HPLC DAD Research Articles

Biostimulant effect of Cladophora glomerata extract on garden cress plant growth

Biostimulants that improve agriculture's productivity and environmental responsibility are widely favored and used. Algae has emerged as a viable option for sustainable agriculture. In the present study, the effects of the Cladophora glomerata (L.) Kütz. aqueous extract in three concentrations (2.5, 5, and 10 mg extract per mL of tap water) was tested on the growth and productivity of garden cress (GCR) Lepidium sativum L., plants over 12 days, and the results were compared with those of the control group (irrigated only with tap water). The effect of the algae extract was studied by assessing the changes in phenolic compounds using HPLC–DAD and determining the chlorophyll content as well as the total antioxidant capacity of the GCR using DPPH and ABTS tests. Total phenolics, as well as total flavonoids, were measured. The potential role of algae extract in promoting cress plant growth was attained when compared to the control, mainly at a concentration of 2.5 mg/mL, which exhibited the highest yield growth after 12 days and presented the highest antioxidant capacity at 13.53 ± 1.16 mg Trolox equivalent (TE)/g dry weight (DW), compared to 10.44 ± 0.33 mgTE/g DW for the control. The total phenolic content significantly increased (p < 0.05) from 14.34 ± 0.84 mg gallic acid equivalent (GAE)/ g dry weight (DW) to 23.14 ± 0.55 mg GAE/g DW. Eleven phenolic compounds were identified in different tested samples of GCR, whether treated or not treated with algae extracts. Chrysin was only identified in the treated plants. Therefore, the use of algae (C. glomerata) presents promising potential as a biostimulant in agriculture, contributing to increased plant growth and improved resistance to environmental stress.

A novel stability-indicating chromatographic quantification of the antiparkinsonian drug safinamide in its pharmaceutical formulation employing HPTLC densitometry and ion-pair HPLC–DAD

Parkinson's disease (PD) emerges as a notable health concern among the elderly population. Safinamide mesylate (SAF) is a novel and emerging add-on therapy in PD treatment. The stability of innovative drug formulations and the development of appropriate stability-indicating methods are of great importance to modern pharmaceutical analysis. The current work has established novel comprehensive stability-indicating chromatographic approaches, HPTLC coupled with densitometric quantification and HPLC–DAD, for the selective assay of SAF in pharmaceutical formulation along with its synthetic precursor impurity; 4-hydroxy benzaldehyde (4-HBD) in presence of its stress induced degradation products. The stability of SAF was investigated under different stress conditions. It was found that SAF is likely to undergo acid, base hydrolysis, and oxidative degradation. Using mass spectrometry and infrared spectroscopy, the structures of the forced degradation products were confirmed and elucidated. The dissolution behavior of Parkimedine® Tablets was also monitored in the FDA suitable medium. Multiple assessment tools were used to evaluate the environmental sustainability of the proposed methods and the reported one. The greenness tools included Complex-GAPI and AGREE metrics. In addition, the innovative concepts of "blueness" and "whiteness" evaluation were incorporated through the newly introduced BAGI and RGB12 algorithms, respectively.

Determination of phytochemical quality of leaves Vaccinium vitis-idaea L. and Vaccinium myrtillus L. from the polluted and non-polluted areas

Elemental composition of Vaccinium myrtillus L. and Vaccinium vitis-idaea L. has not been determined yet in detail. In our study, a scanning electron microscope coupled with an energy dispersive X-ray analyser (SEM–EDX) ensured the determination of 15 elements in the leaves of Vaccinium sp. growing in the control and the mine polluted locality. The soil elemental analyses showed a higher content of 11 elements from all determined in the mine-influenced samples. Elemental analyses of the control leaves showed the highest contents of all determined elements for V. vitis-idaea, except for carbon. The impact of pollution on V. myrtillus leaves caused significant increase in oxygen, natrium, magnesium, sulphur, chlorine, potassium and calcium content. The contents of carbon, nitrogen and silicon decreased substantially. In the case of the V. vitis-idaea leaves, the content of most of elements reduced, and in the case of six elements, even significantly. A significant increase was recorded for carbon, iron and copper. The effect of the environment has influenced the production of phytochemicals, assessed by HPLC–DAD. The production of flavonoids (hyperoside, isoquercetin) increased significantly in V. myrtillus and decreased to a detectable minimum in V. vitis-idaea. In the case of chlorogenic acid, arbutin and hydroquinone, their levels changed minimally in V. myrtillus, but significantly more in the V. vitis-idaea leaves. The importance of elemental analyses lies in detecting the presence of toxic elements or their harmful concentrations in plants that are the source of food or dietary supplements.

A new Fe3O4-MWCNTs-reinforced hollow fiber solid/liquid phase microextraction-based natural deep eutectic solvent for determination of trace phthalate esters in aqueous samples using HPLC–DAD

A new Fe3O4-MWCNTs-reinforced hollow fiber solid/liquid phase microextraction-based natural deep eutectic solvent for determination of trace phthalate esters in aqueous samples using HPLC–DAD

Mathematical Modelling and Chemical Analysis to Characterise Anthocyanin Self-association Interactions Influencing Colour Expression and Stability in Young Red Wines

AbstractAnthocyanins are phenolic compounds that provide colour to young red wines following extraction from grape skins, and their reaction kinetics are not well understood as they exist within a complex pH-dependent multistate system. For commercial wineries to best control anthocyanin colour expression that is a major determinant for red wine quality, there is a critical need for behaviours of all pH-dependent monomeric and self-associated anthocyanins to be considered together. In response, the current study employed mathematical and experimental techniques to reveal kinetic and steady-state behaviours of all species of a model anthocyanin, malvidin-3-O-β-D-glucopyranoside (M3G), within three Shiraz red wines throughout fermentation. Investigator-developed models represented the following: (i) acid–base reactions that form red, purple, and blue anthocyanin monomers; (ii) hydration reactions that result in monomeric colour loss; and (iii.) physical self-association interactions that temporally stabilise anthocyanin colour. Simulations were validated experimentally using high-performance liquid chromatography with diode array detector (HPLC–DAD) and colourimetric analysis of wine samples. Moreover, a unique predictive modelling framework was developed that takes in measured values for anthocyanin concentration, pH, and temperature at the onset of fermentation, and output data describing corresponding wine colour expression and stability characteristics that would arise after fermentation has progressed. Results of the current study elucidate complex reaction kinetics occurring between anthocyanin species under dynamic red wine conditions and provide meaningful predictions for future wine colour and shelf-life characteristics. Outputs may be used to inform key winemaking decisions influencing red wine quality and to optimise the use of winery resources throughout the fermentation process.

Phytochemical Analysis and Antioxidant, Antimicrobial, and Antibiofilm Effects of a New Himalayan Lichen Placidium deosaiense Usman and Khalid Growing in Pakistan

Phytochemical composition and antimicrobial, antibiofilm, and antioxidant effects of a newly described Himalayan lichen Placidium deosaiense Usman and Khalid growing in Pakistan were investigated. HPLC–DAD methods were used for identification of secondary metabolites in acetone and methanol extracts. The total phenolics content was measured using a spectrophotometric method. The study investigated the antioxidant (DPPH-scavenging activity assay and reducing-power assay), antibacterial (minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC)), and antibiofilm (inhibition of biofilm formation and reduction in mature biofilm) activities of extracts of the lichen P. deosaiense and isolated parietin. The chemical constituents olivetol, olivetolic acid, haematommic acid, fallacinol, and parietin were identified as major compounds in the tested extracts of the lichen. Parietin was isolated from the acetone extract on a separation column. The methanol extract had higher values of TPC (21.67 mg GAE/g) than the acetone extract. Isolated parietin showed the best antioxidant activity measures, according to the DPPH-scavenging activity assay (IC50 = 51.616 μg/mL) and reducing-power assay. Although the extracts showed the best antibacterial activity (especially against Proteus mirabilis ATCC 12453), parietin demonstrated superior antibiofilm activity (especially against Staphylococcus aureus ATCC 25923). This is the first report on the phytochemical composition of the lichen Placidium deosaiense and the first description of the chemical composition of some of the 45 species of the genus Placidium. This research will pave the way for further exploration of new activities of this lichen and its metabolites, which are important for medicine and pharmacy.

Valorization of Pineapple Core Waste for Sequential Extraction of Phenolic Compounds and Carotenoids: Optimization Through Ultrasound-Assisted Method and Box–Behnken Design

AbstractIn this study, a pioneering cascade method involving ultrasound-assisted extraction (UAE) and Box–Behnken Design (BBD) was optimized to valorize pineapple core waste by the sequential extraction of, firstly phenolic compounds and, secondly, carotenoids. The effectiveness of the extraction was evaluated based on total polyphenol content (TPC) and antioxidant activity using 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and ferric reducing antioxidant power (FRAP) assays. Characterization of the carotenoids was performed using high-performance liquid chromatography with a diode array detector (HPLC–DAD). The initial characterization of dry pineapple core (DPC) samples revealed their nutritional composition, including protein, lipid, and carbohydrate weight percentages of 1.20 ± 0.05%, 5.3 ± 0.4%, and 88.6 ± 0.5%, respectively. The high extractives content (40.0 ± 4.5%) suggests a substantial presence of phenolic compounds, making the pineapple core a valuable source of natural antioxidants. The optimal UAE conditions for phenolic compound extraction were 70% amplitude, 5 min extraction time, and 2 cycles, yielding an antioxidant extract rich in phenolic compounds with a desirability value of 81.2%. Therefore, DPC was considered a valuable source of natural antioxidants. The extraction of β-carotene also showed promising results with optimal UAE conditions of 20% amplitude, 3 min extraction time, and 2 cycles. This research promotes the sustainable use of pineapple waste and demonstrates the potential to obtain valuable additives for the food, pharmaceutical, and cosmetic industries, encouraging a more circular and efficient use of resources in the pineapple processing industry.

Polycyclic aromatic hydrocarbons (PAHs) in black crusts on stone monuments in Milan: detection, quantification, distributions, and source assessment

In the field of conservation of cultural heritage, one must always consider the environmental conditions in which the works of art are located and the level of atmospheric pollution to which they are exposed, especially in the case of monuments stored outdoors. The present study is focused on the detection and the quantification of polycyclic aromatic hydrocarbons (PAHs) in black crust samples from the Monumental Cemetery of Milan (Italy), and the assessment of their sources through the analysis of the distributions of the different compounds in the samples, together with the use of diagnostic ratios. Six black crust samples taken from funerary monuments were analyzed. Fourteen polycyclic aromatic hydrocarbons were identified (naphthalene, acenaphthylene, acenaphthene, fluorene, phenanthrene, anthracene, fluoranthene, pyrene, chrysene, benzo[a]anthracene, benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene, indeno[1,2,3-cd]pyrene) by high-performance liquid chromatography with a diode-array detector (HPLC–DAD), with a total concentration from 0.72 to 3.81 μg/g (mean of 1.87 μg/g). The known carcinogenic benzo[a]pyrene accounted for 5–10% of the total polycyclic aromatic hydrocarbons in the samples analyzed, with concentrations up to 0.20 μg/g. Moreover, the study of the distribution and diagnostic ratios allowed us to confirm that anthropogenic sources such as traffic and the proximity of the train station are the major causes of the degradation of the monuments contained in this Cemetery.

Analysis of antioxidant nutrients, anti-HIV and anticancer metabolic fingerprints of Pelargonium quercifolium (L.f) L'Hér

Pelargonium quercifolium is an aromatic herb with a strong resinous scent. It is used traditionally to treat heart disease, hypertension, stress, rheumatism, combat hidden hunger, and inhibit HIV-1. The present study aimed to evaluate the toxicity, anti-HIV, antioxidant nutrients and anticancer potentials of the ethanol extract of P. quercifolium. In this study, high-resolution ultra-performance liquid chromatography-mass spectrometry (UPLC-MS), high-performance liquid chromatography with diode-array detection (HPLC-D AD), xCELLigence real-time cell analyser single plate (RTCA-SP), MTT assays and luciferase-based assay were employed to elucidate the phytochemicals, cytotoxicity, anti-HIV, antioxidant nutrients and anticancer metabolites in P. quercifolium. Results show that the characterized metabolites in P. quercifolium most of which were flavonoids, phenolic derivatives, alkaloids and coumarins could arrest cancer cell proliferation and inhibit HIV infection given the high level of cytotoxicity of the tested plant extract. It was also observed and reported for the first time, the nutrient profile of P. quercifolium is higher than most vegetables consumed as a staple food. This study underpins the potential of using anticancer and antiviral metabolites from leaf extract of P. quercifolium as a pharmacological option.

A sustainable HPLC method coupled with diode array detection for versatile quantification of telmisartan, chlorthalidone and amlodipine in a fixed-dose antihypertensive formulation and dissolution studies

Cardiovascular diseases, especially hypertension, stand as prominent contributors to global mortality. Hypertension, often referred to as a silent killer syndrome, necessitates the use of multiple medications for effective control and management. A new environmentally friendly HPLC–DAD method is introduced in this study for the concurrent analysis of telmisartan (TEL), chlorthalidone (CHT) and amlodipine besylate (AML), in both pure forms and combined pharmaceutical dosage form. An isocratic elution mode was employed to achieve chromatographic separation, utilizing an Inertsil C18 column (250 × 4.6 mm, 5.0 µm) and a mobile phase mixture of acetonitrile and phosphate buffer (pH 3.0 ± 0.1) with ratio of 35:65, v/v. The separation was achieved within 10 min at a flow rate of 1.0 mL/min. The proposed method's validation was carried out following the guidelines outlined by the International Council for Harmonisation (ICH). The achieved linearity range was 1.0–140.0 μg/mL for TEL and 1.0–100.0 μg/mL for CHT and AML with quantification limits of 0.061, 0.177, and 0.313 μg/mL for TEL, CHT, and AML, respectively. The fixed combination tablet dosage form demonstrated acceptable release profile, as indicated by the in-vitro dissolution studies. The studied dissolution media were phosphate buffer pH 7.5, 0.01 N HCl, and water, utilizing a USP type II apparatus at 37 ± 0.5 °C with a stirring rate of 75 rpm. The proposed method was applied successfully for the quality assessment of Telma-ACT® Tablets with good precision and accuracy. Various tools were used for evaluating the level of greenness, including Green Analytical Procedure Index (GAPI), Analytical Greenness Metric for Sample Preparation (AGREEprep), Analytical Eco-Scale (AES), and Analytical Method Greenness Score (AMGS). These tools had confirmed the eco-friendliness of the proposed method. Additionally, the newly introduced White Analytical Chemistry (WAC), and the Blue Applicability Grade Index (BAGI) have been specifically developed to evaluate the sustainability and the applicability of the method.

Effect of salt stress on growth and phenolic compounds production in callus suspension culture of the dioecious species thyrse sorrel (Rumex thyrsiflorus Fingerh.)

The sex-dependent differences in the response to salt stress of the dioecious Rumex thyrsiflorus and the influence of different concentrations of sodium chloride (NaCl) on the biosynthesis of phenolic compounds in callus suspension cultures were evaluated. The cultures originated from callus obtained on hypocotyls isolated from male and female seedlings were cultured on Murashige and Skoog medium supplemented with 0.4 mg/L 6-benzylaminopurine and 1 mg/L 2,4-dichlorophenoxyacetic acid. The results showed that the response to salt stress depends on both the sex of the plant and the NaCl concentration in the culture medium. The analysis of the main morphometric parameters showed that callus tissue derived from the hypocotyls of female seedlings was less sensitive to salt stress than that from male seedlings, which was correlated with higher concentrations of phenolic compounds. The optimal NaCl concentration to increase the production of phenolic compounds was 129 mM for females and 43 mM for males. In the methanolic tissue extracts 22 compounds were determined using the HPLC–DAD method. In general, higher amounts of compounds were detected in the extracts from the female tissue. The following dominated: catechin (max. 213.31), cryptochlorogenic acid (max. 76.35) and epicatechin (max. 54.84) (mg/100 g DW). This comprehensive phytochemical analysis of the sex-related aspects of the response to salt stress was performed for the first time in this dioecious model species. The results revealed potential application of NaCl as the ecological friendly and inexpensive elicitor to increase the production of pharmaceutically valuable compounds and highlight the importance of dioecy in phytochemistry.

Method development and validation of phloroglucinol and dieckol in Ecklonia cava using HPLC–DAD

Method development and validation of phloroglucinol and dieckol in Ecklonia cava using HPLC–DAD

QuEChERS Method for Simultaneous Analysis of Tetracyclines from Chicken by HPLC–DAD

QuEChERS Method for Simultaneous Analysis of Tetracyclines from Chicken by HPLC–DAD

The Production of Useful Phenol Compounds with Antioxidant Potential in Gametophytes and Sporophytes from In Vitro Cultures in Four Ornamental Ferns Species

Ferns are vascular plants with significant ornamental value, and in recent years, they have also been considered for the potential application in different economic sectors like nutrition, medicine and cosmetics. This study aimed to obtain in vitro fern gametophytes and sporophytes and evaluate their potential as secondary metabolites sources. For this study four fern species were used: Polypodium vulgare L. (PV), Asplenium trichomanes L. (AT), Athyrium filix-femina L. Roth (AF), and Osmunda regalis L. (OR). The polyphenols, flavonoids, tannins and terpenoids content and antioxidant activity were estimated by different colorimetric methods. For metabolite identification and their antioxidant activity, HP-TLC separations were used. Also, quantitative HPLC–DAD analysis was performed to estimate the content of certain metabolites. The results showed that in vitro generation of gametophytes registered higher growth rates in OR, PV, and AF, while the regeneration of sporophytes was better for PV, AT, and AF. The OR gametophyte produced the highest quantities in all analyzed metabolite classes and also had the highest antioxidant values. We report for the first time the presence of metabolites such as catechin, caffeic acid, gallic acid, ferulic acid, cinnamic acid, or rutin in OR. In this study, we demonstrated the possibility of producing potent antioxidant metabolites by in vitro cultures in ferns.

Extraction optimization and reactivity of 7α-acetoxy-6β-hydroxyroyleanone and ability of its derivatives to modulate PKC isoforms

Protein kinase C is a family of kinases that play important roles in carcinogenesis. Medicinal plants from Plectranthus spp. (Lamiaceae) are a well-known source of interesting abietanes, such as 7α-acetoxy-6β-hydroxyroyleanone (Roy). This study aimed to extract and isolate Roy from P. grandidentatus Gürke, comparing two extraction methods (CO2 supercritical and ultrasound-assisted acetonic extraction), and design new royleanone derivatives for PKC modulation focusing on breast cancer therapy. The concentration of Roy in the extracts was determined by HPLC–DAD. The supercritical extraction method yielded 3.6% w/w, with the presence of 42.7 μg mg−1 of Roy (yield of 0.13%), while ultrasound-assisted acetonic extraction yielded 2.3% w/w, with the presence of 55.2 μg mg−1 of Roy (yield of 0.15%). The reactivity of Roy was investigated aiming at synthetizing new ester derivatives through standard benzoylation and esterification reactions. The benzoylated (Roy-12-Bz) and acetylated (Roy-12-Ac) derivatives in the C12 position were consistently prepared with overall good yields (33–86%). These results indicate the 12-OH position as the most reactive for esterification, affording derivatives under mild conditions. The reported di-benzoylated (RoyBz) and di-acetylated (RoyAc) derivatives were also synthesized after increasing the temperature (50 °C), reaction time, and using an excess of reagents. The cytotoxic potential of Roy and its derivatives was assessed against breast cancer cell lines, with RoyBz emerging as the most promising compound. Derivatization at position C-12 did not offer advantages over di-esterification at positions C-12 and C-6 or over the parent compound Roy and the presence of aromatic groups favored cytotoxicity. Evaluation of royleanones as PKC-α, βI, δ, ε, and ζ activators revealed DeRoy’s efficacy across all isoforms, while RoyPr showed promising activation of PKC-δ but not PKC-ζ, highlighting the influence of slight structural changes on isoform selectivity. Molecular docking analysis emphasized the importance of microenvironmental factors in isoform specificity, underscoring the complexity of PKC modulation and the need for further exploration.

The effect of extraction conditions on the chemical profile of obtained raw poplar propolis extract

Various conditions of extraction were applied to obtain active extracts of raw poplar-type propolis. The extraction efficiency of traditional maceration was evaluated in terms of used solvent: ethanol (70 and 96% v/v), acetone (pure and 70%), propylene glycol, glycerol (50%), water and water with extraction modifiers: PEG 400 and lecithin. For obtained extracts, the total amounts of extracted phenolics and flavonoids as well as antioxidant activity were compared. For the most active extracts, the profile of volatile organic compounds with the use of GC × GC–MS and selected polyphenols content by HPLC–DAD was compared. To increase the activity of water propolis, extract ultrasound and microwave-assisted extraction were applied and obtained water extracts were compared regarding the main polyphenolic compounds content quantified by HPLC method. The recovery of 70% ethanolic extraction and the effect of the extension of extraction time were also examined by the HPTLC method. Based on conducted analyses, 70% ethanolic extract was found as the most aromatic and bioactive, followed by pure acetone and polypropylene glycol extracts. Compared to the classic maceration, water extraction assisted by microwaves and ultrasounds did not provide a higher extraction yield. In the case of 70% ethanolic extraction, the 5-day duration is recommended which allows to recovery of close to 80% of bioactive components of raw propolis.

Comparative Study of the Phytochemical Profile and Biological Activity of Ajuga reptans L. Leaf and Root Extracts

In this work, the phytochemical composition and the biological activity of the ethanolic extracts obtained from Ajuga reptans L. (Lamiaceae) leaves and roots (growing in Lesser Poland Voivodeship, Poland) were compared. The phytochemical composition of the extracts were determined by the high-performance liquid chromatography with diode-array detection (HPLC–DAD) method. The dominant compounds in both extracts were verbacoside, isoverbacoside, 3,4-dihydroxyphenylacetic acid and rosmarinic acid. The antioxidant capacity of the extracts was evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH), ferric reducing antioxidant power (FRAP) and 2,2’-azobis(3-ethylbenzothiazoline-6-sulfonate (ABTS) tests. The Folin–Ciocalteu assay was used to determine the total polyphenolic content. Additionally, enzyme (tyrosinase, collagenase) inhibition tests and metal chelating ability were studied to assess the anti-aging properties of the extracts. Moreover, the A. reptans extracts’ capacity to absorb the whole range of ultraviolet radiation and high-energy visible (HEV) light was evaluated. The skin irritation test (SIT) EpiDerm was applied to evaluate the safety of the bugle extracts. The noteworthy point is that there is a lack of literature on the assessment of A. reptans root extract activity. Our study is the only one that compares the quantitative composition and biological activity of extracts from the root and leaves of A. reptans. The obtained results indicated that both of the extracts exhibit high antioxidant, chelating and photoprotective activity, but the extract from A. reptans roots showed a higher enzyme inhibition effect for mature skin. The A. reptans root extract, similarly to the leaf extract, could be applied as potentially multi-functional, safe and sensitive cosmetic raw materials, especially in anti-aging and anti-pollution cosmetics.

Improved Antioxidant Properties and Vitamin C and B12 Content from Enrichment of Kombucha with Jujube (Ziziphus jujuba Mill.) Powder

(1) Objectives: Traditional kombucha (K) is a fermented beverage obtained from black or green tea infusion. Besides traditional substrates, the possibility of using alternative ingredients resulted in changes in metabolic profile and biological activity. The aim of this work was to study an alternative kombucha (KJ) prepared by the addition of jujube powder to black tea. (2) Materials and Methods: Changes in pH, protein, sugars, phenolic (TPC), flavonoid (TFC), and vitamin C and B12 content were evaluated at different time points over a period of 45 days. The identification of polyphenols by HPLC DAD and the antioxidant capacity by DPPH, ABTS, and FRAP tests of all samples was also carried out. (3) Results: The results showed higher protein, total phenolic content, and antioxidant capacity in KJ samples than in K ones. Vitamin C content increased during fermentation and reached its maximum concentration on day 45 (7.1 ± 0.3 mg/100 mL) for KJ. Caffeine in the supplemented samples was the main biocompound among those identified. Vitamin B12 formed on day 4 in K and after 24 h in KJ samples, remaining constant at the initial value of 2.30 ± 0.01 mg/100 mL up to day 45. (4) Conclusions: The results highlight that the fortification of kombucha with jujubes improved its biological activity and the content of bioactive compounds.

Green HPLC–DAD method for the analysis of analgesic and antihistamine drugs adulterated in herbal mixtures

This study presents a HPLC-DAD method developed for the simultaneous detection of histamine H1 antagonist drugs and analgesic compounds, including paracetamol, cetirizine hydrochloride, cinnarizine, chlorpheniramine maleate, and loratadine, in herbal formulations. This method systematically investigates critical parameters for enhancing separation and quantification using HPLC-DAD. Chemometric techniques were employed to optimize the method for detecting adulterants, with sample preparation involving liquid-phase extraction and a mobile phase for separation using HPLC-DAD. The calibration curves were linear over concentrations ranging from 10 to 40 µg/mL, with correlation coefficients exceeding 0.99. The method exhibits good accuracy, with mean recoveries ranging from 90 to 105 %, and precision (% RSD) for intraday and interday precision: 0.71 to 2.94 %. The limits of detection (LODs) and limits of quantification (LOQs) for each compound are provided. The HPLC-DAD method ensures rigorous quality control, detecting potential adulteration in 32 traditional medicine samples and emphasizing its safety and efficacy in managing allergic conditions. The proposed approach is considered environmentally friendly based on evaluations from commonly used greenness assessment tools, Green analytical chemistry metrics (ComplexGAPI, AGREE, BAGI) confirm the method's sustainability and practicality.

GC–MS based antioxidants characterization in Saussurea heteromalla (D. Don) Hand-Mazz by inhibition of nitric oxide generation in macrophages

For centuries, medicinal plants have served as the cornerstone for traditional health care systems and same practice is still prevalent today. In the Himalayan region, Saussurea heteromalla holds a significant place in traditional medicine and is used to address various health issues. Despite its historical use, little exploration has focused on its potential for scavenging free radicals and reducing inflammation. Hence, our current study aims to investigate the free radical scavenging capabilities of S. heteromalla extracts. The n-hexane extract of entire plant revealed promising activity. This extract underwent extensive extraction on a larger scale. Subsequent purification, employing column chromatography, HPLC–DAD techniques, led to the identification of active compounds, confirmed via GC–MS and the NIST database as 1-O-butyl 2-O-octyl benzene-1,2-dicarboxylate and 2,4-ditert-butylphenol. Assessing the free radical scavenging properties involved utilizing RAW-264.7 macrophages activated by lipopolysaccharides. Notably, the compound 2,4-di-tert-butylphenol exhibited remarkable scavenging abilities, demonstrating over 80% inhibition of Nitric oxide. This study stands as the inaugural report on the isolation of these compounds from S. heteromalla.