HMG-CoA reductase (Ki = 1.3 nM/L)
Primary Target: 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase [1]
.
- Downstream Effectors: Ras and RhoA. By inhibiting HMG-CoA reductase, Cerivastatin prevents the synthesis of farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP), which are required for the prenylation and membrane translocation of Ras and RhoA, respectively. This inhibits their cell signaling functions related to proliferation and invasion [1]
.

Treatment with cerivastatin sodium (5–50 ng/mL; 3 days; MDA-MB-231 cells) reduces MDA-MB-231 cell proliferation in a dose-dependent manner (up to 40% inhibition at 25 ng/mL) [1]. After 36 hours of treatment, cerivastatin sodium (25 ng/mL; 18–36 hr; MDA-MB-231 cells) caused a cell cycle plot in the G1/S phase. The administration of cerivastatin sodium (25 ng/mL; 18 hours; MDA-MB-231 cells) significantly raises the levels of p21Waf1/Cip1 [1]. Cerivastatin sodium (MDA-MB-; 25 ng/mL; 12 hours). Matrigel-based p21 staining in MDA-MB-231 cells is modulated by cerivastatin sodium (10–25 ng/mL; 18 hours) [1]. 231 cells) therapy raises MDA-MB-231 cells' p21 levels [1]. (25 ng/mL; 18–36 hours) causes morphological alterations and delocalizes Ras and RhoA from the cell membrane to the cytoplasm [1]. Cerivastatin sodium (25 ng/mL; 4-36 hours) increases IκB while inducing NFκB inactivation in a RhoA blocking regulatory mechanism, resulting in decreased metalloprotein 9 expression and urine instability[1].

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Inhibition of Cell Proliferation: Cerivastatin induced a dose-dependent decrease in the proliferation of highly invasive MDA-MB-231 breast cancer cells, reaching a 40% inhibition at 25 ng/mL. This effect was reversed by co-incubation with mevalonate (MVA, 100 μM) or geranylgeranyl pyrophosphate (GGPP, 10 μM), but not by farnesyl pyrophosphate (FPP, 10 μM). In contrast, Cerivastatin did not significantly modify the proliferation of poorly invasive MCF-7 cells at concentrations up to 25 ng/mL [1]
.
- Absence of Apoptosis: The anti-proliferative effect was not due to apoptosis. Cerivastatin (25 ng/mL) did not induce Annexin V binding, propidium iodide incorporation, DNA fragmentation into oligonucleosomal ladders, or an increase in the sub-G1 cell population in flow cytometry. Higher concentrations (50 ng/mL) exhibited cytotoxic effects, including cell detachment [1]
.
- Cell Cycle Arrest: Cerivastatin (25 ng/mL) induced a cell cycle arrest at the G1/S phase in MDA-MB-231 cells after 36 hours of treatment (67.1% of cells in G0/G1 vs. 58.9% in controls) [1]
.
- Induction of p21Waf1/Cip1: Cerivastatin treatment (25 ng/mL, 12-18 h) markedly increased the level of the cyclin-dependent kinase inhibitor p21Waf1/Cip1 in MDA-MB-231 cells, as shown by Western blot, ELISA, and RT-PCR. This increase occurred at the transcriptional level. The effect was reversed by MVA and GGPP, but not by FPP, linking it to RhoA inhibition [1]
.
- Inhibition of Cell Invasion: Cerivastatin dose-dependently inhibited the invasion of MDA-MB-231 cells through Matrigel in a Transwell assay. At 25 ng/mL, invasion was inhibited by 54 ± 5.1% after 18 hours. This inhibitory effect was rescued by MVA and GGPP, but not by FPP [1]
.
- Delocalization of RhoA and Ras: Confocal microscopy showed that Cerivastatin (25 ng/mL) caused the delocalization of RhoA from the cell membrane to the cytosol after 18 hours of treatment. This effect was reversed by GGPP but not FPP. Delocalization of Ras was only partial and observed after 36 hours of treatment and was not reversed by either GGPP or FPP [1]
.
- Disruption of Actin Cytoskeleton: Cerivastatin treatment (25 ng/mL) induced dramatic changes in cell morphology, causing disorganization of actin stress fibers and loss of focal adhesion sites. These effects were prevented by GGPP but not by FPP [1]
.
- Inhibition of NFκB Activity: Cerivastatin decreased the constitutive NFκB activity in MDA-MB-231 cells. EMSA showed a time-dependent decrease in NFκB DNA-binding activity (complete by 36 h). Immunofluorescence demonstrated the translocation of the RelA (p65) subunit from the nucleus to the cytoplasm (starting at 18 h). This inhibition was associated with an increase in IκBα protein in the cytoplasm. The effect on NFκB was reversed by GGPP but not by FPP [1]
.
- Downregulation of Invasion-Related Genes: Cerivastatin decreased the expression of several NFκB-regulated genes involved in invasion. It reduced tissue factor (TF) expression on the cell surface by 63% after 18 h at 25 ng/mL. It also decreased urokinase-type plasminogen activator (u-PA) surface expression (by 59% after 2 days at 25 ng/mL) and reduced the secretion of metalloproteinase-9 (MMP-9) after 36 hours. Secretion of the MMP-9 inhibitor TIMP-1 was unaffected [1]
.

After penetrating the epidermis, cerivastatin sodium is rapidly absorbed and achieves its peak concentration in one to three hours. With an elimination half-life of 2-4 hours, cerivastatin sodium is substantially bound to rabbit protein (99.5%) in the bloodstream. The three respiratory chemicals in the epidermis that cerivastatin sodium mostly activates are inert, whereas the third respiratory molecule is somewhat active when combined with the parent medication. All respiratory compounds had cutoff concentrations that were lower than those of the parent medication. Waste (20–25%) and matrix (66-73%) are the methods of elimination, with nearly minimal parent material consumption [2].

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Efficacy in Patients with Primary Hypercholesterolemia: In a 4-week, randomized, double-blind, placebo-controlled, multicenter study, Cerivastatin at a total daily dose of 0.2 mg (administered as either 0.1 mg twice daily, 0.2 mg once daily with the evening meal, or 0.2 mg once daily at bedtime) significantly reduced total cholesterol, LDL cholesterol, and apolipoprotein B, and increased HDL cholesterol compared to placebo in patients with primary hypercholesterolemia [2]
.
- Onset of Action: A treatment response in LDL-C reduction was observed after 1 week of therapy, with maximal effects achieved by 3 weeks [2]
.
- Subgroup Analysis: Greater LDL-C reductions were correlated with female sex, increasing age, and lower body weight. Current cigarette smoking was correlated with lesser LDL-C reductions. The LDL-C response was not significantly associated with baseline Lp(a) levels [2]
.

HMG-CoA Reductase Inhibition Assay: The inhibitory potency of Cerivastatin against HMG-CoA reductase was determined in a bioassay. The inhibition constant (Ki) was calculated to be 1.3 nM/L, indicating a highly potent inhibition. For comparison, lovastatin exhibited a Ki of 150 nM/L under the same assay conditions [2]
.

Cell Proliferation Analysis[1]
Cell Types: MDA-MB-231 cells
Tested Concentrations: 5 ng/mL, 10 ng/mL, 25 ng/mL, 50 ng/mL
Incubation Duration: 3 days
Experimental Results: Induction of MDA-MB-231 cells There was a dose-dependent decrease in cell proliferation.

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Cell cycle analysis [1]
Cell Types: MDA-MB-231 Cell
Tested Concentrations: 25 ng/mL
Incubation Duration: 18 hrs (hours), 36 hrs (hours)
Experimental Results: Induced cell cycle arrest in G 1/S phase.

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Western Blot Analysis[1]
Cell Types: MDA-MB-231 Cell
Tested Concentrations: 25 ng/mL
Incubation Duration: 18 hrs (hours)
Experimental Results: A significant increase in p21Waf1/Cip1 levels was induced.

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RT-PCR[1]
Cell Types: MDA-MB-231 Cell
Tested Concentrations: 25 ng/mL
Incubation Duration: 12 hrs (hours)
Experimental Results: Increased p21Waf1/Cip1 mRNA levels.

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Cell Proliferation Assay: Cells (MDA-MB-231 or MCF-7) were seeded in 24-well plates at 5×10⁴ cells/well in medium with 2% fetal calf serum. They were incubated with varying concentrations of Cerivastatin for 3 (MDA-MB-231) or 5 (MCF-7) days. The medium for MCF-7 cells was renewed on day 3. After incubation, cells were detached with a non-enzymatic cell dissociation solution, and the total cell number (adherent + supernatant) was counted using a particle counter [1]
.
- Apoptosis Analysis (Annexin V/PI): Cells were washed and incubated with FITC-conjugated Annexin V and propidium iodide. Staining was analyzed by flow cytometry to detect early apoptosis (Annexin V positive) and membrane permeability (PI positive) [1]
.
- DNA Fragmentation Assay: Genomic DNA was isolated from cells, separated by electrophoresis on a 2% agarose gel, and stained with ethidium bromide. A ladder pattern of DNA fragments indicates apoptosis [1]
.
- Cell Cycle Analysis by Flow Cytometry: Nuclei were prepared using the Vindelov technique. Cells were treated with trypsin, trypsin inhibitor, and RNase A, then stained with propidium iodide. The DNA content was analyzed by flow cytometry to determine the distribution of cells in sub-G1, G0/G1, S, and G2/M phases [1]
.
- p21Waf1/Cip1 ELISA: The level of p21Waf1/Cip1 protein in nuclear extracts was measured using a commercial ELISA kit according to the manufacturer's instructions [1]
.
- p21Waf1/Cip1 and β-actin RT-PCR: Total RNA was extracted. RT-PCR was performed using specific primers for p21Waf1/Cip1 and β-actin. PCR products were analyzed on a 2% agarose gel stained with ethidium bromide [1]
.
- Matrigel Invasion Assay: Cells were detached and resuspended in serum-free medium with BSA. 5×10⁴ cells were seeded in the upper chamber of a Transwell insert coated with Matrigel. The lower chamber contained medium with BSA and basic fibroblast growth factor as a chemoattractant. After 18 hours, non-migrated cells were removed, and cells that had invaded through the Matrigel to the lower surface of the membrane were stained and counted [1]
.
- Immunofluorescence and Confocal Microscopy: Cells were fixed with paraformaldehyde, permeabilized with Triton X-100, and incubated with primary antibodies against Ras or RhoA. After washing, cells were incubated with a FITC-conjugated secondary antibody. Actin filaments were visualized with TRITC-labeled phalloidin. Images were captured using a confocal scanning laser microscope [1]
.
- Electrophoretic Mobility Shift Assay (EMSA): Nuclear extracts were incubated with a ³²P-labeled oligonucleotide containing an NFκB binding site. The DNA-protein complexes were resolved by PAGE and visualized by autoradiography. For supershift assays, nuclear extracts were pre-incubated with antibodies against RelA (p65), p50, RelB, or c-Rel [1]
.
- NFκB Localization by Immunofluorescence: Cells were fixed, permeabilized, and incubated with a primary antibody against RelA (p65). After washing, cells were incubated with a FITC-conjugated secondary antibody. Fluorescence was visualized by microscopy. Nuclei were identified by propidium iodide staining [1]
.
- Western Blot Analysis: Cytoplasmic extracts were prepared, and equal amounts of protein were subjected to SDS-PAGE. Proteins were transferred to a membrane and probed with primary antibodies against IκBα or p21Waf1/Cip1. Binding was detected with a horseradish peroxidase-coupled secondary antibody and chemiluminescence [1]
.
- Flow Cytometry Analysis of TF and u-PA: For TF, cells were incubated with a FITC-conjugated anti-TF antibody. For u-PA, cells were incubated with a primary anti-u-PA antibody, followed by a FITC-conjugated secondary antibody. Surface expression was analyzed by flow cytometry [1]
.
- Gelatin Zymography for MMP-9: Cells were incubated in serum-free medium with Cerivastatin. Conditioned medium was collected and subjected to electrophoresis on a polyacrylamide gel containing gelatin. Gels were washed, incubated overnight to allow gelatinase activity, and stained with Coomassie Blue. Gelatinolytic activity appeared as clear bands [1]
.
- TIMP-1 ELISA: The level of TIMP-1 secreted into the conditioned medium of Cerivastatin-treated cells was measured using a commercial ELISA kit [1]
.

Reduction of serum cholesterol, most notably low-density lipoprotein cholesterol is associated with reductions in cardiovascular morbidity and mortality. Statins have been shown to effectively reduce low-density lipoprotein cholesterol via inhibition of the hydroxymethyl-coenzyme A (HMG-CoA) reductase. Cerivastatin is the most potent HMG-CoA reductase inhibitor currently under study in the United States. METHODS AND RESULTS: A parallel group, randomized, placebo-controlled, double-blind, multicenter study was conducted to compare the efficacy and safety of three different dosing regimens of 0.2 mg/day of cerivastatin, a new HMG-CoA reductase inhibitor, in patients with hypercholesterolemia. After a 10-week diet-placebo lead-in period, 319 patients with low-density lipoprotein cholesterol >160 mg/dL were randomized to 4 weeks of treatment with one of the following regimens: cervastatin 0.1 mg twice daily, cerivastatin 0.2 mg once daily with the evening meal, cerivastatin 0.2 mg once daily at bedtime or placebo. All three active treatment groups produced statistically significant (P <.05) changes compared to aseline and placebo in total cholesterol (0.1 mg twice daily _18.9%; 0.2 mg once daily with the evening meal: _21.9%; 0.2 mg once daily at bedtime: _22.1%; placebo: 0.0%), low-density lipoprotein cholesterol (0.1 mg twice daily: _25.7%; 0.2 mg once daily with the evening meal: _29.4%; 0.2 mg once daily at bedtime: _30.4%; placebo: 1.4%) and high-density lipoprotein cholesterol (0.1 mg twice daily: 5.3%; 0.2 mg once daily with the evening meal: baseline and placebo, were also reduced by all active treatments (0.1 mg twice daily: _11.6% [P =.05]; 0.2 mg once daily with the evening meal: _11.6% [P =.05]; and 0.2 mg at bedtime: _10.9% [P =.07]). The percentage change in total cholesterol and low-density lipoprotein cholesterol after 4 weeks of therapy for the once-daily cerivastatin groups was statistically significantly greater (P <.05) than the cerivastatin twice daily regimen. A treatment responser was seen by 1 week of therapy and was maximal by 3 weeks. The drug was well tolerated in all three dosing regimens and resulted in no significant increase in biochemical or clinical side effects compared to placebo. CONCLUSION: Cerivastatin is a novel, highly potent, well-tolerated HMG-CoA reductase inhibitor that produces low-density lipoprotein cholesterol reductions of approximately 30% when administered at 0.2 mg once a day in the evenings.[2]

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Absorption, Distribution and Excretion
The mean absolute oral bioavailability is 60% (range 39% - 101%). Protein binding: Very high (>99%) (80% bound to albumin). Bioavailability: 60% (range 39% - 10%). Excretion: Feces (bile): 70%. Kidney: 24%. Time to peak concentration: Approximately 2.5 hours. For more complete data on absorption, distribution, and excretion of cerivastatin (8 items), please visit the HSDB record page. Metabolism/Metabolites Hepatic metabolism. The biotransformation pathways of cerivastatin in humans include: demethylation of benzyl methyl ether to M1, and methyl hydroxylation. The 6'-isopropyl moiety forms M23. Administered in its active (open acid) form. Biotransformation occurs via demethylation and hydroxylation. Certain metabolites (M1 and M23) possess pharmacological activity, with relative potencies of 50% and 100% of the parent compound, respectively. Cerivastatin is metabolized by CYP3A4 and CYP2C8; however, the drug appears to have a higher affinity for the latter enzyme. /HMG-CoA reductase inhibitor/ Known human metabolites of cerivastatin include (E)-7-[4-(4-fluorophenyl)-5-(hydroxymethyl)-2,6-di(propyl-2-yl)pyridin-3-yl]-3,5-dihydroxyhept-6-enoic acid and (E)-7-[4-(4-fluorophenyl)-6-(1-hydroxypropyl-2-yl)-5-(methoxymethyl)-2-propyl-2-ylpyridin-3-yl]-3,5-dihydroxyhept-6-enoic acid. Biological half-life: 2–3 hours. Elimination: 2–3 hours.

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Absorption: Cerivastatin is well absorbed, reaching maximal plasma levels within 1-3 hours following oral administration [2]
.
- Distribution: In the circulation, Cerivastatin is highly bound to plasma proteins (99.5%) [2]
.
- Metabolism: Cerivastatin is metabolized predominantly in the liver to three polar metabolites. Two of these metabolites are pharmacologically active, but to a lesser extent than the parent drug, while the third metabolite is inactive. Plasma concentrations of all metabolites are substantially lower than those of the parent drug [2]
.
- Elimination: The elimination half-life of Cerivastatin is 2-4 hours. Elimination of its metabolites occurs via urine (20-25%) and feces (66-73%). Essentially no parent compound is excreted unchanged [2]
.
- Accumulation: Multiple daily dosing of 0.1 to 0.4 mg indicates no significant effect on absorption or metabolism and thus no accumulation of the drug [2]
.
- Special Populations: Pharmacokinetic studies in male and female subjects in specific age groups (18-45 years and 65-85 years) indicate no relevant differences in Cerivastatin metabolism [2]
.

Protein Binding
Over 99% of circulating drugs are bound to plasma proteins (80% to albumin).

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Cerivastatin Sodium was withdrawn from the world market due to 52 reported deaths attributed to drug-related rhabdomyolysis leading to kidney failure. In the United States, there were 31 fatalities and 385 nonfatal cases of rhabdomyolysis reported among an estimated 700,000 users, most of whom required hospitalization. [3]

The risk of rhabdomyolysis was found to be higher among patients who received the full dose of Cerivastatin Sodium (0.8 mg/day). [3]

A significant drug-drug interaction was identified with concomitant use of gemfibrozil. This interaction was implicated in 12 of the 31 fatalities in the United States. [3]

Rhabdomyolysis was reported to be 10 times more common with Cerivastatin Sodium than with the other five approved statins (atorvastatin, fluvastatin, lovastatin, pravastatin, simvastatin) at the time. [3]

[1]. Cerivastatin, an inhibitor of HMG-CoA reductase, inhibits the signaling pathways involved in the invasiveness and metastatic properties of highly invasive breast cancer cell lines: an in vitro study. Carcinogenesis. 2001 Aug;22(8):1139-48.

[2]. Cerivastatin, a New Potent Synthetic HMG Co-A Reductase Inhibitor: Effect of 0.2 mg Daily in Subjects With Primary Hypercholesterolemia. J Cardiovasc Pharmacol Ther. 1997 Jan;2(1):7-16.

[3]. Withdrawal of cerivastatin from the world market. Curr Control Trials Cardiovasc Med. 2001;2(5):205-207.

Cerivastatin is (3R,5S)-3,5-dihydroxyhept-6-enoic acid, where the (7E)-hydrogen is replaced by 4-(4-fluorophenyl)-2,6-diisopropyl-5-(methoxymethyl)pyridin-3-yl. It was previously used in sodium form to lower cholesterol and prevent cardiovascular disease, but was withdrawn globally in 2001 due to reports of severe muscle toxicity. It belongs to the pyridine class of compounds, dihydroxy monocarboxylic acids, and statins (synthetic). It is the conjugate acid of cerivastatin (1-). On August 8, 2001, the U.S. Food and Drug Administration (FDA) announced that Bayer Pharmaceuticals had voluntarily withdrawn Baycol from the U.S. market due to reports of fatal rhabdomyolysis associated with the cholesterol-lowering (lipid-lowering) product. The drug had also been withdrawn from the Canadian market. Cerivastatin is a synthetic lipid-lowering drug. Cerivastatin competitively inhibits hepatic hydroxymethylglutaryl-CoA (HMG-CoA) reductase, which catalyzes the conversion of HMG-CoA to mevalonate, a key step in cholesterol synthesis. This drug lowers plasma cholesterol and lipoprotein levels and modulates the immune response by inhibiting major histocompatibility complex II on interferon-γ-stimulated antigen-presenting cells (such as human vascular endothelial cells). Muscle toxicity (myopathy and rhabdomyolysis) limits its Khellinical application. Drug Indications For adjunctive dietary therapy to reduce elevated total cholesterol and low-density lipoprotein cholesterol levels in patients with primary hypercholesterolemia and mixed dyslipidemia (Fredrickson type IIa and IIb), particularly when dietary restriction of saturated fat and cholesterol alone is ineffective, as are other non-pharmacological treatments.
FDA Label

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Mechanism of Action
Cerivastatin competitively inhibits hydroxymethylglutaryl-CoA (HMG-CoA) reductase, the enzyme responsible for converting HMG-CoA to mevalonate in the liver. Since mevalonate is a precursor to sterols such as cholesterol, this leads to decreased cholesterol levels in hepatocytes, upregulation of low-density lipoprotein (LDL) receptors, and increased uptake of LDL cholesterol from the circulation by the liver.
When statins are used in combination with fibrates or niacin, myopathy may be due to enhanced inhibition of skeletal muscle sterol synthesis (a pharmacodynamic interaction). /Statins/
Statins are a class of lipid-lowering drugs that competitively inhibit hydroxymethylglutaryl-CoA (HMG-CoA) reductase, which catalyzes the conversion of HMG-CoA to mevalonate, an early precursor to cholesterol. These drugs have structures similar to HMG-CoA and selectively and reversibly competitively inhibit HMG-CoA reductase. The high affinity of statins for HMG-CoA reductase is likely due to their binding to two different sites on the enzyme. HMG-CoA reductase inhibitors

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Cerivastatin Sodium (also known as Baycol® or Lipobay®) is a synthetic statin (HMG-CoA reductase inhibitor) that was approved by regulatory agencies based on its surrogate efficacy in lowering serum lipoproteins (e.g., LDL cholesterol). At the time of its market withdrawal, it held slightly less than 4% of the U.S. statin market. Its withdrawal highlights the limitations of relying solely on surrogate markers for drug approval, as long-term efficacy and safety data for synthetic statins like Cerivastatin Sodium were weak or nonexistent compared to the fermentation-derived statins. [3]

The withdrawal of Cerivastatin Sodium demonstrated that not all statins are interchangeable in terms of safety. It was found to be clearly inferior to other statins due to its higher risk of fatal rhabdomyolysis. In the presence of safer alternatives like simvastatin and pravastatin, the decision was made to withdraw Cerivastatin Sodium from the market to ensure patient safety. [3]

459.242

C, 64.85; H, 6.91; F, 3.95; N, 2.91; Na, 4.77; O, 16.61

143201-11-0

Cerivastatin;145599-86-6

446156

White to off-white solid powder

646.3ºC at 760 mmHg

197-199ºC

344.7ºC

1.37E-17mmHg at 25°C

4.88

3

7

11

33

620

2

COCC1=C(C(C)C)N=C(C(C)C)C(/C=C/[C@@H](O)C[C@@H](O)CC(O)=O)=C1C1=CC=C(F)C=C1

GPUADMRJQVPIAS-QCVDVZFFSA-M

InChI=1S/C26H34FNO5.Na/c1-15(2)25-21(11-10-19(29)12-20(30)13-23(31)32)24(17-6-8-18(27)9-7-17)22(14-33-5)26(28-25)16(3)4/h6-11,15-16,19-20,29-30H,12-14H2,1-5H3,(H,31,32)/q+1/p-1/b11-10+/t19-,20-/m1./s1

sodium (3R,5S,E)-7-(4-(4-fluorophenyl)-2,6-diisopropyl-5-(methoxymethyl)pyridin-3-yl)-3,5-dihydroxyhept-6-enoate

BAY-w-6228; BAY-w 6228; CERIVASTATIN SODIUM; Baycol; 143201-11-0; cerivastatin sodium salt; Rivastatin; BAY-w6228; Cerivastatin Sodium; Rivastatin; Lipobay

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: (1). This product requires protection from light (avoid light exposure) during transportation and storage.  (2). Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture.

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

H2O : ~100 mg/mL (~207.67 mM)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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