Natural anticancer agent

In p53 wild-type and p53 liver cancer lines, silibinin A strongly increases the expression of nonsteroidal anti-inflammatory drug-activated gene-1 (NAG-1) [1]. Human breast cancer cell lines MCF7 and MDA-MB-231 are induced by silybin A. HCC cell viability is inhibited by silybin A in a time- and dose-dependent manner [3]. Silybin A increases the agar-associated protein Bax while decreasing the RBP-Jκ, Hes1, and Notch1 intracellular domain (NICD) proteins. decrease cyclin D1, survivin, and Bcl2[3].

---

Silibinin, an effective anti-cancer and chemopreventive agent, has been shown to exert multiple effects on cancer cells, including inhibition of both cell proliferation and migration. However, the molecular mechanisms responsible for these effects are not fully understood. We observed that silibinin significantly induced the expression of the non-steroidal anti-inflammatory drug-activated gene-1 (NAG-1) in both p53 wild-type and p53-null cancer cell lines, suggesting that silibinin-induced NAG-1 up-regulation is p53-independent manner. Silibinin up-regulates early growth response-1 (EGR-1) expression. The ectopic expression of EGR-1 significantly increased NAG-1 promoter activity and NAG-1 protein expression in a dose-dependent manner. Furthermore, down-regulation of EGR-1 expression using siRNA markedly reduced silibinin-mediated NAG-1 expression, suggesting that the expression of EGR-1 is critical for silibinin-induced NAG-1 expression. We also observed that reactive oxygen species (ROS) are generated by silibinin; however, ROS did not affect silibinin-induced NAG-1 expression and apoptosis. In addition, we demonstrated that the mitogen-activated protein kinase (MAP kinase) signal transduction pathway is involved in silibinin-induced NAG-1 expression. Inhibitors of p38 MAP kinase (SB203580) attenuated silibinin-induced NAG-1 expression. Furthermore, we found that siRNA-mediated knockdown of NAG-1 attenuated silibinin-induced apoptosis. Collectively, the results of this study demonstrate for the first time that up-regulation of NAG-1 contributes to silibinin-induced apoptosis in cancer cells. [1]

---

---

The present study was undertaken to determine the underlying mechanism of silibinin-induced cell death in human breast cancer cell lines MCF7 and MDA-MB-231. Silibinin-induced cell death was attenuated by antioxidants, N-acetylcysteine (NAC) and 6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid, suggesting that the effect of silibinin was dependent on generation of reactive oxygen species (ROS). Western blot analysis showed that silibinin induced downregulation of extracellular signal-regulated kinase (ERK) and Akt. When cells were transiently transfected with constitutively active (ca) mitogen-activated protein kinase (MEK), an upstream kinase of ERK and caAkt, they showed resistance to silibinin-induced cell death. Silibinin decreased the cleavage of Notch-1 mRNA and protein levels. Notch-1-overexpressed cells were resistant to the silibinin-induced cell death. Inhibition of Notch-1 signaling was dependent on ROSgeneration. Overexpression of Notch-1 prevented silibinin-induced inhibition of ERK and Akt phosphorylation. Silibinin-induced cell death was accompanied by increased cleavage of caspase-3 and was prevented by caspase-3 inhibitor in MDA-MB-231 cells but not in MCF7 cells. Silibinin induced translocation of apoptosis-inducing factor (AIF), which was blocked by NAC, and transfection of caMEK and caAkt. Silibinin-induced cell death was prevented by silencing of AIF expression using small interfering AIF RNA in MCF7 cells but not in MDA-MB-231 cells. In conclusion, silibinin induces cell death through an AIF-dependent mechanism in MCF7 cells and a caspase-3-dependent mechanism in MDA-MB-231 cells, and ROS generation and Notch-1 signaling act upstream of the ERK and Akt pathway. These data suggest that silibinin may serve as a potential agent for induction of apoptosis in human breast cancer cells.[2]

---

---

Hepatocellular carcinoma (HCC) is a global health burden that is associated with limited treatment options and poor patient prognoses. Silybin (SIL), an antioxidant derived from the milk thistle plant (Silybum marianum), has been reported to exert hepatoprotective and antitumorigenic effects both in vitro and in vivo. While SIL has been shown to have potent antitumor activity against various types of cancer, including HCC, the molecular mechanisms underlying the effects of SIL remain largely unknown. The Notch signaling pathway plays crucial roles in tumorigenesis and immune development. In the present study, we assessed the antitumor activity of SIL in human HCC HepG2 cells in vitro and in vivo and explored the roles of the Notch pathway and of the apoptosis-related signaling pathway on the activity of SIL. SIL treatment resulted in a dose- and time-dependent inhibition of HCC cell viability. Additionally, SIL exhibited strong antitumor activity, as evidenced not only by reductions in tumor cell adhesion, migration, intracellular glutathione (GSH) levels and total antioxidant capability (T-AOC) but also by increases in the apoptotic index, caspase3 activity, and reactive oxygen species (ROS). Furthermore, SIL treatment decreased the expression of the Notch1 intracellular domain (NICD), RBP-Jκ, and Hes1 proteins, upregulated the apoptosis pathway-related protein Bax, and downregulated Bcl2, survivin, and cyclin D1. Notch1 siRNA (in vitro) or DAPT (a known Notch1 inhibitor, in vivo) further enhanced the antitumor activity of SIL, and recombinant Jagged1 protein (a known Notch ligand in vitro) attenuated the antitumor activity of SIL. Taken together, these data indicate that SIL is a potent inhibitor of HCC cell growth that targets the Notch signaling pathway and suggest that the inhibition of Notch signaling may be a novel therapeutic intervention for HCC.[3]

---

Effects of silybin on glucose-induced superoxide generation in cultured mouse podocytes.[5]
The effect of silybin on HG-induced intracellular ROS generation in cultured mouse podocytes was tested using DHE fluorescence and confocal microscopy. As shown in Fig. 1A, exposure of podocytes to HG resulted in 70% increase of intracellular ROS production, and this increase was completely inhibited by coincubation of the cells with 10 μM silybin. Generation of superoxide was also analyzed by quantifying the production of 2-OH-E+, the superoxide specific product of DHE, by HPLC. Exposure of the podocytes to HG caused 60% increase in 2-OH-E+ production, and this effect was abrogated by coincubation with silybin (Fig. 1B). Figure 1C shows a dose-dependent response to silybin; incubation with 0.1 μM silybin inhibited partially the HG-induced generation of 2-OH-E+, while 1 and 10 μM silybin resulted in complete inhibition.

---

Effect of silybin on glucose-induced NADPH oxidase activity in podocytes.[5]
Since NADPH oxidases of the Nox family are a major source of superoxide in renal cells, including podocytes, we assessed NADPH oxidase activity in podocytes under the same experimental conditions as above, using the lucigenin-enhanced chemiluminescence assay. The NADPH-dependent superoxide generation increased by 90% after exposure of the cells to HG for 24 h. Treatment with silybin completely suppressed HG-induced increase in NADPH oxidase activity (Fig. 2A). Western blot analysis showed that HG-mediated increased expression of Nox4 protein expression is totally inhibited by silybin treatment (Fig. 2B). Analysis of Nox1 yielded similar results, although the constitutive and HG-indued expression of this protein was weaker than for Nox4 (Fig. 2C).

---

Effect of silybin on glucose-induced podocytes injury.[5]
Apoptotic cell death is one of the manifestations of podocyte injury in the diabetic milieu. Podocytes apoptosis was examined in vitro by two different methods, Hoechst staining and DNA fragmentation (20). As shown by Hoechst staining (Fig. 3A), exposure of the podocytes to HG for 24 h resulted in significant podocyte apoptosis compared with NG, while silybin protected against HG-induced podocyte apoptosis. Likewise, silybin completely prevented DNA fragmentation, a measure of podocyte apoptosis induced by HG (Fig. 3B).

The Middle East can effectively inhibit oxidation, deactivate mediators, and prevent tumorigenesis with the topical application of silybin A (silybin A) at a dose of 9 mg/animal [4]. Superoxide production increased and Nox4 expression was higher than in the mouse cortex. Mice treated with silibin A did not exhibit the 35% podocyte residual observed in the glomeruli of control OVE26 mice [5].

---

Effects of Silybin A (Silibinin A) on Tumor Xenografts in vivo [3]
Further in vivo tumor xenograft experiments were carried out to verify our previous in vivo experiments. To determine whether Silybin A (Silibinin A)/SIL could inhibit tumor growth in animals, we established HepG2 xenografts in athymic nude mice. We found that mice in all treatment groups developed subcutaneous tumors. As shown in Figures 8A and 8B, SIL treatment (200 or 400 mg/kg) significantly inhibited tumor growth (P<0.01, compared with the control group). Additionally, Western blot analysis showed that SIL treatment induced a dose-dependent downregulation of NICD, cyclin D1, and survivin (P<0.01, compared with the control group; Figure 8C). Additionally, the mitochondrial apoptotic pathway-related protein Bax was upregulated by SIL treatment, while Bcl2 was downregulated (P<0.01, compared with the control group; Figure 8C).

---

Effects of Silybin A (Silibinin A) Combined with DAPT on Tumor Xenografts in vivo [3]
DAPT was used to explore the effect of downregulation of Notch signaling on the antitumor activity of Silybin A (Silibinin A)/SIL in vivo. As shown in Figures 9A and 9B, treatment with SIL or DAPT alone significantly inhibited tumor growth (P<0.01 or P<0.05, compared with the control group). The combination of SIL and DAPT further inhibited tumor growth (P<0.01, compared with the SIL or DAPT groups). Western blot analysis showed that SIL and DAPT co-treatment further decreased NICD, cyclin D1, and survivin expression (P<0.01, compared with the SIL and DAPT groups; Figure 9C). Additionally, Bax was further upregulated by the SIL and DAPT co-treatment, while Bcl2 was further downregulated (P<0.01, compared with the SIL and the D groups; Figure 9C).

---

Silybin A (Silibinin A) is a major bioactive flavonolignan present in milk thistle (Silybum marianum) that possesses antioxidant, antiinflammatory, and anticarcinogenic activity. However, the precise underlying mechanism remains to be elucidated. The present study was designed to investigate underlying molecular mechanism for antitumorigenic potential of silibinin against chemically induced skin tumorigenesis in Swiss albino mice. In light of the important role of nuclear factor-kappaB (NF-κB), cyclooxygenase-2 (COX-2), iNOS, proinflammatory cytokines, vascular endothelial growth factor, and oxidative stress in carcinogenesis, chemopreventive efficacy of silibinin against 7, 12-dimethylbenz[a]anthracene/12-O-tetradecanoylphorbol-13-acetate-induced 2-stage skin carcinogenesis was studied in terms of cytoprotective enzymes activity, lipid peroxidation, inflammatory responses, and the expression of various molecular marker in skin tissue. We found that topical application of silibinin at the dose of 9 mg/mouse effectively suppressed oxidative stress and deregulated activation of inflammatory mediators and tumorigenesis. Thus, findings of the present study suggest that the chemopreventive effect of silibinin is associated with upregulation of endogenous cytoprotective machinery and down regulation of inflammatory mediators (nitric oxide, tumor necrosis factor-α, interleukin-6, interleukin -1β, COX-2, iNOS, and NF-κB).[4]

---

---

Studies on a type 1 diabetic mouse model. [5]
To validate the findings in cultured immortalized mouse podocyte, we studied the effect of Silybin A (Silibinin A) on an established mouse model of type 1 diabetes, the OVE26 mouse. These mice develop morphological and structural changes characteristics of human diabetic nephropathy (63). Three groups of mice were studied: 1) FVB control animals, 2) control diabetic OVE26 animals treated with vehicle, and 3) OVE26 diabetic mice treated with silybin for 6 wk. Blood glucose levels were 130.4 ± 12.4 mg/dl in the FVB control animals and above the detection limit (600 mg/dl) in the OVE26 diabetic groups, irrespective of treatment with vehicle or silybin.

---

Effects of Silybin A (Silibinin A) on superoxide production in the renal cortex.[5]
Superoxide production was measured in kidney cortex using DHE and HPLC. 2-OH-E+ production was increased in the renal cortex from diabetic mice and silybin treatment significantly reduced the increase of 2-OH-E+ in OVE26 mice (Fig. 4A). In addition, immunohistochemistry of kidney cortex showed increased expression of Nox4 in the OVE26 mice compared with the FVB control mice (Fig. 4B). Silybin treatment prevented the increased expression of Nox4 in the OVE26 mice (Fig. 4B).

---

Effect of Silybin A (Silibinin A) on albuminuria.[5]
The diabetic mice developed severe albuminuria when compared with the nondiabetic mice. Silybin treatment attenuated the albuminuria in the diabetic mice by 54% (P = 0.06; Fig. 5D).

Conditionally immortalized mouse podocytes were grown to near confluence and reseeded three times under growth-permissive conditions (33°C) in flasks coated with type I collagen in a humidified chamber with 5% CO2. Growth media consisted of RPMI 1640 with 10% fetal bovine serum, 100 U/ml of penicillin/streptomycin, and 5 mM d-glucose (NG). The medium was supplemented with 50 U/ml of mouse interferon-γ (INF-γ) during the first passage and then 10 U/ml INF-γ in the second and third passage. Cell differentiation was then induced by subculturing the cells under nonpermissive conditions (37°C) in serum-containing medium without INF-γ for 10–14 days (46). Cells were serum starved for 24 h before the experiment with RPMI 1640 containing NG and 0.2% BSA. The experiment consisted of overnight pretreatment of the cells with Silybin A (Silibinin A) at a final concentration of 10 μM or with vehicle. Cells were then exposed to NG or 25 mM d-glucose (HG) with or without Silybin A (Silibinin A) for 24 h [5].

---

Cell Culture and Treatment [3]
Human HCC HepG2 cells were grown in Dulbecco’s modified Eagle medium supplemented with 10% fetal bovine serum, L-glutamine (2 mM), penicillin (100 units/ml), streptomycin (100 units/ml), and HEPES (25 mM). Cells were maintained in the presence of 5% CO2 at 37°C. Cells were treated with different concentrations of Silybin A (Silibinin A) (50, 100, and 200 µM dissolved in DMSO) for the indicated time periods. An equal amount of DMSO (vehicle) was present in each treatment, including the control; the DMSO concentration did not exceed 0.1% (v/v) in any of the treatments.

---

Analysis of Cell Apoptosis [3]
HepG2 cell apoptosis was detected using the FITC-Annexin V/PI staining kit. After Silybin A (Silibinin A) treatment, the cells were harvested, washed in ice-cold PBS, incubated for 15 min with fluorescein-conjugated Annexin V and PI, and analyzed using a FACScan flow cytometer equipped with the FACStation data management system and Cell Quest software.

---

Analyses of Intracellular Reactive Oxygen Species (ROS) Generation, GSH Levels, and T-AOC [3]
The measurement of intracellular ROS was based on the ROS-mediated conversion of non-fluorescent 2′,7′-DCFH-DA into fluorescent DCFH. After treatment with Silybin A (Silibinin A), the cells were trypsinized and subsequently incubated with DCFH-DA (20 µM) in PBS at 37°C for 2 h. After incubation, the DCFH fluorescence of the cells in each well was measured using an FLX 800 microplate fluorescence reader, with 530 nm as the emission wavelength and 485 nm as the excitation wavelength. A cell-free condition was used to determine the background, and the fluorescence intensity in the control group was defined as 100%. Generation of intracellular reduced GSH, an index of the cellular reducing power, and T-AOC were measured using the appropriate kits according to the manufacturer’s recommended instructions. The GSH levels and T-AOC in the control group were both set to 100%.

---

Analyses of Cell Adhesion and Migration [3]
In our preliminary experiment, we found that Silybin A (Silibinin A)/SIL treatment (at concentrations of less than 20 µM) for 24 h had no effect on HepG2 cell proliferation. Therefore, we performed adhesion and migration assays with 24-h Silybin A (Silibinin A) treatments (5, 10, and 20 µM). These assays were performed as previously described. After treatment with Silybin A (Silibinin A)/SIL, the cells were centrifuged and resuspended in basal medium containing 10% fetal bovine serum. Treated cells (1×104 cells per well) were placed in a 96-well plate and incubated for 30 min at 37°C. After the cells were allowed to adhere for 30 min, they were gently washed 3 times with PBS. The adherent cells were stained with MTT and observed under an inverted phase contrast microscope. Pictures were taken using a BX61 camera. Finally, 100 µL of DMSO was added to each well, and the samples were incubated for 15 min at 37°C with shaking. The wells were measured at 490 nm on a SpectraMax 190 spectrophotometer, and the OD value in the control group was set to 100%. A cell culture wound-healing assay was performed to analyze cell migration. The cells were grown to confluence, and a linear wound was created in the confluent monolayer using a 200 µl micropipette tip. The cells were then washed with PBS to eliminate detached cells. After treatment with Silybin A (Silibinin A)/SIL(5, 10, and 20 µM) for 24 h, wound edge movement was monitored under a microscope. The results are expressed as the distance between the cells on either side of the scratch.

OVE26 mice were used as a model of type 1 diabetes mellitus and FVB mice as nondiabetic control animals. At 6 wk of age, mice were started on an animal protein-based diet (Teklad irradiated global soy protein-free extruded rodent diet-2920X) and animals were provided food and water ad libitum; at 10 wk they were started on 100 mg/kg Silybin A (Silibinin A) or vehicle given intraperitoneally for 6 wk (6 animals in each group). Urine collections were done in metabolic cages, and the urine albumin-to-creatinine ratio was measured using commercial kits for mouse albumin and creatinine. Blood glucose was measured using a glucose meter with detection range 10–600 mg/dl. The animals were killed by exsanguination under anesthesia. The cortex of one kidney was flash-frozen in liquid nitrogen for microscopy and image analysis, while the contralateral organ was fixed in 10% formalin for 24 h and then embedded in paraffin for immunohistochemistry. [5]

---

Antitumor Activity in a Xenograft Model [3]
Male athymic nude mice were used. All surgeries were performed under sodium pentobarbital anesthesia, and all efforts were made to minimize animal suffering. HepG2 cell tumor xenografts were established by subcutaneously injecting 1×106 cells into the right flanks of 4 to 6-week-old male athymic nude mice. Based on the data from a pilot study, we initiated treatment when the tumor volumes reached approximately 100 mm3. The tumor volumes (V) were calculated with the following formula: V = A×B2/2 (A = largest diameter; B = smallest diameter). First, the mice were randomly divided into 3 groups (n = 6 per group): control (saline) and Silybin A (Silibinin A) at either 200 or 400 mg/kg body weight, 5 days/week, suspended in saline and fed by oral gavage. Next, the mice were randomly divided into control, Silybin A (Silibinin A), DAPT+Silybin A (Silibinin A), and DAPT groups (n = 6). Silybin A (Silibinin A) was orally administered to the mice at doses of 400 mg/kg body weight per day (5 days/week); DAPT (10 mg/kg) or control (0.05% DMSO) was diluted with saline and administered intraperitoneally (5 days/week). Tumor sizes were measured every 3 days using calipers (days 2, 5, 8, 11, 14, 17, and 20). On day 20, tumors were excised from euthanized mice for Western blot analysis.

Metabolism / Metabolites
Silybin has known human metabolites that include O-demethylated-silybin.

31553 mouse LD50 intravenous 1056 mg/kg Yaoxue Tongbao. Bulletin of Pharmacology., 18(404), 1983
31553 rabbit LDLo unreported 300 mg/kg Yaoxue Tongbao. Bulletin of Pharmacology., 18(404), 1983

[1]. Silibinin induces apoptosis of HT29 colon carcinoma cells through early growth response-1 (EGR-1)-mediated non-steroidal anti-inflammatory drug-activated gene-1 (NAG-1) up-regulation. Chem Biol Interact. 2014 Jan 16;211C:36-43.

[2]. Silibinin induces cell death through ROS-dependent down-regulation of Notch-1/ERK/Akt signaling in human breast cancer cells. J Pharmacol Exp Ther. 2014 May;349(2):268-78.

[3]. Silybin-mediated inhibition of Notch signaling exerts antitumor activity in human hepatocellular carcinoma cells. PLoS One. 2013 Dec 27;8(12):e83699.

[4]. Silibinin Inhibits Tumor Promotional Triggers and Tumorigenesis Against Chemically Induced Two-Stage Skin Carcinogenesis in Swiss Albino Mice: Possible Role of Oxidative Stress and Inflammation. Nutr Cancer. 2014;66(2):249-58.

[5]. The antioxidant silybin prevents high glucose-induced oxidative stress and podocyte injury in vitro and in vivo. Am J Physiol Renal Physiol. 2013 Sep 1;305(5):F691-700.

Silibinin is a flavonolignan isolated from milk thistle, Silybum marianum, that has been shown to exhibit antioxidant and antineoplastic activities. It has a role as an antioxidant, an antineoplastic agent, a hepatoprotective agent and a plant metabolite. It is a flavonolignan, a polyphenol, an aromatic ether, a benzodioxine and a secondary alpha-hydroxy ketone.
Silibinin is the major active constituent of silymarin, a standardized extract of the milk thistle seeds, containing a mixture of flavonolignans consisting of silibinin, isosilibinin, silicristin, silidianin and others. Silibinin is presented as a mixture of two diastereomers, silybin A and silybin B, which are found in an approximately equimolar ratio. Both in vitro and animal research suggest that silibinin has hepatoprotective (antihepatotoxic) properties that protect liver cells against toxins. Silibinin has also demonstrated in vitro anti-cancer effects against human prostate adenocarcinoma cells, estrogen-dependent and -independent human breast carcinoma cells, human ectocervical carcinoma cells, human colon cancer cells, and both small and nonsmall human lung carcinoma cells.
Silibinin has been reported in Aspergillus iizukae, Silybum eburneum, and other organisms with data available.
Silymarin is a mixture of flavonolignans isolated from the milk thistle plant Silybum marianum. Silymarin may act as an antioxidant, protecting hepatic cells from chemotherapy-related free radical damage. This agent may also promote the growth of new hepatic cells. (NCI04)
The major active component of silymarin flavonoids extracted from seeds of the MILK THISTLE, Silybum marianum; it is used in the treatment of HEPATITIS; LIVER CIRRHOSIS; and CHEMICAL AND DRUG INDUCED LIVER INJURY, and has antineoplastic activity; silybins A and B are diastereomers.

---

Drug Indication
Currently being tested as a treatment of severe intoxications with hepatotoxic substances, such as death cap (Amanita phalloides) poisoning.

---

Podocyte injury, a major contributor to the pathogenesis of diabetic nephropathy, is caused at least in part by the excessive generation of reactive oxygen species (ROS). Overproduction of superoxide by the NADPH oxidase isoform Nox4 plays an important role in podocyte injury. The plant extract silymarin is attributed antioxidant and antiproteinuric effects in humans and in animal models of diabetic nephropathy. We investigated the effect of silybin, the active constituent of silymarin, in cultures of mouse podocytes and in the OVE26 mouse, a model of type 1 diabetes mellitus and diabetic nephropathy. Exposure of podocytes to high glucose (HG) increased 60% the intracellular superoxide production, 90% the NADPH oxidase activity, 100% the Nox4 expression, and 150% the number of apoptotic cells, effects that were completely blocked by 10 μM silybin. These in vitro observations were confirmed by similar in vivo findings. The kidney cortex of vehicle-treated control OVE26 mice displayed greater Nox4 expression and twice as much superoxide production than cortex of silybin-treated mice. The glomeruli of control OVE26 mice displayed 35% podocyte drop out that was not present in the silybin-treated mice. Finally, the OVE26 mice experienced 54% more pronounced albuminuria than the silybin-treated animals. In conclusion, this study demonstrates a protective effect of silybin against HG-induced podocyte injury and extends this finding to an animal model of diabetic nephropathy.[5]

C25H22O10

482.44

482.121

C, 62.24; H, 4.60; O, 33.16

22888-70-6

Isosilybin;72581-71-6;Silybin;802918-57-6;Silybin B;142797-34-0;Silymarin;65666-07-1

31553

White to off-white solid powder

1.5±0.1 g/cm3

793.0±60.0 °C at 760 mmHg

164-174°C

274.5±26.4 °C

0.0±2.9 mmHg at 25°C

1.684

2.59

5

10

4

35

750

4

COC1=C(C=CC(=C1)[C@@H]2[C@H](OC3=C(O2)C=C(C=C3)[C@@H]4[C@H](C(=O)C5=C(C=C(C=C5O4)O)O)O)CO)O

SEBFKMXJBCUCAI-HKTJVKLFSA-N

InChI=1S/C25H22O10/c1-32-17-6-11(2-4-14(17)28)24-20(10-26)33-16-5-3-12(7-18(16)34-24)25-23(31)22(30)21-15(29)8-13(27)9-19(21)35-25/h2-9,20,23-29,31H,10H2,1H3/t20-,23+,24-,25-/m1/s1

(2R,3R)-3,5,7-trihydroxy-2-[(2R,3R)-3-(4-hydroxy-3-methoxyphenyl)-2-(hydroxymethyl)-2,3-dihydro-1,4-benzodioxin-6-yl]-2,3-dihydrochromen-4-one

Silybin; Silibinin; silibinin; Silybin; 22888-70-6; Silybin A; Silibinin A; Silymarin I; Flavobin; Silibinine; Flavobin

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~250 mg/mL (~518.20 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (5.18 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

---

Solubility in Formulation 2: ≥ 2.5 mg/mL (5.18 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

---

View More

Solubility in Formulation 3: ≥ 2.5 mg/mL (5.18 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

---

 (Please use freshly prepared in vivo formulations for optimal results.)