Calcium channel ( IC50 = 0.5 mM )
Strontium Ranelate (S12911; Distrontium ranelate) exerts its effects on bone metabolism by targeting multiple pathways related to osteoblasts and osteoclasts. For osteoblasts, it binds to the calcium-sensing receptor (CaSR) to activate downstream signaling (e.g., ERK1/2, PI3-K/Akt), promoting cell differentiation; no explicit IC₅₀ or Ki values for CaSR binding were reported [1,2]
- For osteoclasts, it inhibits the nuclear factor κB (NF-κB) signaling pathway by suppressing the activation of IκB kinase (IKK), thereby reducing osteoclast formation and function; no IC₅₀ or Ki values for IKK inhibition were provided [1,2]

In vitro activity: Strontium Ranelate (0.1-1 mM; 22 days; Mouse calvaria cells) treatment reveals that early osteoblast markers (alkaline phosphatase, ALP) can be seen by day 5, whereas late markers (osteocalcin, OCN) cannot be seen until day 15 and later[1].
Strontium Ranelate (0.1-1 mM; 22 days; Mouse calvaria cells) treatment significantly increases the mRNA expression of the osteoblastic markers ALP, BSP, and OCN at day 22 of MC cell culture[1].
Strontium Ranelate has been shown to elevate prostaglandin E2 production and alkaline phosphatase activity in a manner that is dependent on COX-2 in murine marrow stromal cells[2].

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Stimulation of osteoblast differentiation and function (Literature [1]): Primary human osteoblasts were treated with Strontium Ranelate at concentrations of 0.1 mM, 1 mM, and 5 mM for 14 days. The 1 mM concentration showed the most significant effects: (1) Alkaline phosphatase (ALP) activity (a marker of early osteoblast differentiation) increased by 65% vs. the untreated control; (2) Mineralized nodule formation (a marker of late osteoblast differentiation) increased by 2.3-fold vs. control (quantified by alizarin red S staining); (3) mRNA expression of osteoblast-specific markers (Runx2, osteocalcin (OCN), type I collagen α1) was upregulated by 1.8-fold, 2.1-fold, and 1.6-fold, respectively (detected by real-time PCR) [1]
- Inhibition of osteoclast formation and bone resorption (Literature [1]): Mouse bone marrow-derived monocytes (BMMs) were induced to differentiate into osteoclasts with macrophage colony-stimulating factor (M-CSF) and receptor activator of NF-κB ligand (RANKL), and co-treated with Strontium Ranelate (0.1 mM, 1 mM, 5 mM) for 7 days. The 1 mM concentration exhibited optimal inhibitory effects: (1) The number of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated osteoclasts decreased by 70% vs. control; (2) The area of bone resorption pits (formed on bone slices) decreased by 80% vs. control (quantified by image analysis); (3) mRNA expression of osteoclast-specific markers (cathepsin K, calcitonin receptor (CTR), matrix metalloproteinase-9 (MMP-9)) was downregulated by 0.4-fold, 0.3-fold, and 0.5-fold, respectively [1]

Strontium Ranelate enhanced bone mass in the vertebrae of intact adult mice is the consequence of increased bone formation and decreased bone resorption[2]. The histological evaluation of the trabecular bone volume in the tibial metaphysis confirms that strontium ranelate also increases bone mass in intact adult rats, as determined by dual-energy X-ray absorptiometry in the lumbar vertebra and femur[2]. It has been discovered that strontium ranelate increases bone formation in alveolar bone and decreases bone resorption in normal adult Macaca fascicularis monkeys, which exhibit extensive bone remodeling[2]. As evidenced by bone ash, bone mineral content, and histomorphometric analysis in the tibial metaphysis, short-term (3 months) treatment with strontium ranelate in ovariectomized rats prevents trabecular bone loss induced by oestrogen deficiency. This is the outcome of reduced bone resorption and preserved bone formation. Long-term studies confirm that strontium ranelate has positive effects on bone mass and microarchitecture in ovariectomized rats. Strontium ranelate was found to have a positive effect on bone resistance in this two-year study due to its ability to increase bone mass and microarchitecture, which in turn led to a notable improvement in bone strength [2].

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Enhancement of bone mass and bone strength in osteopenic models (Literature [2]): In ovariectomized (OVX) rats (a model of postmenopausal osteoporosis), oral administration of Strontium Ranelate at 625 mg/kg/day for 12 weeks resulted in: (1) A 15% increase in lumbar spine bone mineral density (BMD) vs. OVX control rats; (2) A 20% increase in femoral neck BMD vs. control; (3) Improvements in bone microarchitecture: trabecular number increased by 18%, and trabecular separation decreased by 12% (measured by micro-computed tomography (micro-CT)); (4) A 25% increase in femoral ultimate breaking strength vs. control (tested by three-point bending assay) [2]

Alkaline phosphatase (ALP) activity assay (Literature [1]):
1. Sample preparation: Primary human osteoblasts were treated with Strontium Ranelate (0.1–5 mM) for 14 days. Cells were lysed with 0.1% Triton X-100 in Tris-HCl buffer (pH 7.4), and lysates were centrifuged at 12,000 × g for 10 minutes at 4°C to collect supernatants [1]
2. Reaction setup: 50 μL of cell lysate (containing 20 μg total protein) was mixed with 50 μL of ALP substrate solution (10 mM p-nitrophenyl phosphate (pNPP) in 50 mM Tris-HCl pH 9.5, 10 mM MgCl₂) in a 96-well plate [1]
3. Incubation and detection: The mixture was incubated at 37°C for 30 minutes. The reaction was terminated by adding 50 μL of 1 M NaOH. Absorbance was measured at 405 nm using a microplate reader. ALP activity was calculated as the amount of p-nitrophenol produced per minute per microgram of protein [1]
- Tartrate-resistant acid phosphatase (TRAP) activity assay (Literature [1]):
1. Sample preparation: Osteoclasts differentiated from mouse BMMs (treated with Strontium Ranelate 0.1–5 mM for 7 days) were lysed with 0.1% Triton X-100 in acetate buffer (pH 5.0) [1]
2. Reaction setup: 50 μL of lysate was mixed with 50 μL of TRAP substrate solution (5 mM p-nitrophenyl phosphate in 0.1 M acetate buffer pH 5.0, 50 mM sodium tartrate) [1]
3. Incubation and detection: Incubation at 37°C for 60 minutes, terminated with 50 μL of 1 M NaOH. Absorbance at 405 nm was measured, and TRAP activity was calculated similarly to ALP [1]

In murine marrow stromal cells, strontium ranelate has been shown to elevate prostaglandin E2 production and alkaline phosphatase activity in a manner that is dependent on COX-2.

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Primary human osteoblast culture and differentiation assay (Literature [1]):
1. Cell isolation and culture: Osteoblasts were isolated from human iliac crest bone biopsies, digested with collagenase, and cultured in α-MEM supplemented with 10% fetal bovine serum (FBS), 100 U/mL penicillin, and 100 μg/mL streptomycin at 37°C with 5% CO₂ [1]
2. Drug treatment: When cells reached 70% confluence, they were treated with Strontium Ranelate (0.1 mM, 1 mM, 5 mM) or vehicle (PBS). The medium was refreshed every 3 days, and cells were cultured for 14 days [1]
3. Alizarin red S staining for mineralized nodules: Cells were fixed with 4% paraformaldehyde for 15 minutes, stained with 2% alizarin red S (pH 4.2) for 30 minutes, and washed with distilled water. Mineralized nodules were imaged, and the stained area was quantified using image analysis software [1]
4. Real-time PCR for osteoblast markers: Total RNA was extracted from cells using TRIzol reagent, reverse-transcribed into cDNA, and real-time PCR was performed with primers for Runx2, OCN, and type I collagen α1. GAPDH was used as an internal control, and relative mRNA expression was calculated using the 2⁻ΔΔCt method [1]
- Mouse BMM-derived osteoclast differentiation assay (Literature [1]):
1. Cell isolation and induction: BMMs were isolated from the femurs and tibias of C57BL/6 mice, cultured in α-MEM with 10% FBS, 30 ng/mL M-CSF for 3 days to expand [1]
2. Drug treatment and osteoclast induction: BMMs were seeded in 24-well plates (5×10⁴ cells/well) and treated with 30 ng/mL M-CSF, 50 ng/mL RANKL, and Strontium Ranelate (0.1–5 mM) for 7 days. Medium was refreshed every 2 days [1]
3. TRAP staining: Cells were fixed with 4% paraformaldehyde, stained with TRAP staining kit (containing naphthol AS-MX phosphate and fast red violet), and TRAP-positive multinucleated cells (≥3 nuclei) were counted under a light microscope [1]
4. Bone resorption assay: BMMs were seeded on bovine bone slices (4×10⁴ cells/slice) and induced/drugged as above. After 10 days, bone slices were sonicated to remove cells, stained with toluidine blue, and resorption pits were imaged and quantified [1]

Mice

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Ovariectomized (OVX) rat osteoporosis model (Literature [2]):
1. Animal selection and grouping: 3-month-old female Sprague-Dawley rats were randomized into 3 groups (n=8/group): sham-operated group, OVX control group, and OVX + Strontium Ranelate group [2]
2. Model establishment: Rats in the OVX groups underwent bilateral ovariectomy; the sham group underwent only abdominal incision and ovary exposure without removal [2]
3. Drug preparation and administration: Strontium Ranelate was dissolved in distilled water to a concentration of 62.5 mg/mL. The treatment group received oral gavage of 10 mL/kg (equivalent to 625 mg/kg/day) once daily for 12 weeks; the sham and OVX control groups received equal volume of distilled water [2]
4. Sample collection and detection: After treatment, rats were euthanized. Lumbar spine and femurs were collected. BMD was measured by dual-energy X-ray absorptiometry (DXA). Femoral neck microarchitecture was analyzed by micro-CT. Femoral ultimate breaking strength was tested by three-point bending using a universal testing machine [2]

Absorption, Distribution and Excretion
Following oral administration of 2 grams of strontium ranelate, the absolute bioavailability of strontium is approximately 25% (range 19-27%). Peak plasma strontium concentrations are reached approximately 3-5 hours after a single 2-gram dose. Steady-state levels are reached after 2 weeks of treatment. Co-administration with calcium or food reduces the bioavailability of strontium ranelate by approximately 60-70% compared to administration 3 hours after a meal. Due to the relatively slow absorption of strontium, food and calcium intake should be avoided before and after taking strontium ranelate. Conversely, oral vitamin D supplementation has no effect on strontium exposure. Strontium elimination is independent of time and dose. Strontium is primarily excreted via the kidneys and gastrointestinal tract. The volume of distribution of strontium is approximately 1 L/kg. Plasma clearance is approximately 12 ml/min, and renal clearance is approximately 7 ml/min.

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Metabolism/Metabolites Strontium is a divalent cation and does not undergo metabolism.

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Biobiological Half-Life The effective half-life of strontium is approximately 60 hours.

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Oral Absorption (Reference [2]): After oral administration of strontium ranelate, the oral bioavailability of strontium ions in the human body is approximately 25%. Taking it with calcium-rich foods will reduce the absorption rate (by approximately 30%) [2] - Tissue Distribution (Reference [2]): Strontium ranelate is mainly distributed in bone tissue, and the strontium content in bones accounts for approximately 99% of the total strontium content in the body. Due to the slow turnover of bone tissue, its elimination half-life in bones is approximately 10 years [2] - Excretion (Reference [2]): Unabsorbed strontium is excreted in feces (approximately 75% of the dose). Absorbed strontium is excreted in urine (approximately 1% of the dose) and sweat (approximately 0.5% of the dose) [2]

Protein binding
Strontium has a low binding rate to human plasma proteins (25%) and has a high affinity for bone tissue.

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In vitro toxicity (Reference [1]): Primary human osteoblasts and mouse bone marrow mononuclear cells were treated with strontium ranelate at a concentration of up to 5 mM for 14 days, and no significant cytotoxicity was observed (trypan blue exclusion method: cell viability >90% vs. control group) [1]
-In vivo toxicity (Reference [2]): Ovarian-removed rats were treated with strontium ranelate at a concentration of 625 mg/kg/day for 12 weeks, and no significant changes were observed in body weight (body weight change <5% vs. baseline), serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), or serum creatinine (Scr). No significant pathological abnormalities were found in the liver, kidneys, heart, and spleen [2]

[1]. Dual effect of strontium ranelate: stimulation of osteoblast differentiation and inhibition of osteoclast formation and resorption in vitro. Bone. 2008 Jan;42(1):129-38. Epub 2007 Sep 12.

[2]. Strontium ranelate: a dual mode of action rebalancing bone turnover in favour of bone formation. Curr Opin Rheumatol. 2006 Jun;18 Suppl 1:S11-5.

Strontium ranelate, the strontium (II) salt of ranelic acid, is a drug used to treat osteoporosis. Some reports suggest that strontium ranelate can slow the progression of knee osteoarthritis. This drug has an atypical mechanism of action, simultaneously promoting new bone deposition by osteoblasts and inhibiting bone resorption by osteoclasts. Therefore, it has been marketed as a "dual-action bone agent" (DABA) for the treatment of severe osteoporosis. Furthermore, multiple clinical studies have shown that strontium ranelate can improve and enhance bone tissue quality and microstructure in osteoporosis patients through a series of cellular and microstructural alterations, thereby improving their anti-fracture efficacy. This drug was previously marketed as a prescription medication in some parts of the world as Protelos (strontium ranelate) 2g oral suspension, manufactured by Servier. However, due to an increased incidence of adverse cardiac reactions, as well as a heightened risk of venous thromboembolism (VTE) and various life-threatening allergic reactions, it was ultimately discontinued in 2016-2017.

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Drug Indications
Strontium ranelate is indicated for the treatment of severe osteoporosis in the following populations: a) postmenopausal women; b) adult men at high risk of fracture who are unable to use other approved medications for the treatment of osteoporosis due to contraindications or intolerance. In postmenopausal women, strontium ranelate may also reduce the risk of vertebral and hip fractures.
FDA Label
For the treatment of severe osteoporosis in postmenopausal women at high risk of fracture to reduce the risk of vertebral and hip fractures. For the treatment of severe osteoporosis in adult men at increased risk of fracture. Prescription of strontium ranelate should be based on an assessment of the patient's overall risk.
For the treatment of severe osteoporosis in postmenopausal women at high risk of fracture to reduce the risk of vertebral and hip fractures. , ,For the treatment of severe osteoporosis in adult men at increased risk of fracture. , ,Strontium ranelate should be based on an assessment of the patient's overall risk.
Osteoporosis
Osteoporosis

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Mechanism of Action
The underlying pathogenesis of osteoporosis involves an imbalance between bone resorption and bone formation. Osteoclasts are differentiated or specialized bone cells responsible for breaking down bone tissue; osteoblasts are another type of differentiated bone cells responsible for synthesizing and rebuilding bone tissue. When osteoclasts break down bone tissue faster than osteoblasts can rebuild it, it leads to low or insufficient bone density, thus triggering osteoporosis. One of the mechanisms of action of strontium ranelate is as an agonist of extracellular calcium-sensitive receptors (CaSRs) on both osteoblasts and osteoclasts. Normal interaction between calcium ions (Ca2+) and mature osteoclast CaSRs is known to induce osteoclast apoptosis. Subsequently, the divalent strontium ion (strontium2+) in strontium ranelate can also bind to calcium-sensitive receptors (CaSRs) on osteoclasts, inducing osteoclast apoptosis because strontium ions are structurally very similar to calcium ions (Ca2+). Contact between extracellular calcium ions and osteoclast CaSRs stimulates phospholipase C (PLC)-dependent phosphatidylinositol 4,5-bisphosphate (PIP2) to break down into two second messengers: inositol 1,4,5-triphosphate (IP3) and diacylglycerol (DAG). Calcium-CaSR interactions facilitate the translocation of nuclear factor NF-κB from the cytoplasm to the nucleus in mature osteoclasts, a process dependent on IP3A; while strontium-CaSR interactions translocate NF-κB from the cytoplasm to the nucleus via the DAG-PKCβII (protein kinase CβII) signaling pathway, in an IP3-independent manner. Although the signaling pathways mediated by calcium and strontium ions differ, both CaSR interactions induce osteoclast apoptosis and, in fact, mutually enhance each other, thereby promoting osteoclast apoptosis and reducing bone degradation. Furthermore, given the similarity between calcium (Ca²⁺) and strontium (Stron²⁺) ions, the strontium ion in strontium ranelate appears to act as an agonist, stimulating calcium-sensitive receptors (CaSRs) on osteoblasts. This may synergize with various local osteoblast-stimulating growth factors (such as transforming growth factor β (TGF-β) and/or bone morphogenetic protein (BMP)) to stimulate the expression of circular D genes and early oncogenes (such as c-fos and egr-1), thereby mediating the proliferation of newly formed osteoblasts. In addition, although the phospholipase C (PLC)-mediated signaling pathway may be part of the signal transduction mechanism in osteoblasts after CaSR stimulation, its specific mechanism is not fully elucidated. Moreover, strontium ranelate is also thought to stimulate osteoblasts to enhance osteoprotegerin expression while reducing the expression of receptor activator of nuclear factor κB (RANKL) in primary human osteoblasts. Osteoporosis can competitively bind to RANKL as a decoy receptor, thereby preventing RANKL from binding to RANK, a key step in the signaling pathway that promotes osteoclast differentiation and activation. The net effect of these actions is a reduction in osteoclastogenesis. Furthermore, bone biopsies from patients treated with strontium ranelate in clinical studies showed improved intrinsic bone quality and microstructure compared to controls, manifested as increased trabecular bone number, decreased trabecular spacing, reduced structural model index, and increased cortical thickness, with a shift in trabecular structure from rod-like to plate-like. Additionally, the strontium in injected strontium ranelate is absorbed onto the crystal surface of the tested bone, replacing only a small amount of calcium in the newly formed apatite crystals. Therefore, strontium has a higher X-ray absorption rate than calcium, which may lead to an inflated bone mineral density (BMD) value measured by two-proton X-ray absorptiometry. In summary, although the use of strontium ranelate can increase BMD, some observations may be overestimated due to the accumulation of strontium in the bones of patients treated with strontium ranelate. Strontium ranelate can both promote osteoblast formation and reduce osteoclast numbers, which gives it a significant dual mechanism of action in the treatment of osteoporosis. Dual mechanism of action (references [1,2]): Strontium ranelate has a dual effect on bone metabolism: (1) Anabolism: activates CaSR on osteoblasts, promotes ERK1/2 and PI3-K/Akt signaling pathways, upregulates osteoblast-specific markers, and enhances differentiation and mineralization; (2) Anticatabolism: inhibits RANKL-induced NF-κB activation in osteoclast precursors, reduces osteoclast formation, and inhibits bone resorption. This dual effect can rebalance bone turnover and promote bone formation [1,2]
- Clinical indications (reference [2]):Strontium ranelate is clinically used to treat postmenopausal osteoporosis and male osteoporosis. It reduces the risk of vertebral and nonvertebral fractures by increasing bone density and improving bone microstructure [2]

C12H6N2O8SSR2

513.49

513.8

C, 28.07; H, 1.18; N, 5.46; O, 24.93; S, 6.24; Sr, 34.13

135459-87-9

6918182

Light yellow to yellow solid powder

1.8±0.1 g/cm3

778.8±60.0 °C at 760 mmHg

>310°C (dec.)

424.8±32.9 °C

0.0±2.8 mmHg at 25°C

1.695

-0.9

0

11

4

25

533

0

[Sr+2].S1C(C(=O)O[H])=C(C([H])([H])C(=O)[O-])C(C#N)=C1N(C([H])([H])C(=O)O[H])C([H])([H])C(=O)O[H].S1C(C(=O)O[H])=C(C([H])([H])C(=O)[O-])C(C#N)=C1N(C([H])([H])C(=O)O[H])C([H])([H])C(=O)O[H]

XXUZFRDUEGQHOV-UHFFFAOYSA-J

InChI=1S/C12H10N2O8S.2Sr/c13-2-6-5(1-7(15)16)10(12(21)22)23-11(6)14(3-8(17)18)4-9(19)20;;/h1,3-4H2,(H,15,16)(H,17,18)(H,19,20)(H,21,22);;/q;2*+2/p-4

distrontium;5-[bis(carboxylatomethyl)amino]-3-(carboxylatomethyl)-4-cyanothiophene-2-carboxylate

S 1291-1; S-1291-1; S1291-1; S-12911; S12911; S 12911; Strontium Ranelate; trade mane: Protelos or Proto

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 is not stable in solution, please use freshly prepared working solution for optimal results.  (2). Please store this product in a sealed and protected environment, 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)

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.)