Concentrations Of Silymarin Research Articles

Improved dissolution and bioavailability of silymarin delivered by a solid dispersion prepared using supercritical fluids

The objective of this study was to improve the dissolution and bioavailability of silymarin (SM). Solid dispersions (SDs) were prepared using solution-enhanced dispersion by supercritical fluids (SEDS) and evaluated in vitro and in vivo, compared with pure SM powder. The particle sizes, stability, and contents of residual solvent of the prepared SM-SDs with SEDS and solvent evaporation (SE) were investigated. Four polymer matrix materials were evaluated for the preparation of SM-SD-SEDS, and the hydrophilic polymer, polyvinyl pyrrolidone K17, was selected with a ratio of 1:5 between SM and the polymer. Physicochemical analyses using X-ray diffraction and differential scanning calorimetry indicated that SM was dispersed in SD in an amorphous state. The optimized SM-SD-SEDS showed no loss of SM after storage for 6 months and negligible residual solvent (ethanol) was detected using gas chromatography. In vitro drug release was increased from the SM-SD-SEDS, as compared with pure SM powder or SM-SD-SE. In vivo, the area under the rat plasma SM concentration-time curve and the maximum plasma SM concentration were 2.4-fold and 1.9-fold higher, respectively, after oral administration of SM-SD-SEDS as compared with an aqueous SM suspension. These results illustrated the potential of using SEDS to prepare SM-SD, further improving the biopharmaceutical properties of this compound.

Protective activity of Panduratin A against thioacetamide-induced oxidative damage: demonstration with in vitro experiments using WRL-68 liver cell line.

BackgroundChalcone Panduratin A (PA) has been known for its antioxidant property, but its merits against oxidative damage in liver cells has yet to be investigated. Hence, the paper aimed at accomplishing this task with normal embryonic cell line WRL-68.MethodsPA was isolated from Boesenbergia rotunda rhizomes and its 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging and ferric reducing power (FRAP) activities were measured in comparison with that of the standard reference drug Silymarin (SI). Oxidative damage was induced by treating the cells with 0.04 g/ml of toxic thioacetamide for 60 minutes followed by treatment with 1, 10 and 100 μg/ml concentrations of either PA or SI. The severities of oxidative stress in the control and experimental groups of cells were measured by Malondialdehyde (MDA) levels, superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) activities.ResultsPA exhibited an acceptable DPPH scavenging and FRAP activities close to that of Silymarin. Treating the injured cells with PA significantly reduced the MDA level and increased the cell viability, comparable to SI. The activities of SOD, CAT and GPx were significantly elevated in the PA-treated cells in a dose dependent manner and again similar to SI.ConclusionCollectively, data suggested that PA has capacity to protect normal liver cells from oxidative damage, most likely via its antioxidant scavenging ability.

Silymarin-loaded solid nanoparticles provide excellent hepatic protection: physicochemical characterization and in vivo evaluation

BackgroundThe purpose of this study was to develop a novel silymarin-loaded solid nanoparticle system with enhanced oral bioavailability and an ability to provide excellent hepatic protection for poorly water-soluble drugs using Shirasu porous glass (SPG) membrane emulsification and a spray-drying technique.MethodsA silymarin-loaded liquid nanoemulsion was formulated by applying the SPG membrane emulsification technique. This was further converted into solid state nanosized particles by the spray-drying technique. The physicochemical characteristics of these nanoparticles were determined by scanning electron microscopy, differential scanning calorimetry, and powder X-ray diffraction. Their dissolution, bioavailability, and hepatoprotective activity in rats were assessed by comparison with a commercially available silymarin-loaded product.ResultsFormulation of a silymarin-loaded nanoemulsion, comprising silymarin, castor oil, polyvinylpyrrolidone, Transcutol HP, Tween 80, and water at a weight ratio of 5/3/3/1.25/1.25/100 was accomplished using an SPG membrane emulsification technique at an agitator speed of 700 rpm, a feed pressure of 15 kPa, and a continuous phase temperature of 25°C. This resulted in generation of comparatively uniform emulsion globules with a narrow size distribution. Moreover, the silymarin-loaded solid nanoparticles, containing silymarin/castor oil/polyvinylpyrrolidone/Transcutol HP/Tween 80 at a weight ratio of 5/3/3/1.25/1.25, improved about 1,300-fold drug solubility and retained a mean size of about 210 nm. Silymarin was located in unaltered crystalline form in the nanoparticles. The drug dissolved rapidly from the nanoparticles, reaching nearly 80% within 15 minutes, indicating three-fold better dissolution than that of the commercial product. Further, the nanoparticles showed a considerably shorter time to peak concentration, a greater area under the concentration-time curve, and a higher maximum concentration of silymarin compared with the commercial product (P < 0.05). In particular, the area under the concentration-time curve of the drug provided by the nanoparticles was approximately 1.3-fold greater than that of the commercial product. In addition, the silymarin-loaded nanoparticles significantly reduced carbon tetrachloride-induced hepatotoxicity, indicating improved bioactivity compared with silymarin powder and the commercial product.ConclusionSilymarin-loaded nanoparticles developed using SPG membrane emulsification and spray-drying techniques could be a useful system for delivery of poorly water-soluble silymarin while affording excellent hepatic protection.

Improved silymarin content in elicited multiple shoot cultures of Silybum marianum L.

Silybum marianum L. extracts are being used as antihepatotoxic therapy for liver diseases. Silymarin is a polyphenolic flavonoid mixture isolated from milk thistle which is believed to be responsible for the plant's hepatoprotective action. Regeneration of Silybum marianum plants from shoot tip explants and assessment of their morphogenic potential, silymarin total concentration and its major constituents upon exposure to medium composition alteration and different elicitors' application was targeted. Different concentrations of NaCl, quercetin, gamma irradiation and dried fungal extracts were used to elicit silymarin production in the cultures. The chemical composition of silymarin and its total concentration was investigated through HPLC at all the experiment stages. Multiple shoots were recorded after 3weeks of culture on MS medium containing various concentrations of BA and/or NAA. IAA was more effective than NAA and IBA in inducing robust roots in shoot cultures. The flowering plants recorded 20% and 40% of the total plants number in the multiplication and rooting stages respectively. The highest total silymarin concentration reached its peak with (10Gy) gamma-irradiation to be 6.598% dry weight in the in vitro regenerated shoot tip explants. The in vitro grown flowers showed 1.7 times more sylimarin productivity as compared to that of the wild grown congruent.

First detailed quantification of silymarin components in the leaves ofSilybum marianumcultivated in egypt during different growth stages

Summary A high performance liquid chromatographic method was developed for the quantitative determination of the active components of silymarin in the leaves of Silybum marianum during different growth stages. In this study, taxifolin and six main active constituents in silymarin, including silydianin, silychristin, diastereomers of silybin (silybin A and B) and diastereomers of isosilybin (isosilybin A and B) were completely separated on a 5-μm ODS column (Luna, Phenomenex, USA) with a mobile phase consisting of methanol and 5 mM NaH2PO4 (pH 3.5 adjusted with phosphoric acid) in a ratio of 45:55 v/v. Quantitation was performed with UV detection at 280 nm, based on peak area. The concentration of each component, as well as the total silymarin concentration was determined and compared with those of the seeds, aiming at optimizing the utilization of the cultivated plant. The developed method was validated with respect to linearity, range, specificity, accuracy, precision, and robustness. The leaves were fou...

Chemotherapeutic activity of silymarin combined with doxorubicin or paclitaxel in sensitive and multidrug-resistant colon cancer cells

The milk thistle extract silymarin, alone or in combined chemotherapy, is now under investigation in anticancer research, with particular interest for its possible employ in the treatment of chemoresistant tumours. So far, the consequences of a silymarin pre-treatment have not been thoroughly investigated. We studied whether silymarin pre-treatment synergized with chemotherapy, exploring the dose-dependence of the interaction in sensitive and multidrug-resistant cells. We studied cell cycle perturbations induced by silymarin in two colon carcinoma cell lines, LoVo and the multidrug-resistant isogenic LoVo/DX. Synergism/additivity/antagonism of silymarin-doxorubicin silymarin-paclitaxel combined treatments were evaluated by isobologram/combination index analysis, in the whole spectrum of active and sub-active concentrations of all drugs. The mechanisms of silymarin interaction with the other drugs were investigated by measuring drug uptake and cell cycle perturbations. Silymarin had similar antiproliferative activity against both cell lines. Pre-treatment with low silymarin concentrations synergised with both doxorubicin and paclitaxel in LoVo but not in LoVo/DX. Higher silymarin concentrations were additive with doxorubicin and paclitaxel in both cell lines. Silymarin favourably interfered with uptake and cell cycle effects of the chemotherapeutics in LoVo but not in LoVo/DX. These findings confirm activity of silymarin against colon carcinoma, including multidrug-resistant types, at relatively high but clinically achievable concentrations. In view of its low toxicity, two schedules based on low- and high-dose silymarin pre-treatment might offer a valuable option for combined treatment.

Elicitation of silymarin in cell cultures of Silybum marianum: effect of subculture and repeated addition of methyl jasmonate

Production of silymarin and the effect of the elicitor, methyl jasmonate (MeJA), was monitored in cell cultures of Silybum marianum over 4 years. Silymarin concentrations gradually declined after prolonged subculture, making the success of elicitor strategy limited in long-term cultures. The continuous presence of MeJA in cultures for an extended period was necessary for induction of silymarin accumulation. A repeated elicitor strategy was not a good option for improving silymarin productivity in batch cultures. Removal of medium from elicited cultures and addition of fresh medium avoided the toxic effects of elicitor accumulation, allowing the system to respond to a repeated MeJA treatment without loss of productivity.

Effects of silymarin on the spontaneous proliferation and cell cycle of human peripheral blood leukemia T cells

Silymarin is a polyphenolic flavonoid that has a strong antioxidant activity and exhibits anti-carcinogenic, anti-inflammatory, and cytoprotective effects. Although its hepatoprotective effect has been well documented, the effect of silymarin on T cells is largely unknown. The purpose of this study was to analyze the effects of the silymarin on the proliferation and cell cycle progression of Jurkat cells, a human peripheral blood leukemia T cell line. Cells were incubated with various concentrations of silymarin for 24-72 h and examined for cell growth and proliferation using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and DNA 5-bromo 2'-deoxyuridine (BrdU) colorimetric assays. Cell cycle analysis by flow cytometry was also performed using propidium iodide staining. Results of the study revealed that silymarin increased proliferation of Jurkat cells at 50-400 microM concentrations with 24 h exposure, confirmed by both MTT and BrdU assays. However, Jurkat incubation with silymarin at higher concentrations of 400 microM for 48 h and 200-400 microM for 72 h caused inhibition of DNA synthesis, cell cycle arrest at the G2/M phase and significant cell death. Results of the present study also revealed a similarity of cell growth patterns between Jurkat, U937 and RPMI 8866 cells. In conclusion, this study demonstrated an in vitro growth stimulatory effect of silymarin on leukemia cells with monocyte, T and B cell origin that has not been previously reported for either solid tumors or other leukemia cells, suggesting a possible specific stimulatory effect of silymarin on the key cells of the immune system.

Effects of silymarin on the proliferation and glutathione levels of peripheral blood mononuclear cells from β-thalassemia major patients

Iron toxicity in β-thalassemia major is the main cause of oxidative stress and cell mediated immune deficiencies. Despite indicative signs of severe oxidative deficiencies associated with β-thalassemia major, such as decreased level of plasma antioxidants and depletion of erythrocyte glutathione, little is known about intracellular redox status of immune cells. Since glutathione is a primary intracellular antioxidant and plays an essential role in several functions in T cells, in this study intracellular glutathione (GSH) levels as well as proliferation of PHA-activated peripheral blood mononuclear cells (PBMC) were investigated in 28 β-thalassemia major patients and 28 healthy age-matched individuals. Considering the potential benefits of flavonoids in the therapy of oxidative stress, the effects of silymarin on the GSH levels and proliferation of PBMC from normal and thalassemia individuals were further examined. Quantitative determination of intracellular GSH and proliferative response of PBMC to PHA were performed before and after 72 h incubation of PBMC with various concentrations of silymarin (0, 5, 10, or 20 μg/ml). Results demonstrated a significant reduction of GSH and proliferation in β-thalassemia major cells; however treatment with silymarin led to restoration of both GSH levels and PBMC proliferation in thalassemia patients. Considerably low levels of GSH and depressed proliferative response of PBMC in β-thalassemia major may be responsible for the cell mediated immune abnormalities in iron overload conditions. Moreover, the GSH restoration and improvement of PBMC growth by silymarin is a possible explanation for its recently reported antioxidant and immunostimulatory activities. These data suggest the benefit of using flavonoids to normalize immune dysfunction in β-thalassemia major. The immunomodulatory effects of silymarin in β-thalassemia major are currently under further investigation in a double blind clinical trial.

Enhancement of mammary carcinogenesis in two rodent models by silymarin dietary supplements

Silymarin is a mixture of polyphenolic flavonoids isolated from milk thistle (Silybum marianum) with anticancer activities reported for several organ sites. The present study tested the efficacy of dietary silymarin against mammary carcinogenesis in two rodent models. In the Sprague-Dawley rat model, female rats were fed a purified diet supplemented with none, 0.03, 0.1, 0.3 or 1% (w/w) of silymarin from 21 days of age (DOA) and carcinogenesis was initiated by a single i.p. injection of 1-methyl-1-nitrosourea (MNU) at 51 DOA. Mammary tumor (MT) development was followed till 110 days after carcinogen injection. In the MMTV-neu/HER2 transgenic mouse mammary carcinogenesis model, homozygous transgenic females were fed a purified diet supplemented with none or 0.3% silymarin, either from 28 or 120 DOA and MT development was followed to approximately 300 DOA. The results showed that dietary silymarin increased the plasma concentration of free and total silibinin, a major component of silymarin, in a dose-dependent manner in the rat, but did not decrease either MT incidence or number. Instead silymarin modestly increased the number of MNU-induced MTs in rats. Similarly, silymarin increased MT incidence and multiplicity and non-MTs in the neu-transgenic mice. In cell culture, treatment of human MCF-7 breast cancer cells with serum-achievable concentrations of silymarin in the rodent models stimulated their growth, in part through an estrogen-like activity. Because silymarin is being used in the treatment of liver cirrhosis and a variety of other human ailments, and is sold as a dietary supplement, our findings add a cautionary note to its application in breast cancer prevention.

Preparation of silymarin proliposome: A new way to increase oral bioavailability of silymarin in beagle dogs

The aim of the present study was to find a method to increase oral bioavailability of silymarin, that is to say, by the preparation of silymarin proliposome and to compare the pharmacokinetic characteristics and bioavailability after oral administration of silymarin proliposome and silymarin in beagle dogs. Silymarin proliposome was prepared by the film-deposition on carriers. After the proliposome was contacted with water, the silymarin liposome suspensions formed automatically. The tests of physicochemical properties including SEM, TEM, encapsulation efficiency, dissolution studies, particle size of the reconstituted liposome and stability of the silymarin proliposome were determined by laser-particle-sizer, HPLC, etc. The concentrations of silymarin in plasma of beagle dogs and its pharmacokinetic behaviors after oral administration of silymarin liposome suspensions and silymarin were studied by RP-HPLC. The pharmacokinetic parameters were computed by software program 3p97. The encapsulation efficiency of silymarin liposome could be more than 90%, with an average particle size of about 196.4 nm and the proliposome appeared a very stability at 40 °C during 3 months. It was found that mean plasma concentration–time curves of silymarin after oral administration of liposome suspensions and silymarin in beagle dogs were both in accordance with open two-compartments model and first-order absorption. Pharmacokinetic parameters of silymarin proliposome and silymarin in beagle dogs were T max both 30 min; C max 472.62 and 89.78 ng mL −1; and AUC0-∞ 2606.21 and 697 ng mL −1 h, respectively. The high bioavailability of silymarin proliposome could be obtained by oral administration. Silymarin proliposome was stable and did enchance the gastrointestinal absorption of silymarin.

Protective Effects of Silymarin against Free Radical-Induced Erythrocyte Lysis

The oxidative hemolysis of rat erythrocytes induced by 2,2’-azobis–(2-amidinopropane) (AAPH) and its inhibition by silymarin studied. Different concentrations of silymarin showed no significant hemolysis compared with phosphate buffer solution. AAPH (25 mM and 50 mM) induced hemolysis in a time-dependent manner. Silymarin inhibited AAPH (25 mM)-induced hemolysis concentration-dependently. However, in the presence of 50 mM of AAPH, only the two higher concentrations of silymarin inhibited hemolysis. Addition of silymarin 3h after incubation with AAPH (25 mM), had no significant effects on the time course of cell lysis. It is concluded that, in addition to its well-established antioxidant effects, silymarin displays cytoprotective properties.

Hepatoprotective activity of Thunbergia laurifolia Linn extract in rats treated with ethanol: In vitro and in vivo studies

Primary cultures of rat hepatocyte and rats were used as the in vitro and in vivo models to evaluate the hepatoprotective activity of aqueous extract from Thunbergia laurifolia (TLE). Ethanol was selected as hepatotoxin. Silymarin (SL) was the reference hepatoprotective agent. In the in vitro study, MTT reduction assay and release of transaminases (ALT and AST) were the criteria for cell viability. Primary cultures of rat hepatocyte (24 h culturing) were treated with ethanol (96 μl/ml) and various concentrations of TLE (2.5, 5.0, 7.5 and 10.0 mg/ml) or SL (1, 2 and 3 mg/ml) for 2 h. Ethanol decreased MTT (%) nearly by half. Both TLE and SL increased MTT reduction and brought MTT (%) back to normal. Ethanol induced release of ALT and AST was also reduced by TLE (2.5 and 5.0 mg/ml) and SL (1 mg/ml). In the in vivo study, serum transaminases, serum triglyceride (STg) together with hepatic triglyceride (HTg) and histopathological examination were the criteria for evidences of liver injury. Ethanol (4 g/(kg day), po for 14 days) caused the increase in ALT, AST, HTg and centrilobular hydropic degeneration of hepatocytes. TLE at 25 mg/(kg day), po, or SL at 5 mg/(kg day), po, for 7 days after ethanol enhanced liver cell recovery by bringing HTg, ALT and/or AST back to normal. These results suggest that TLE and SL possess the hepatoprotective activity against ethanol induced liver injury in both primary cultures of rat hepatocyte and rats.

Effect of milk thistle (Silybum marianum) on the pharmacokinetics of irinotecan

2096 Background: Milk thistle is one of the most commonly used non-traditional herbal therapies, particularly in Western Europe. The main component of the mixture of flavonolignans in milk thistle, silybin, significantly reduce the activities of CYP3A4 (Ki, ∼32 μM) and UGT1A1 (Ki, ∼1 μM) in human hepatocyte cultures in vitro. Here, we investigated whether milk thistle affects the pharmacokinetics of irinotecan, a substrate for both CYP3A4 and UGT1A1. Methods: Six cancer patients were treated with irinotecan at a dose of 125 mg/m2, given as a 90-min infusion in cycles of once every week. Four days before the second irinotecan administration, patients received standardized capsules from a single batch (GNC, Pittsburgh, PA), each containing 200 mg milk thistle seed extract, three times a day with water, for 14 consecutive days. Pharmacokinetic parameters of irinotecan and its metabolites SN-38, SN-38-glucuronide (SN-38G), and APC were obtained during the first, second, and third weekly administration. Results: During the first irinotecan administration (in the absence of milk thisle), the mean (± SD) irinotecan clearance was 31.2 ± 19.6 L/h. Short term (i.e., 4 days) or more prolonged intake of milk thistle (i.e., 12 days) had no significant effect on irinotecan clearance (cycle 2, 25.4 ± 14.6 L/h; cycle 3, 25.6 ± 13.6 L/h; P = 0.161). The AUC ratio of SN-38 and irinotecan was slightly decreased by milk thistle (cycle 1, 0.0258 vs cycle 2, 0.0223 vs cycle 3, 0.0217; P = 0.047). The relative extent of glucuronidation (AUC ratio of SN-38G and SN-38), as a measure of the potential of milk thistle to modulate the activity of UGT1A1, was similar in the three cycles of treatment (10.8 vs 13.5 vs 13.1; P = 0.636). Likewise, the AUC ratio of the CYP3A4-mediated metabolite APC and irinotecan was unaffected by milk thistle (0.331 vs 0.285 vs 0.337; P = 0.533). The maximum plasma concentrations of the silymarin component silybin at steady state, determined using LC/MS/MS, ranged between 0.0249μM and 0.257μM. Conclusions: This study suggests that silymarin concentrations after intake of recommended doses of milk thistle are too low to significantly affect the function of CYP3A4 and UGT1A1 in vivo, and that milk thistle does not affect the disposition of irinotecan. No significant financial relationships to disclose.

Extraction of nutraceuticals from milk thistle: part II. Extraction with organic solvents.

Seeds from milk thistle (Silybum marianum Gaert L.) contain flavanolignan and dihydroflavanol compounds that have interesting and important therapeutic activities. The recovery of these silymarin compounds generally involves a two-step defatting and extraction process using organic solvents. This study examined the batch, single-stage extraction of whole and defatted seeds using ethanol, methanol, acetonitrile, and acetone as the solvents. In extracting defatted milk thistle seeds with organic solvents, extraction with ethanol resulted in the highest silymarin yield, although some potential degradation was observed. The maximum yields of taxifolin, silychristin, silydianin, silybinin A, and silybinin B in ethanol were 0.6, 4.0, 0.4, 4.0, and 7.0 mg/g of defatted seed, respectively. However, if silybinin A were the diastereoisomer of choice, methanol would be the preferred extraction solvent because it yielded the highest silybinin A to silybinin B ratio. Interestingly, lipid removal is an important extraction step, because defatted material yields twice the silymarin concentration.

The effects of silymarin on the attachment and viability of primary cultures of rat hepatocytes

The effects of the hepatoprotective compound silymarin on hepatocytes in primary culture were studied. Exposure of cells in primary culture (both conventional cells and perivenous or periportal cells isolated by the digitonin-collagenase perfusion technique) to high concentrations of silymarin surprisingly demonstrated that silymarin per se was cytotoxic. Incubation of cells for 18 hr with silymarin at concentrations exceeding 25 μ m abruptly increased cell damage, whereas viability decreased in a more linear fashion with increasing concentrations of its major constituent, silybin. Morphologically, cell cultures exposed to silymarin concentrations lower than 20 μ m appeared normal, but at higher concentrations intercellular contacts were lost; cells appeared granulated and took up eosin. Silymarin and silybin at these doses were also found to prevent cell attachment. The mechanism responsible for this effect at relatively low concentrations of silymarin during prolonged exposure in the primary cell culture system is not clear at present. The effects of low doses on cell attachment to the matrix suggest an action on the cell membrane and/or on the cytoskeleton.

Effect of silymarin on plant tissue cultures

The effect of silymarin complex on various types of expiants differing in their nutrition requirements was investigated. The growth of tumorous periwinkle (Catharanthus roseus [L.] G. Don) callus tissue was still identical with the control tissue at the silymarin concentration of 35 mg in 1000 ml of the nutrient medium. However, this silymarin concentration totally inhibited the growth of habituated periwinkle callus tissue; in the presence of 10 mg of silymarin, the growth of this tissue was similar to that of the corresponding tissue grown without silymarin. The growth of tobacco callus tissue (D-strain) requiring for its growth kinetin was reduced by 46.2% at the concentration 10 mg of silymarin in 1000 ml of nutrient medium, but its dry weight was increased by 21% in comparison with the control. Silymarin was most effective on the growth of callus derived from tobacco (Nicotiana glauca Grah:) stem pieces; callogenesis was observed in control tissue in 89.5% cases while in the presence of silymarin (10 mg) only in 48.6%. The primary callus growth was strongly inhibited, too (by 89.9%). The organogenesis onset was never observed on tobacco stem pieces cultured on a nutrient medium with kinetin and IAA in the presence of silymarin. When all types of expiants were transferred from the medium with silymarin on control medium, normal growth appeared very soon and the differences between the experimental and control expiants were smoothed out during two months. These results indicate that the observed changes might be due to the blocking of membrane system permeability leading to an insufficient supply of cells with nutrients and growth substances.