Humulus lupulus aqueous extract and hydrolate as a potential ingredient for cosmetics: chemical characterization and in vitro antimicrobial, cytotoxicity, antioxidant and anti-inflammatory assessment.

Humulus lupulus extracts have in their composition different molecules, such as polyphenols, α-acids, ß-acids, and hydrocarbons, which contribute to the plant's medicinal properties. These molecules are associated with antimicrobial, antioxidant and anti-inflammatory activities. OBJECTIVE: This work focuses on the evaluation of H. lupulus biological activities, with the aim of evaluating its potential for inclusion in cosmetic formulations. METHODS: Two distinct aqueous extracts and two hydrolates obtained via hydrodistillation were evaluated. These include the flower parts (FE, FH) and the mix of aboveground parts (ME, MH). The chemical profiles for both aqueous extracts and hydrolates were identified by high performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC-MS). Antimicrobial, antioxidant, cytotoxicity, and anti-inflammatory activity were tested in vitro using standard methods. RESULTS: Rutin was the major compound found in FE (40.041 µg mg-1 of extract) and ME (2.909 µg mg-1 of extract), while humulenol II was the most abundant compound in hydrolates (FH: 20.83%; MH: 46.80%). Furthermore, FE was able to inhibit the growth of Staphylococcus aureus and Staphylococcus epidermis with MIC values of 50% and 25% (v/v), respectively. FH showed the same effect in Staphylococcus aureus (50% v/v). FH evidenced poor antioxidant potential in DPPH scavenging test and demonstrated significant antioxidant and anti-inflammatory effects by reducing (***p < 0.001) intracellular reactive oxygen species (ROS), NO (nitric oxide) levels (***p < 0.001) and cyclooxygenase-2 (COX-2) protein expression (***p < 0.001) in lipopolysaccharide (LPS)-stimulated macrophages. Nevertheless, it is important to note that FH exhibited cytotoxicity at high concentrations in 3T3 fibroblasts and RAW 264.7 macrophages. CONCLUSION: The studied H. lupulus aqueous extracts and hydrolates revealed that FH stands out as the most promising bioactive source for cosmetic formulations. However, future research addressing antimicrobial activity is necessary to confirm its potential incorporation into dermatological and cosmetic formulations.

Assessment of the phytochemical profiles of novel hop (Humulus lupulus L.) cultivars: A potential route to beer crafting.

This study investigated the volatile phytochemical diversity of 30 samples obtained from experimental hybrid and commercial H. lupulus L. plants. Essential oils distilled from these samples were analysed by high resolution gas chromatography coupled with accurate mass time-of-flight mass spectrometry (GC-accTOFMS). A total of 58 secondary metabolites, mainly comprising 18 esters, 6 monoterpene hydrocarbons, 2 oxygenated monoterpenes, 20 sesquiterpene hydrocarbons, 7 oxygenated sesquiterpenes, and 4 ketones, were positively or tentatively identified. A total of 24 metabolites were detected in all samples, but commercial cultivars (selected for brewing performance) had fewer compounds identified compared to experimental genotypes. Chemometrics analyses enabled distinct differentiation of experimental hybrids from commercial cultivars, discussed in terms of the different classes of compounds present in different genotypes. Differences among the mono- and sesquiterpenoids, appear to be related to either: i) the genetic origin of the plants; or ii) the processes of bioaccumulation of the identified secondary metabolites.

Xanthohumol from hop cones (Humulus lupulus L.) prevents ADP-induced platelet reactivity.

Hop cones (Humulus lupulus L.), very rich source of phenolic compounds, possessing anticancer, antioxidant and anti-inflammatory activities, are considered as beneficial diet ingredients improving human health. In this study, the antiplatelet action of xanthohumol (XN), the principal flavonoid in hop cones, was investigated. XN significantly attenuated ADP-induced blood platelet aggregation (97.2 ± 35.7 AU for 6 µg/ml of XN vs. 120.4 ± 30.1 AU for 0.17% dimethyl sulfoxide (DMSO), p < 0.001) and significantly reduced the expression of fibrinogen receptor (activated form of GPIIbIIIa) on platelets' surface (47.6 ± 15.8 for 1.5 µg/ml XN, 44.6 ± 17.3% for 3 µg/ml XN vs. 54.5 ± 19.2% for control or 43.3 ± 18.4% for 6 µg/ml XN vs. 49.7 ± 19.4% for 0.17% DMSO, p < 0.05 or less). These findings suggest that the phenolic compounds originating from hops (XN) have a novel role as antiplatelet agents and can likely be used as dietary supplements in prophylactic approaches.

Pharmacokinetics of prenylated hop phenols in women following oral administration of a standardized extract of hops.

SCOPE: Women seeking alternatives to hormone-replacement therapy for menopausal symptoms often try botanical dietary supplements containing extracts of hops (Humulus lupulus L.). Hops contain 8-prenylnaringenin (8-PN), a potent phytoestrogen, the related flavanones 6-prenylnaringenin and isoxanthohumol (IX), and the prenylated chalcone xanthohumol (XN). METHODS AND RESULTS: After chemically and biologically standardizing an extract of spent hops to these marker compounds, an escalating dose study was carried out in menopausal women to evaluate safety and pharmacokinetics. 8-PN, 6-prenylnaringenin, IX, and XN, sex hormones, and prothrombin time were determined in blood samples and/or 24 h urine samples. There was no effect on sex hormones or blood clotting. The maximum serum concentrations of the prenylated phenols were dose-dependent and were reached from 2 to 7 h, indicating slow absorption. The marker compounds formed glucuronides that were found in serum and urine. Secondary peaks at 5 h in the serum concentration-time curves indicated enterohepatic recirculation. The serum concentration-time curves indicated demethylation of IX to form 8-PN and cyclization of XN to IX. Slow absorption and enterohepatic recirculation contributed to half-lives exceeding 20 h. CONCLUSION: This human study indicated long half-lives of the estrogenic and proestrogenic prenylated phenols in hops but no acute toxicity.

Electrochemical versus spectrophotometric assessment of antioxidant activity of hop (Humulus lupulus L.) products and individual compounds.

Antioxidant (AO) activity of extracts of hop cones (Serbian domestic varieties) and commercial hop products (Saaz, Spalter, Spalter select, and Magnum pellets) was determined by parallel application of recently developed direct current (DC) polarographic and widely used DPPH assay. Correlations between 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) scavenging and total phenolics (TPC) determined by the Folin-Ciocalteu assay (FC) (0.99), and between H2O2 scavenging (HPS) and humulone content (H) determined by conductometric method (0.94), total resins (TR) (0.85), and hop storage index (HIS) (-0.90), were found statistically significant at p < 0.05 level while complete lack of HPS correlation with TPC and DPPH was observed. To obtain an insight into differences between results of AO assays applied, activity of individual compounds, prevalent hop phenolics, and bitter acids was determined. By far superior HPS activity of humulone was followed by catechin, quercetin, xanthohumol, lupulone, and rutin. In contrast, DPPH scavenging activity of phenolics (quercetin > catechin > rutin > xantohumol) was found substantially higher than activity of bitter acids. According to ferric reducing antioxidant power (FRAP) and scavenging of 2,2-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS), higher AO activity was ascribed to phenolics, while almost neglecting humulone. Besides reliability, low cost, and an easy-to-handle procedure, an ability to recognize humulone as the major contributor of hop AO activity could allow DC polarographic assay to be applied in analysis of various hop-derived products.

A semi-automatic microextraction in packed sorbent, using a digitally controlled syringe, combined with ultra-high pressure liquid chromatography as a new and ultra-fast approach for the determination of prenylflavonoids in beers.

In this work a highly selective and sensitive analytical procedure based on semi-automatic microextraction by packed sorbents (MEPS) technique, using a new digitally controlled syringe (eVol(®)) combined with ultra-high pressure liquid chromatography (UHPLC), is proposed to determine the prenylated chalcone derived from the hop (Humulus lupulus L.), xanthohumol (XN), and its isomeric flavonone isoxanthohumol (IXN) in beers. Extraction and UHPLC parameters were accurately optimized to achieve the highest recoveries and to enhance the analytical characteristics of the method. Important parameters affecting MEPS performance, namely the type of sorbent material (C2, C8, C18, SIL, and M1), elution solvent system, number of extraction cycles (extract-discard), sample volume, elution volume, and sample pH, were evaluated. The optimal experimental conditions involves the loading of 500µL of sample through a C18 sorbent in a MEPS syringe placed in the semi-automatic eVol(®) syringe followed by elution using 250µL of acetonitrile (ACN) in a 10 extractions cycle (about 5min for the entire sample preparation step). The obtained extract is directly analyzed in the UHPLC system using a binary mobile phase composed of aqueous 0.1% formic acid (eluent A) and ACN (eluent B) in the gradient elution mode (10min total analysis). Under optimized conditions good results were obtained in terms of linearity within the established concentration range with correlation coefficients (R) values higher than 0.986, with a residual deviation for each calibration point below 12%. The limit of detection (LOD) and limit of quantification (LOQ) obtained were 0.4ngmL(-1) and 1.0ngmL(-1) for IXN, and 0.9ngmL(-1) and 3.0ngmL(-1) for XN, respectively. Precision was lower than 4.6% for IXN and 8.4% for XN. Typical recoveries ranged between 67.1% and 99.3% for IXN and between 74.2% and 99.9% for XN, with relative standard deviations %RSD no larger than 8%. The applicability of the proposed analytical procedure in commercial beers, revealed the presence of both target prenylchalcones in all samples being IXN the most abundant with concentration of between 0.126 and 0.200µgmL(-1).

Application of plant-based antimicrobials for the growth inhibition of clostridia in pressed beet pulp silage.

BACKGROUND: In this study the inhibition of hop beta acids on the growth of clostridia in soil-contaminated pressed sugar beet pulp silages was investigated. Hop beta acids are natural substances which display their effect at low concentrations. Fresh pressed beet pulp material was mixed with soil to artificially contaminate it with clostridia. Laboratory silos were filled with the substrate, stored at 25 °C and opened for sampling at 0, 2, 8, 15, 30, 60, and 90 days. The impact on clostridial growth during silage fermentation was monitored by determination of the pH value and dry matter content, as well as chemical analysis of the fermentation products. Throughout the experiments, the effect of a commercial silage inoculant based on lactic acid bacteria (LAB) and hop-resistant LAB were examined with and without the combination of plant-based antimicrobials. RESULTS: Results indicate that in contaminated silage samples without any additives high butyric acid contents occurred due to clostridial growth. This spoilage could not be suppressed by the application of LAB, whereas the combined application of LAB and hop beta acids significantly improved silage quality, which was reflected by favourable organic acid composition (P < 0.05). CONCLUSION: The experimental data indicate that the application of hop beta acids improves the preservation effect of LAB in suppressing clostridial growth in silages and thus demonstrates some potential for the combined use of plant-based antimicrobials and LAB.