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Melatonin improves plant resistance to multiple stresses by participating in the biosynthesis of metabolites. Flavonoids are an important family of plant secondary metabolites and are widely recognized to be involved in resistance; however, the crosstalk between melatonin and flavonoid is largely unknown. We found that the resistance of pigeon pea (Cajanus cajan) to salt, drought, and heat stresses were significantly enhanced by pre-treatment with melatonin. Combined transcriptome and LC-ESI-MS/MS metabolomics analyses showed that melatonin significantly induced the enrichment of flavonoids and mediated the reprogramming of biosynthetic pathway genes. The highest fold-increase in expression in response to melatonin treatment was observed for the CcF3´H family, which encodes an enzyme that catalyses the biosynthesis of luteolin, and the transcription factor CcPCL1 directly bonded to the CcF3´H-5 promoter to enhance its expression. In addition, salt stress also induced the expression of CcPCL1 and CcF3´H-5 , and their overexpression in transgenic plants greatly enhanced salt tolerance by promoting the biosynthesis of luteolin. Overall, our results indicated that pre-treatment of pigeon pea with melatonin promoted luteolin biosynthesis through the CcPCL1 and CcF3´H-5 pathways, resulting in salt tolerance. Our study shows that melatonin enhances plant tolerance to multiple stresses by mediating flavonoid biosynthesis, providing new avenues for studying the crosstalk between melatonin and flavonoids. [ABSTRACT FROM AUTHOR]

The present study was designed to evaluate the chemical extraction, chemical composition, and antioxidant and antibacterial properties of the total flavonoids in Willow Buds (TFW). We investigated the optimal extraction of TFW using response surface methodology (RSM). Chemical compounds were analyzed using Q-Orbitrap LC–MS/MS. The DPPH radical scavenging capacity, hydroxy radical inhibitory ability, and superoxide anion radical inhibitory ability were explored to determine the antioxidant properties of flavonoid extractions. The antibacterial effect was assessed via minimal inhibitory concentration. The results demonstrated that the optimal extraction conditions were an ethanol concentration of 50%, a time of 35 min, and a liquid/material ratio of 70:1 mL/g. Under these conditions, the yield of TFW was 7.57%. Eight flavonoids, a phenolic glycoside, and an alkaloid were enriched in the Willow Buds. The TFW exhibited significant antioxidant activity, with IC50 values of 0.18-0.24 mg/mL and antimicrobial activity against Escherichia coli, Salmonella enterica, Staphylococcus aureus, and Streptococcus pneumoniae. TFW may be explored as potential and natural compounds in food and pharmacological applications. [ABSTRACT FROM AUTHOR]

Artemisia absinthium L. is a highly medicinal plant with a broad range of biomedical applications. A. absinthium callus cultures were established in response to bio-fabricated single NPs (Ag and Cu) or a combination of both NPs (Ag and Cu) in different ratios (1:2, 2:1, 1:3, and 3:1) along with thidiazuron (TDZ) (4 mg/L) to elicit the biomass accumulation, production of non-enzymatic compounds, antioxidative enzymes, and antioxidant activity. Silver and copper nanoparticles (Ag and Cu NPs) were synthesized using the leaves of Moringa oleifera as reducing and capping agent and further characterized through UV-Visible spectroscopy and SEM. The 30 µg/L suspension of Ag and Cu NPs (1:2, 2:1) and 4 mg/L TDZ showed 100% biomass accumulation as compared to control (86%). TDZ in combination with Ag NPs enhanced biomass in the log phases of growth kinetics. The Cu NPs alone enhanced the superoxide dismutase activity (0.56 nM/min/mg FW) and peroxidase activity (0.31 nM/min/mg FW) in callus cultures. However, the combination of Ag and Cu NPs with TDZ induced significant total phenolic (7.31 µg/g DW) and flavonoid contents (9.27 µg/g DW). Furthermore, the antioxidant activity was highest (86%) in the Ag and Cu NPs (3:1) augmented media. The present study provides the first evidence of bio-fabricated single NPs (Ag and Cu) or a combination of both NPs (Ag and Cu) in different ratios (1:2, 2:1, 1:3, and 3:1) along with TDZ (4 mg/L) on the development of callus culture, production of endogenous enzymes, non-enzymatic components, and further antioxidant activity in callus cultures of A. absinthium. [ABSTRACT FROM AUTHOR]

SUMMARY: Calcineurin B‐like (CBL)‐interacting protein kinases (CIPKs) play a central role in Ca2+ signalling and promote drought tolerance in plants. The CIPK gene family in pigeon pea (Cajanus cajan L.), a major food crop affected by drought, has not previously been characterised. Here, we identified 28 CIPK genes in the pigeon pea genome. Five CcCIPK genes were strongly upregulated in roots upon drought treatment and were selected for further characterisation. Overexpression of CcCIPK13 and CcCIPK14 increased survival rates by two‐ to three‐fold relative to controls after 14 days of drought. Furthermore, the three major flavonoids, genistin, genistein and apigenin, were significantly upregulated in the same transgenic plants. Using CcCIPK14 as bait, we performed a yeast two‐hybrid screen and identified six interactors, including CcCBL1. CcCIPK14 exhibited autophosphorylation and phosphorylation of CcCBL1 in vitro. CcCBL1‐overexpressed plants displayed higher survival rates upon drought stress as well as higher expression of flavonoid biosynthetic genes and flavonoid content. CcCIPK14‐overexpressed plants in which CcCBL1 transcript levels were reduced by RNA interference had lower survival rates, which indicated CcCBL1 in the same pathway as CcCIPK14. Together, our results demonstrate a role for the CcCIPK14‐CcCBL1 complex in drought stress tolerance through the regulation of flavonoid biosynthesis in pigeon pea. Significance Statement: This study systematically elucidates the mechanism by which the CIPK‐CBL complex regulates flavonoid accumulation and drought resistance in pigeon pea, which lays a foundation for in‐depth exploration of relevant molecular mechanisms in plants. [ABSTRACT FROM AUTHOR]

Fig (Ficus carica L.) leaves are produced each year and often disposed, resulting in a waste of resources. Fig waste leaves are rich in flavonoids, which have strong antioxidant activity; however, the variety and chemical structure of antioxidants in fig leaves have not been reported in detail. To take full advantage of fig waste leaves, antioxidant capacity of different extracts (petroleum ether, ethyl acetate, and water) was evaluated by 1, 1-diphenyl-2-picrylhydrazyl (DPPH), 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic) acid (ABTS), and ferric-ion-reducing antioxidant power (FRAP) methods. The results showed that flavonoids in ethyl acetate extraction had the highest content (83.92 ± 0.01 mg/g), maximum DPPH scavenging activity (IC50 0.54 mg/mL), highest ABTS scavenging rate (80.28%), and FRAP (3.46 mmol/g). Furthermore, an HPLC-QTOF-MS/MS-DPPH method was developed to identify 11 flavonoids in fig waste leaves. This rapid and efficient method can not only be used for screening the antioxidant components in fig waste leaves, but also can be combined with mass spectrometry to identify the compounds with antioxidant capacity. There are three flavonoids with significant antioxidant capacity, which are 3-O-(rhamnopyranosyl-glucopyranosyl)-7-O-(glucopyranosyl)-quercetin, isoschaftoside, and rutin. The results confirmed that fig waste leaves contain a variety of antioxidant components, which contributed to increase the value of fig waste leaves as antioxidants. [ABSTRACT FROM AUTHOR]

Flavonoids are important secondary metabolites in the plant growth and development process. As a medicinal plant, pigeon pea is rich in secondary metabolites. As a flavonoid, there are few studies on the regulation mechanism of naringenin in plant stress resistance. In our study, we found that naringenin can increase the pigeon pea's ability to tolerate salt and influence the changes that occur in flavonoids including naringenin, genistein and biochanin A. We analyzed the transcriptome data after 1 mM naringenin treatment, and identified a total of 13083 differentially expressed genes. By analyzing the metabolic pathways of these differentially expressed genes, we found that these differentially expressed genes were enriched in the metabolic pathways of phenylpropanoid biosynthesis, starch and sucrose metabolism and so on. We focused on the analysis of flavonoid biosynthesis related pathways. Among them, the expression levels of enzyme genes CcIFS , CcCHI and CcCHS in the flavonoid biosynthesis pathway had considerably higher expression levels. By counting the number of transcription factors and the binding sites on the promoter of the enzyme gene, we screened the transcription factors CcMYB62 and CcbHLH35 related to flavonoid metabolism. Among them, CcMYB6 2 has a higher expression level than the others. The hairy root transgene showed that CcMYB62 could induce the upregulation of CcCHI , and promote the accumulation of naringenin, genistein and biochanin A. Our study revealed the molecular mechanism of naringenin regulating flavonoid biosynthesis under salt stress in pigeon pea, and provided an idea for the role of flavonoids in plant resistance to abiotic stresses. • Naringenin enhanced the salt tolerance of pigeon pea. • Naringenin can induce the accumulation of downstream secondary metabolites. • Naringenin induced significant upregulation of CcCHS , CcCHI and CcIFS. • Revealing the potential regulation between CcMYB62 and flavonoid biosynthesis. [ABSTRACT FROM AUTHOR]

In this study, different types of natural deep eutectic solvents (NADES) were used to extract flavonoids from sweet tea. A total of 7 flavonoids, primarily dihydrochalcones, were successfully extracted. Among the NADESs tested, a ternary deep eutectic solvent Bet-PG-Ur (betaine: propanediol: urea = 1: 2: 1, molar ratio) demonstrated the highest extraction efficiency. The chemical and morphological changes of both the NADES and extracted compounds were analyzed using Fourier transform infrared spectroscopy (FT-IR) and scanning electron microscopy (SEM). To optimize the extraction process and evaluate the total flavonoid yield, response surface methodology (RSM), artificial neural network (ANN), and random forest (RF) techniques were employed. The NADES demonstrated consistent extraction performance even after three cycles of recovery using the SPE method. The results revealed that all three models successfully predicted the TFC in the green extraction ultrasonic process. The ANN model exhibited superior result consistency compared to the others, which can be attributed to the enhanced statistical parameters obtained. The highest total flavonoid content (TFC) of 167.561 mg/g was predicted by ANN and achieved under the following conditions: liquid/solid ratio of 19, water content of 50 %, temperature of 45 °C, and ultrasonic power of 400 W. [Display omitted] • A ternary DES-ultrasonic assisted was used for extraction of 7 flavonoids from sweet tea. • For the first time, machine learning was used to forecast the parameters for the TFC in sweet tea. • In terms of process modeling and prediction, ANN is better than RF and RSM. • The extraction efficiency of DES can reach 80% after 3 cycles of recovery by SPE. • Ultrasound-assisted extraction of 7 flavonoids from sweet tea using NADESs. [ABSTRACT FROM AUTHOR]

A novel approach for synchronous extraction and enrichment of hydrophilic component and hydrophobic component with CO 2 -based switchable-solvent (SHS) from pigeon pea leaves was developed in the present study. After screening of SHS, N, N-dimethyl-cyclohexylamine (DMCHA)/water system was selected as the properest SHS for the extraction and enrichment process. To optimize this process, we conducted single factor experiments with the Box-Behnken design. The optimal parameters were as follows: solid-liquid ratio R (S/L) 40:1 (mg/mL), ultrasound time 15 min and bubbling CO 2 time 79 min (0.5 mL/s). Under the circumstances, the extraction yields of representative flavonoid vitexin and stilbene cajaninstilbenes acid (CSA) reached 1.80 ± 0.12 mg/g and 9.32 ± 0.08 mg/g, respectively, which were 1.06 and 1.49 folds to those by the traditional methods. And the contents of target compounds vitexin and CSA in the enriched extracts were 2.3% and 12.1%, which increased above 6.76 and 4.58 folds to that in extracts using 65% ethanol-UAE. And the results suggested that the N, N-dimethyl-cyclohexylamine (DMCHA)/water system can be guaranteed for at least 5 cycles with relatively stable extraction yields of vitexin and CSA. Furthermore, a theoretical study using density functional theory was put into effect to explore the properties of SHS and their relationship with the vitexin and CSA. The mechanism study showed that SHS had higher biocompatibility with target compounds due to the H-bonds. The present method was an eco-friendly, promising, and sustainable alternative to extract and enrich simultaneously hydrophilic and hydrophobic phytochemicals from plant materials. Image 1 • This paper developed a novel approach for efficient and recyclable extraction and enrichment of phytochemicals from the disused pigeon pea leaves in one solvent system. • The mechanism study showed that SHS had higher biocompatibility with target compounds due to the H-bonds using the density functional theory. • The switchable solvents have usually been used in the extraction of oil. Moreover, the exploit of SHS considering bioactive compounds from plants material has hardly been reported yet. [ABSTRACT FROM AUTHOR]