed in 50 μg L−1 application. In this application, the callus structure was compact and its colour was green. However, the aromatic compounds, neral and geranial increased significantly in 25 μg L−1 application. The maximum increase in phenolic compounds such as caffeic acid, chlorogenic acid, proto‐catechic acid, hesperidin and p‐coumaric acid was observed in the 75 μg L−1 AgNP and the highest increase in rosmarinic acid compound was determined in the 50 μg L−1 application. The study found that AgNP applications are an effective method for increasing the production of secondary metabolites in medicinal and aromatic plants, such as lemon balm, in vitro.

Impact of zinc oxide nanoparticles on biosynthesis of thymoquinone in cell cultures of Nigella sativa

The rising market interest for Nigella sativa (Black seeds) necessitates the development of cultivation strategies to enhance metabolites production. Zinc oxide nanoparticles (ZnO-NPs) have drawn global attention as efficient and bio safe elicitors for in vitro cultures, to enhance secondary metabolites production in medicinal plants. In this study, ZnO-NPs were utilized for establishment of callus and cell cultures in black seeds for the first time. Hypocotyl explants were cultured on Murashige and Skoog (MS) media with varying levels of ZnO-NPs, resulted in callus induction and biomass formation. Optimal response in callus growth parameters were observed when explants were grown on MS media supplemented with 60 mg/L ZnO-NPs, resulting in 71.2 % callus induction frequency, 28.2 g/L fresh biomass, 9.7 g/L dry biomass, and 63 % water content. A substantial increase in callus growth was observed when ZnO-NPs were combined with 6-Benzylaminopurine (BA) at ratio of 45:1.5 mg/L, resulting in 91.2 % callus induction frequency and 42.2 g/L fresh biomass. In cell suspension cultures, ZnO-NPs alone at 45 mg/L produced optimum callus biomass (60.9 g/L). However, in combination with BA, callus biomass did not increase significantly in cell cultures. Maximum accumulation of total phenolic content (TPC: 26.8 mg GAE/g DW; Gallic acid equivalent dry weight) and total flavonoid content (TFC: 19.5 mg QE/g DW; Quercetin equivalent dry weight) was observed in cell cultures treated with higher concentration (70 mg/L) of ZnO-NPs in the 5th week of the growth curve. Moreover, ZnO-NPs incremented substantially the Phenylalanine lyase (PAL), Superoxide dismutase (SOD) and Peroxidase (POD) enzyme activities in cell cultures. Nonetheless, Reverse Phase High Performance Liquid Chromatography (RP-HPLC) analysis indicated peak production of thymoquinone (168.5 mg/g FW) in cell cultures treated with 45 mg/L ZnO-NPs alone. This study offers a promising approach for commercial production of Nigella sativa biomass and bioactive metabolites.

Effects Of 2,4-D, BAP, and Sucrose Concentrations in The Callus Induction of White (Clitoria ternatea var. Albiflora) and Blue Butterfly Pea (Clitoria ternatea)

The blue butterfly pea (Clitoria ternatea) and white butterfly pea (Clitoria ternatea var. Albiflora) belong to the Fabaceae family. Both are locally known as “bunga telang” and native to the Southeast Asian regions. The blue flowered variety is traditionally used to treat headaches, fever, and diabetes and is renowned scientifically for its memory-enhancing properties due to the presence of novel pentacyclic triterpenoids. However, farming of C. ternatea is challenged by inconsistent yields of novel secondary metabolites, especially under changing environmental conditions. Callus and cell suspension cultures, on the other hand, offer an alternative for the consistent production of these metabolites. The current study aims to optimize the treatments of 2,4-dichlorophenoxyacetic acid (2,4-D), 6-benzylaminopurine (BAP), and sucrose concentrations for friable callus formation from seedling explants. Sterile cotyledon explants of in vitro seedlings from both types of butterfly pea were subjected to half-strength MS medium supplemented with different concentrations and combinations of 2,4-D and BAP, with sucrose at 15 g/L and 30 g/L. The highest friable callus fresh weight from the white butterfly pea explants (0.064 ± 0.010 g) was achieved in treatments of 0.40 mg/L 2,4-D and 0.50 mg/L BAP. In contrast, the highest fresh weight of friable callus for the blue variety (0.025 ± 0.016 g) was induced in 0.25 mg/L of 2,4-D. Both varieties showed the highest friable callus weight in 15 g/L sucrose supplemented with 1.00 mg/L of 2,4-D (0.146 ± 0.032 g) and 0.25 mg/L of 2,4-D (0.245 ± 0.075 g) for the white and blue variety respectively. The morphology of calli for both varieties were yellowish, watery, and sticky. This study provides an essential basis the establishment of cell suspension cultures, as an efficient alternative to harness the secondary metabolites associated with the mammalian neuroprotective properties.

Analytical insight into caffeine extraction from typica coffee leaves based on crystallinity enhancement, optical phonon vibration upshift, and morphological evolution.

Caffeine extracted from callus cultures by in vitro technique induced from typica coffee (Coffea arabica L. var. typica) leaves was successfully carried out by a simple Soxhlet method. Analysis of X-ray diffraction patterns showed an increase in crystallinity fraction from leaves (13.56%) to callus (14.46%) and then to caffeine (39.18%). Crystallite size also varied, with average sizes of 18±6, 69±51, and 32.5±17 nm for leaves, callus, and caffeine, respectively. Fourier transmission infrared absorption data confirmed the presence of hydroxyl (OH) groups bound to carbon (C─COH), indicating caffeine content. The high stability of the C─COH is indicated by the broad optical phonon vibrations of the leaves: 247cm-1 to caffeine: 963cm-1. Quantitative analysis of dielectric function and electron loss function intensity peaks of each sample showed that leaves efficiently capture and store light energy while caffeine has less potency. Scanning electron microscopy analysis showed irregular shapes of leaves, oval round shapes for callus, and rectangular crystals for caffeine due to crystal orientation during transformation and had a strong correlation with crystallinity fraction. Finally, the structure-based identification, chemistry, optical-dielectric function, and micro-surface properties have been fully studied, thus unmasking the phenomenon of slow transformation from leaves to caffeine form. PRACTICAL APPLICATION: The result of this study can be applied to uncover new methodologies related to the classification, and biotechnological utilization of callus culture based on structural properties, optical-dielectric function, and micro-surface analysis. Methodologically, the resulting callus culture provides a sustainable and controllable supply of plant material for caffeine extraction, thereby reducing traditional methods involving field-grown plants and avoiding the use of pesticides.

Targeted regulation of 5-aminolevulinic acid enhances flavonoids, anthocyanins and proanthocyanidins accumulation in Vitis davidii callus

BackgroundSpine grape (Vitis davidii) is a promising source of high-quality anthocyanins, with vast potential for application in food, pharmaceutical, and cosmetic industries. However, their availability is limited by resource constraints. Plant cell culture has emerged as a valuable approach for anthocyanin production and serves as an ideal model to investigate the regulation of anthocyanin biosynthesis. Elicitors are employed to achieve targeted enhancement of anthocyanin biosynthesis. The present study investigated the impact of 5-aminolevulinic acid (ALA) as an elicitor on the accumulation of anthocyanins and flavonoids during spine grape callus growth. Specifically, we examined the effects of ALA on anthocyanin and its component accumulation in callus, and biosynthetic anthocyanin gene expression.ResultsALA at 25 µg/L increased the biomass of spine grape callus. ALA induction enhanced the levels of flavonoids, anthocyanins and proanthocyanidins in callus, with maximum values reaching 911.11 mg/100 g DW, 604.60 mg/100 g DW, and 5357.00 mg/100 g DW, respectively, after callus culture for 45 days. Notably, those levels were 1.47-, 1.93- and 1.83-fold higher than controls. ALA induction modulated the flavonoid profile, and among 97 differential flavonoid metabolites differing from controls, 77 were upregulated and 20 were downregulated. Six kinds of anthocyanins, namely cyanidin (8), delphinidin (6), peonidin (5), malvidin (4), petunidin (3) and pelargonidin (3), were detected in callus, with peonidin most abundant. Compared with controls, anthocyanin components were increased in ALA-treated callus. The key genes PAL1, PAL2, PAL4, CHI, CHS3, F3’H, F3H, FLS, DFR, UFGT, MYBA1, LDOX, OMT3, GT1 and ACT involved in anthocyanin biosynthesis were upregulated following ALA treatment, resulting in anthocyanin accumulation.ConclusionThis study revealed a novel mode of ALA-mediated promotion of plant anthocyanin biosynthesis and accumulation at the cellular level, and a strategy for enhancing anthocyanin content in spine grape callus. The findings advance commercial-scale production of anthocyanins via spine grape callus culture. we also explored the accumulation patterns of flavonoids and anthocyanins under ALA treatment. Augmentation of anthocyanins coincided with elevated expression levels of most genes involved in anthocyanin biosynthesis within spine grape callus following ALA treatment.

Optimization of callus induction protocol from leaf explants of P<i>ortulaca oleracea</i> and assessment of fatty acid profiles

A protocol for optimization of callus culture establishment from leaf explants of Portulaca oleracea L. was developed. The effect of ethyl methanesulphonate (EMS) on callus induction and synthesis of fatty acids was evaluated. Callus culture was initiated from leaf explants on Murashige and Skoog’s medium supplemented with 2,4‑dichlorophenoxy acetic acid (2,4‑D), a combination of 2,4‑D + 6‑benzyl adenine (BA) or 2,4‑D + kinetin. The maximum callus biomass was obtained at 2.5 µM BA + 2.5 µM 2,4‑D (20.22 g leaf explant–1 fresh weight). The leaves treated with EMS (0.1 to 0.4% h–1) differentiated callus on this optimized medium combination. The gas chromatography-mass spectrometry analysis of fatty acids indicated that the leaves had a high linolenic acid content (17.72%). The callus cultures synthesized heptadecanoic, oleic, and tricosanoic acids, which were otherwise absent in the leaves. Eicosanoic and docosanoic acids in callus cultures were 2.78 and 3.18 times higher than their content in the leaves, respectively. The diversity of fatty acids in treated callus cultures decreased with increased EMS concentration, but the content of a few individual fatty acids was enhanced. Callus at the dose of 0.1% EMS synthesized linoleic acid, which was two times higher than in the untreated callus, while at 0.2% EMS stearic acid was synthesized, which was absent in the untreated callus. It was concluded that the callus of P. oleracea accumulated different fatty acids, and EMS treatment enhanced the content of a few fatty acids in the cultures.

Влияние биологически активных веществ на тепловой и окислительный стресс модельных объектов Caenorhabditis elegans

Modern medicine strives to prevent age-related diseases. Oxidative stress is associated with development and progression of various diseases. Reactive oxygen species are part of vital physiological processes. High levels of reactive oxygen lead to stress and pathology whereas low ones are associated with healthy physiology. Plant-derived adaptogens demonstrate good results in stress tolerance and homeostasis. Plant materials are a pharmacologically optimal source of chemical compounds to treat various diseases, including those caused by oxidative stress. The research featured biologically active substances isolated from extracts of callus, suspension, and root cultures of medicinal plants. Baicalin and trans-cinnamic acid were obtained from Scutellaria baicalensis while ursolic acid came from Thymus vulgaris. The biologically active substances were tested for neuroprotective properties, as well as for the impact on the expression of SOD-3 and HSP-16.2. Caenorhabditis elegans served as a model to study the accumulation of carbonylated proteins and lipofuscin. The neuroprotective activity of all tested substances decreased as their concentration fell from 200 to 10 μmol/L. C. elegans proved more resistant to thermal stress if pretreated with the biologically active substances. In response to thermal stress, nematodes expressed SOD-3 more actively than HSP-16.2. At 100 μmol/L, the biologically active substances could reduce the level of carbonylated proteins. Ursolic acid was especially effective against protein carbonylation and lipofuscin accumulation in all concentrations. Baicalin, trans-cinnamic acid, and ursolic acid made it possible to reduce oxidative and thermal stress, thus demonstrating good prospects for further studies as part of adaptogenic prepa rations.

Elicitation of callus cultures of the medicinally important plant Embelia ribes Burm f. using biotic and abiotic elicitors for enhanced production of embelin

Elicitation of callus cultures of the medicinally important plant Embelia ribes Burm f. using biotic and abiotic elicitors for enhanced production of embelin

The Influence of Water Deficit on Dehydrin Content in Callus Culture Cells of Scots Pine

Under a water deficit, the protective proteins known as dehydrins (DHNs) prevent nonspecific interactions in protein and membrane structures and their damage, in addition to playing an antioxidant role. The DHNs of a widespread xerophytic species Scots pine (Pinus sylvestris L.) have been poorly studied, and their role in resistance to water deficits has not been revealed. In this paper, we have expanded the list of DHNs that accumulate in the cells of Scots pine under the conditions of water deficits and revealed their relationship with the effects of water deficits. In this investigation, callus cultures of branches and buds of Scots pine were used. A weak water deficit was created by adding polyethylene glycol to the culture medium. Under the conditions of a water deficit, the activity of catalase and peroxidase enzymes increased in the callus cultures. A moderate decrease in the total water content was correlated with a decrease in the growth rate of the callus cultures, as well as with an increase in the activity of lipid peroxidation. The accumulation of Mr 72, 38, and 27 kDa DHNs occurred in the callus cultures of buds, and the accumulation of Mr 72 and 27 kDa DHNs positively correlated with the lipid peroxidation activity. An increase in the content of DHNs was observed in cultures that differed in origin, growth indicators, and biochemical parameters, indicating the universality of this reaction. Thus, previously undescribed DHNs were identified, the accumulation of which is caused by water deficiency and is associated with manifestations of oxidative stress in the kidney cells of Scots pine.

Enhancement of the Production of Tropane Alkaloids in the Hyoscyamus Niger L. Callus Using Different Biotic Elicitors

This study looked at the amount of tropane alkaloids in both the leaves of the mother plant and the callus cultures of Hyoscyamus niger L. It also looked at how biotic elicitation affects the growth of callus cultures and increases the production of alkaloids. The investigation identified and quantified three primary tropane alkaloids (Hyoscyamine, Scopolamine, and Atropine) from both sources. Remarkably, the callus cultures exhibited obviously higher levels of alkaloids in comparison to the leaves of the mother plant, implying their potential as a valuable reservoir for alkaloid production. Concerning the effects of biotic elicitation, the study showed that the application of chitosan (CHT), yeast extract (YE), and fungal extract of Trichoderma asperellum yielded a substantial decrease in callus weight when contrasted with the control group. This decrease became more pronounced with escalating concentrations of the elicitors. In terms of alkaloid synthesis, a clear correlation between the concentration of bio stimulants and tropane alkaloid levels was established. Particularly, CHT elicitation displayed the most pronounced enhancement in levels of tropane alkaloids among the three elicitors. Notably, the highest CHT concentration of 40 mg/L yielded the most elevated levels of alkaloids, measuring at 25.3 µg/g for Hyoscyamine, 31.2 µg/g for Scopolamine, and 21.5 µg/g for Atropine. This represents approximate percentage increases in concentration of 208%, 183%, and 198%, respectively, when compared to the control treatment. Generally, these findings carry significant implications for advancing tropane alkaloid biosynthesis, with potential applications spanning the pharmaceutical and biotechnology sectors.

Influence of Primary Light Exposure on the Morphophysiological Characteristics and Phenolic Compounds Accumulation of a Tea Callus Culture (Camellia sinensis L.).

Tea plant calli (Camellia sinensis L.) are characterized by the accumulation of various phenolic compounds (PC)-substances with high antioxidant activity. However, there is still no clarity on the response of tea cells to light exposure of varying intensity. The purpose of the research was to study tea callus cultures grown under the influence of primary exposure to different light intensities (50, 75, and 100 µmol·m-2·s-1). The cultures' growth, morphology, content of malondialdehyde and photosynthetic pigments (chlorophyll a and b), accumulation of various PC, including phenylpropanoids and flavanols, and the composition of catechins were analyzed. Primary exposure to different light intensities led to the formation of chloroplasts in tea calli, which was more pronounced at 100 µmol·m-2·s-1. Significant similarity in the growth dynamics of cultures, accumulation of pigments, and content of malondialdehyde and various phenolics in tea calli grown at light intensities of 50 and 75 µmol·m-2·s-1 has been established, which is not typical for calli grown at 100 µmol·m-2·s-1. According to data collected using high-performance liquid chromatography, (+)-catechin, (-)-epicatechin, epigallocatechin, gallocatechin gallate, epicatechin gallate, and epigallocatechin gallate were the main components of the tea callus culture's phenolic complex. Its content changed under the influence of primary exposure to light, reaching the greatest accumulation in the final stages of growth, and depended on the light intensity. The data obtained indicate changes in the morphophysiological and biochemical characteristics of tea callus cultures, including the accumulation of PC and their individual representatives under primary exposure to light exposure of varying intensity, which is most pronounced at its highest values (100 µmol·m-2·s-1).

Metabolome and Transcriptome Joint Analysis Reveals That Different Sucrose Levels Regulate the Production of Flavonoids and Stilbenes in Grape Callus Culture.

To reveal the effect of sucrose concentration on the production of secondary metabolites, a metabolome and transcriptome joint analysis was carried out using callus induced from grape variety Mio Red cambial meristematic cells. We identified 559 metabolites-mainly flavonoids, phenolic acids, and stilbenoids-as differential content metabolites (fold change ≥2 or ≤0.5) in at least one pairwise comparison of treatments with 7.5, 15, or 30 g/L sucrose in the growing media for 15 or 30 days (d). Resveratrol, viniferin, and amurensin contents were highest at 15 d of subculture; piceid, ampelopsin, and pterostilbene had higher contents at 30 d. A transcriptome analysis identified 1310 and 498 (at 15 d) and 1696 and 2211 (at 30 d) differentially expressed genes (DEGs; log2(fold change) ≥ 1, p < 0.05) in 7.5 vs. 15 g/L and 15 vs. 30 g/L sucrose treatments, respectively. In phenylpropane and isoflavone pathways, DEGs encoding cinnamic acid 4-hydroxylase, chalcone synthase, chalcone isomerase, and flavanone 3-hydroxylase were more highly expressed at 15 d than at 30 d, while other DEGs showed different regulation patterns corresponding to sucrose concentrations and cultivation times. For all three sucrose concentrations, the stilbene synthase (STS) gene exhibited significantly higher expression at 15 vs. 30 d, while two resveratrol O-methyltransferase (ROMT) genes related to pterostilbene synthesis showed significantly higher expression at 30 vs. 15 d. In addition, a total of 481 DEGs were annotated as transcription factors in pairwise comparisons; an integrative analysis suggested MYB59, WRKY20, and MADS8 as potential regulators responding to sucrose levels in flavonoid and stilbene biosynthesis in grape callus. Our results provide valuable information for high-efficiency production of flavonoids and stilbenes using grape callus.

Feasible biosynthesis of biologically active metabolites in in vitro culture of Macrotyloma uniflorum

Macrotyloma uniflorum, commonly called “Horse gram” is an underutilized pulse crop recognized for its great nutritional significance and a broad range of biological properties. There have been no in vitro studies for the biosynthesis of enhanced bioactive metabolites in the callus culture of M. uniflorum. In the study reported here, we have designed a feasible protocol for high-frequency callus induction and maintenance utilizing a leaf as an explant grown on MS media enriched with various concentrations of different plant growth regulators (PGRs) including α-naphthalene acetic acid (NAA), 6-benzylaminopurine (BAP) and thidiazuron (TDZ) either alone or in combination. Among all the tested PGRs, NAA alone resulted in high-frequency callus induction (97%), and maximum biomass accumulation (Fresh weight (FW): 200 g/L: Dry weight (DW): 20.4 g/L). Moreover, hormonally optimized callus cultures exhibit maximum production of phenolic compounds (166.6 mg/L) and flavonoid compounds (351.6 mg/L). The antioxidant potential of calli extracts was also determined by utilizing various antioxidant activities. Maximum antioxidant activities ((2,2′-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid)) ABTS = 497.9; TEAC μM; (Ferric reducing antioxidant power) FRAP = 823 TEAC μM; (Cellular Antioxidant Activity) CAA: 58.2%) were recorded in leaf-derived calli supplemented with different PGRs treatments. High-performance liquid chromatography (HPLC) analysis further revealed maximum biosynthesis of caffeic acid (1.63 mg/g DW), gallic acid (8.92 mg/g DW), kaempferol (0.71 mg/g DW), myricetin (0.39 mg/g DW), apigenin (0.64 mg/g DW) at 10 mg/L BAP and isorhamnetin (0.68 mg/g DW) at 1 mg/L TDZ + 10 mg/L NAA. The objective of this study was to explore in vitro biosynthesis of enhanced bioactive metabolites in the callus culture of M. uniflorum, leveraging a feasible protocol for high-frequency callus induction and maintenance. The results showcase the remarkable efficacy of NAA in callus induction and biomass accumulation, highlighting the hormonally optimized callus cultures as a potent source for enhanced biosynthesis of bioactive metabolites, paving the way for further applications in pharmaceutical and commercial industries.

Multi-omics signatures of diverse plant callus cultures

Multi-omics signatures of diverse plant callus cultures

Features of cellular selection of tomato plants, resistance against salining

Aim. For selection at the cellular level, nutrient media are offered, which, although not completely, would partially correspond to natural stress conditions. Methods. As explants, immature embryos, cotyledon nodes with meristematic activity, and hypocotyls of tomato plants of the Lagidnyi, Khoryv, Borivskyi. Results. An agar medium of general mineral composition with different concentrations of growth regulators was used for the cultivation of isolated tissues the generally accepted method. Conclusions. When creating in vitro new forms of tomato plants with increased resistance to chloride salinization of the soil, it is necessary to select resistant cell lines using the stepwise selection method according to the scheme, which includes several consecutive actions of a selective agent in different concentrations at the stages of induction – proliferation, morphogenesis of callus cultures. An increase in the content of hydrogen peroxide in callus structures and stimulation of the activity of antioxidant enzymes superoxide dismutase and peroxidase were also found.

Morphological, cytogenetic, and phytochemical estimation of Lychnis flos-cuculi L. cell cultures: can cell biomass be a source of bioactive compounds?

The ragged robin (Lychnis flos-cuculi L.) from the Caryophyllaceae family contains flavonoids, phenolic acids, flavonoids, triterpenoid saponins, and ecdysteroids, making it potential for biological and medicinal application. Plant cell culture technologies could be beneficial for obtaining low-cost biomass sources with valuable phytochemicals sustainably. The current study reports the induction and proliferation of ragged robin cell culture and evaluation of various plant growth regulators (PGRs) impact on the secondary metabolites profile using UPLC-HRMS/MS. Callus was initiated from different explants on the Murashige and Skoog medium with various combinations and concentrations of PGRs. It was found that prolonged proliferation on MS medium enriched with a combination of 1.0 mg l− 1 2,4-Dichlorophenoxyacetic acid and 0.1 mg l− 1 N6-furfuryl adenine (Kinetin) yields the maximum biomass with 504.05% growth index. The results of flow cytometric analysis suggested no genetic alteration between differentiated and dedifferentiated cultures. Furthermore, the UPLC-HRMS/MS result distinguished 101 significant signals that were manually identified as ecdysteroids (16 metabolites), flavonoids (11 metabolites), phenolic acids (10 metabolites), triterpene saponins and sapogenins (53 metabolites), and some primary metabolites. Our study demonstrates that the cell culture of L. flos-cuculi displays significant differences in its phytochemical makeup to plants cultivated in vitro. Moreover, the type of growth regulators used for callus cultures was identified to cause substantial shifts in the identified metabolites.Graphical

Genetic stability evaluation of caladium somaclonal variants by morphological, cytological, and SSR analysis in three successive generations

Somaclonal variants with valuable agronomic traits play a crucial role in crop breeding, provided that they are stably passed down in subsequent generations. Although numerous somaclonal variants have been identified and studied among in vitro regenerated plants in the foliage plant caladium (Caladium × hortulanum Birdsey), no research has been carried out to evaluate their genetic stability in subsequent generations. This study aimed to evaluate the genetic stability of three types of diploid caladium somaclonal variants, previously derived from in vitro callus cultures, over three successive tuber-propagated generations. The analysis of quantitative morphological characteristics revealed a greater degree of variation among the established plants in the first generation (G1) compared to the second (G2) and third generations (G3). Seven plants exhibiting distinct leaf coloration changes were identified in the G1 generation and were found to be stably passed down to the subsequent G2 and G3 generations. These findings indicate a wide range of morphological variation in the G1 generation, followed by relative stability in subsequent generations. A comprehensive cytological and molecular analysis revealed that five of the seven newly observed variants displayed notable differences in relative nuclear DNA content, chromosome number, or SSR (simple sequence repeat) banding patterns compared to their corresponding normal counterparts. The findings of this study will be instrumental in developing new cultivars with distinctive plant morphology through the exploitation of somaclonal variation in caladium.

Cytotoxic activity of callus extract from Vachellia farnesiana (L) Wight & Arn.

The in vitro cultures of Vachellia farnesiana (L) Wight & Arn. have demonstrated cytotoxic activity through callus extract on the HeLa cell line. Explants excised from in vitro-grown seedlings from seeds of two different locations were inoculated on Murashige and Skoog (MS) culture media containing various concentrations of N-6 benzyladenine (BA) or kinetin with 2,4-dichlorophenoxyacetic acid (2,4-D). Optimal efficiency in friable callus induction (100%) was achieved in leaf explants cultured on MS media containing 2.32µM BA + 13.57µM 2,4-D. Plant tissues (callus and leaf) were extracted and subjected to quantitative phytochemical analysis, revealing the highest total alkaloid and phenolic content in leaf extracts from Queretaro adult specimens (339.5 ± 20.9mg atropine equivalents (AE) per g dry extract (DE) and 158.4 ± 12.5mg gallic acid equivalents (GAE) per g DE, respectively). In contrast, callus cultures exhibited significantly higher total triterpene content (356-381mg ursolic acid equivalents (UAE) per g DE) compared to leaf extracts (208-243mg UAE/g DE). Both leaf and callus extracts displayed cytotoxic activity against the HeLa cell line, with a significantly lower half-maximal inhibitory concentration (IC50) for leaf extracts (28-32µg/mL) compared to callus cultures (43-66µg/mL), suggesting that alkaloids were primarily responsible for the cytotoxic activity. Furthermore, this study provides valuable insights into the controlled production of bioactive compounds with cytotoxic activity, with callus serving as a rich source.

Evaluation of phytochemical, antioxidative and antibacterial activities in Hypericum perforatum L. callus cultures

Evaluation of phytochemical, antioxidative and antibacterial activities in Hypericum perforatum L. callus cultures