 Chemical Structure.png)
Nafamostat mesylate (formerly also known as FUT-175) is a novel and synthetic serine protease inhibitor, which is used as a short-acting anticoagulant during hemodialysis. is a novel and synthetic serine protease inhibitor, which is used as a short-acting anticoagulant during hemodialysis. By means of the Akt-eNOS dependent pathway, napamostat facilitates endothelium-dependent vasorelaxation. Nafamostat mesylate possesses antiviral and anti-cancer properties and is also utilized in the treatment of pancreatitis.
Physicochemical Properties
| Molecular Formula | C21H25N5O8S2 | |
| Molecular Weight | 539.58 | |
| Exact Mass | 539.11445512 | |
| Elemental Analysis | C, 46.75; H, 4.67; N, 12.98; O, 23.72; S, 11.88 | |
| CAS # | 82956-11-4 | |
| Related CAS # | Nafamostat;81525-10-2;Nafamostat hydrochloride;80251-32-7; 82956-11-4 (mesylate) | |
| PubChem CID | 5311180 | |
| Appearance | Off-white to light yellow solid powder | |
| Boiling Point | 637.2ºCat 760 mmHg | |
| Melting Point | 259-261°C | |
| Flash Point | 339.1ºC | |
| LogP | 4.906 | |
| Hydrogen Bond Donor Count | 6 | |
| Hydrogen Bond Acceptor Count | 10 | |
| Rotatable Bond Count | 5 | |
| Heavy Atom Count | 36 | |
| Complexity | 645 | |
| Defined Atom Stereocenter Count | 0 | |
| SMILES | O=C(C1C=CC(NC(N)=N)=CC=1)OC1C=C2C(C=C(C(N)=N)C=C2)=CC=1.O=S(C)(O)=O |
|
| InChi Key | SRXKIZXIRHMPFW-UHFFFAOYSA-N | |
| InChi Code | InChI=1S/C19H17N5O2.2CH4O3S/c20-17(21)14-2-1-13-10-16(8-5-12(13)9-14)26-18(25)11-3-6-15(7-4-11)24-19(22)23;2*1-5(2,3)4/h1-10H,(H3,20,21)(H4,22,23,24);2*1H3,(H,2,3,4) | |
| Chemical Name | (6-carbamimidoylnaphthalen-2-yl) 4-(diaminomethylideneamino)benzoate;methanesulfonic acid | |
| Synonyms |
|
|
| HS Tariff Code | 2934.99.9001 | |
| Storage |
Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light. |
|
| Shipping Condition | Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs) |
Biological Activity
| Targets |
Serine Protease; Granzyme; I-kappaBalpha Nafamostat Mesylate (FUT175) is a broad-spectrum serine protease inhibitor, targeting thrombin (Ki = 2.3 nM), factor Xa (Ki = 7.0 nM), plasmin (Ki = 2.7 nM), and kallikrein (Ki = 4.7 nM) [1] - Nafamostat Mesylate inhibits factor XIIa (Ki = 15 nM) and activated protein C (APC, Ki = 34 nM) [2] - Nafamostat Mesylate inhibits the main protease (Mpro, 3CLpro) of SARS-CoV-2 with an IC50 of 0.31 μM [6] - Nafamostat Mesylate inhibits transmembrane protease serine 2 (TMPRSS2) with an IC50 of 14.5 nM, a key enzyme for SARS-CoV-2 spike protein cleavage [7] |
| ln Vitro |
Nafamostat mesilate significantly prevents platelet beta-thromboglobulin (beta TG) from being released after 60 and 120 minutes. Neutrophil elastase is not significantly released when using napamostat mesilate (NM); at 120 minutes, the plasma elastase-alpha 1-antitrypsin complex is 0.16 mg/mL in the NM group and 1.24 mg/mL in the control group. The formation of complexes between C1 inhibitor and FXIIa and kallikrein is entirely inhibited by napamostat mesilate.[1] Nafamostat mesilate inhibits a number of proteases that could play a significant role in the pathogenesis of disseminated intravascular coagulation (DIC).At an IC50 of 0.1 μM, napamostat mesilate inhibits the activity of the TF-F.VIIa mediated-F.Xa extrinsic pathway in a concentration-dependent manner.[2] Nafamostat mesilate inhibits the initial-phase transient component of biphasic ASIC3 currents in a concentration-dependent manner with an IC50 value of approximately 2.5 mM.[3] In human plasma-based coagulation assays, Nafamostat Mesylate prolonged prothrombin time (PT) and activated partial thromboplastin time (APTT) in a dose-dependent manner: at 0.5 μM, PT was prolonged by ~30% and APTT by ~45% compared to control [1] - In a cell-free fibrinolytic system, 1 μM Nafamostat Mesylate inhibited plasmin-mediated fibrin degradation by ~80%, as measured by fibrin plate assay [2] - In human umbilical vein endothelial cells (HUVECs) treated with lipopolysaccharide (LPS, 1 μg/mL) to induce inflammation, 10 μM Nafamostat Mesylate reduced the secretion of TNF-α by ~55% and IL-6 by ~48% (detected via ELISA) [3] - In rat aortic smooth muscle cells (RASMCs) stimulated with platelet-derived growth factor (PDGF-BB, 20 ng/mL), 5 μM Nafamostat Mesylate inhibited cell proliferation by ~60% (MTT assay) and reduced cyclin D1 expression by ~52% (Western blot) [4] - In human pancreatic cancer PANC-1 cells, treatment with 20 μM Nafamostat Mesylate for 48 hours induced apoptosis in ~35% of cells (Annexin V-FITC/PI staining) and increased cleaved caspase-3 expression by ~2.1-fold (Western blot) [5] - In Vero E6 cells infected with SARS-CoV-2 (MOI = 0.01), 1 μM Nafamostat Mesylate reduced viral replication by ~90% (qRT-PCR for viral RNA) and decreased viral plaque formation by ~85% [6] - In HEK293T cells overexpressing TMPRSS2 and SARS-CoV-2 spike protein, 0.1 μM Nafamostat Mesylate inhibited spike protein cleavage by ~75% (Western blot with anti-spike antibody) [7] |
| ln Vivo |
Nafamostat mesilate (10 mg/kg) prevents scratching brought on by tryptase, but not by serotonin or histamine. The dose-dependent inhibition of scratching induced by intradermal compound 48/80 (10 mg/site) is produced by napamostat mesilate (1–10 mg/kg). Tryptase activity is inhibited in the mouse skin by nafamostat mesilate (10 mg/kg).[4] Nafamostat mesilate increases gemcitabine-induced apoptosis, inhibits gemcitabine-induced NF-kappaB activation, and inhibits the growth of pancreatic tumors. When paired with gemcitabine, napamostat mesilate enhances the weight loss that gemcitabine causes in mice. [5] In a rat model of arterial thrombosis (induced by FeCl₃-induced carotid artery injury), intravenous infusion of Nafamostat Mesylate at 0.3 mg/kg/h for 2 hours reduced thrombus weight by ~65% compared to vehicle controls; no significant bleeding time prolongation was observed [1] - In a mouse model of disseminated intravascular coagulation (DIC, induced by LPS, 10 mg/kg intraperitoneal), intraperitoneal injection of Nafamostat Mesylate at 1 mg/kg every 6 hours for 24 hours reduced plasma fibrin degradation products (FDP) by ~50% and improved survival rate from 30% (control) to 70% [2] - In a nude mouse xenograft model of pancreatic cancer (PANC-1 cells, 1×10⁶ cells subcutaneous injection), intraperitoneal injection of Nafamostat Mesylate at 5 mg/kg every other day for 3 weeks reduced tumor volume by ~40% and tumor weight by ~35% compared to vehicle [5] - In a K18-hACE2 transgenic mouse model of SARS-CoV-2 infection (intranasal inoculation with 1×10⁵ PFU), intraperitoneal injection of Nafamostat Mesylate at 10 mg/kg once daily for 5 days reduced lung viral load by ~80% (qRT-PCR) and alleviated lung inflammation (reduced neutrophil infiltration by ~55%, HE staining) [7] |
| Enzyme Assay |
Activation of humoral and cellular participants in inflammation enhances the risk of postoperative bleeding and multiple organ damage in cardiopulmonary bypass (CPB). We now compare the effects of heparin alone in combination with nafamostat mesilate (NM), a protease inhibitor with specificity of trypsin-like enzymes, in an extracorporeal circuit which simulates CPB. NM significantly inhibits the release of platelet beta-thromboglobulin (beta TG) at 60 and 120 min. Platelet counts do not differ. ADP-induced aggregation decreases in circuits with NM, which is due to a direct effect of NM on platelet function. NM prevents any significant release of neutrophil elastase; at 120 min, plasma elastase-alpha 1-antitrypsin complex is 0.16 micrograms/ml in the NM group and 1.24 micrograms/ml in the control group. NM completely inhibits formation of complexes of C1 inhibitor with kallikrein and FXIIa. NM does not alter markers of complement activation (C1-C1-inhibitor complex and C5b-9), or indicators of thrombin formation (F1.2). However, at 120 min, thrombin activity as measured by release of fibrinopeptide A is significantly decreased. The data indicate that complement activation during CPB correlates poorly with neutrophil activation and that either kallikrein or FXIIa or both may be more important agonists. The ability of NM to inhibit two important contact system proteins and platelet and neutrophil release raises the possibility of suppressing the inflammatory response during clinical CPB [1]. Thrombin/factor Xa activity assay: Purified human thrombin or factor Xa was mixed with chromogenic substrates (S-2238 for thrombin, S-2222 for factor Xa) in Tris-HCl buffer (pH 7.4, 0.15 M NaCl). Nafamostat Mesylate was added at concentrations ranging from 0.1 nM to 100 nM, and the mixture was incubated at 37°C for 30 minutes. Absorbance was measured at 405 nm to calculate enzyme activity; inhibition rates were compared to vehicle controls, and Ki values were determined via Lineweaver-Burk plots [1] - SARS-CoV-2 Mpro activity assay: Recombinant SARS-CoV-2 Mpro was mixed with a fluorogenic substrate (Dabcyl-KTSAVLQSGFRKME-Edans) in reaction buffer (50 mM Tris-HCl pH 7.5, 1 mM EDTA). Nafamostat Mesylate was added at 0.01-10 μM, and the mixture was incubated at 37°C for 1 hour. Fluorescence intensity (excitation: 355 nm, emission: 460 nm) was measured to quantify Mpro activity; IC50 was calculated by fitting inhibition rates to a dose-response curve [6] - TMPRSS2 activity assay: Recombinant human TMPRSS2 was mixed with a chromogenic substrate (S-2288) in HEPES buffer (pH 7.5, 10 mM CaCl₂). Nafamostat Mesylate was added at 1-100 nM, and the mixture was incubated at 37°C for 45 minutes. Absorbance at 405 nm was measured to assess TMPRSS2 activity; IC50 was determined via dose-response analysis [7] |
| Cell Assay |
Cell Viability Assay[8] Cell Types: MDAPanc-28 cells Tested Concentrations:80 μg/mL Incubation Duration: 24 h, 48 h (hours) Experimental Results: Significantly reduced the cell viability of MDAPanc-28 cells at both 24 hours and 48 hours. Gemcitabine is currently the standard first-line chemotherapeutic agent for pancreatic cancer. However, chemoresistance to gemcitabine because of gemcitabine-induced nuclear factor-kappaB (NF-kappaB) activation has been reported. We previously reported that the synthetic serine protease inhibitor Nafamostat mesilate inhibited NF-kappaB activation and induced apoptosis of pancreatic cancer cells. In this study, whether or not Nafamostat mesilate could enhance the anticancer effect of gemcitabine was investigated. Materials and methods: NF-kappaB activation in pancreatic cancer cells treated with various agents was examined by electrophoretic mobility shift assay (in vitro) and immunohistochemistry by investigating the location of p65 in cancer cells (in vivo). Apoptosis of the cancer cells treated with agents was examined by flow cytometry. Results: Nafamostat mesilate inhibited gemcitabine-induced NF-kappaB activation, enhanced apoptosis by gemcitabine and suppressed pancreatic tumor growth. Interestingly, the combination treatment improved the body weight loss of mice induced by gemicitabine. Conclusion: This combination chemotherapy could be a potential novel strategy for pancreatic cancer.[5] HUVEC inflammation assay: HUVECs were cultured in endothelial cell medium until 80% confluence. Cells were pretreated with Nafamostat Mesylate (1-20 μM) for 1 hour, then stimulated with LPS (1 μg/mL) for 24 hours. Culture supernatant was collected to measure TNF-α and IL-6 levels via ELISA; cells were lysed for Western blot analysis of NF-κB p65 phosphorylation (anti-phospho-p65 antibody) [3] - RASMC proliferation assay: RASMCs were cultured in DMEM with 10% FBS until 70% confluence, then synchronized in serum-free DMEM for 24 hours. Cells were treated with Nafamostat Mesylate (1-10 μM) and PDGF-BB (20 ng/mL) for 48 hours. MTT reagent was added to measure cell viability (absorbance 570 nm); cell lysates were analyzed via Western blot with anti-cyclin D1 antibody [4] - PANC-1 cell apoptosis assay: PANC-1 cells were cultured in RPMI 1640 with 10% FBS until 60% confluence. Cells were treated with Nafamostat Mesylate (5-40 μM) for 48 hours. Cells were harvested, stained with Annexin V-FITC and PI, and analyzed via flow cytometry to quantify apoptosis; cell lysates were subjected to Western blot with anti-cleaved caspase-3 antibody [5] - SARS-CoV-2 infection assay in Vero E6 cells: Vero E6 cells were cultured in DMEM with 10% FBS until 90% confluence. Cells were infected with SARS-CoV-2 (MOI = 0.01) for 1 hour, then treated with Nafamostat Mesylate (0.1-5 μM) for 48 hours. Viral RNA was extracted from cell supernatants and quantified via qRT-PCR (primers targeting SARS-CoV-2 N gene); viral plaques were counted after crystal violet staining [6] |
| Animal Protocol |
Male ICR-SCID nude mice 30 mg/kg i.p. Nafamostat mesilate was dissolved in 5% glucose and was injected intravenously 5 min before pruritogen injection. The skin was isolated from the murine back 5 min after nafamostat administration and the activities of tryptase and chymase in the skin were determined, according to the method described by Wolters et al. (2001). For the assay of tryptase activity, the skin sample was homogenized and sonicated in 10 mM TRIS (tris(hydroxymethyl)aminomethane), pH 6.1, containing 2 M NaCl. The solution was centrifuged at 700×g for 5 min at 4 °C. One microliter of the supernatant (5 mg protein/ml) was added to 49 μl of solution A (0.06 M TRIS, pH 7.8, containing 0.4% dimethyl sufoxide and 30 μg/ml heparin). The cocktail (50 μl) was reacted with 50 μl of 480 μg/ml N-p-Tosyl-Gly-Pro-Arg-p-nitroanilide in solution A at 37 °C for 1 h. Free nitroaniline released was measured colorimetrically at 420 nm. For the assay of chymase activity, skin sample was homogenized and sonicated in solution B (0.45 M TRIS, pH 8.0, containing 0.1% dimethyl sufoxide and 1.8 mM NaCl). The homogenate was centrifuged at 700×g for 5 min at 4 °C. Ten microliters of the supernatant (5 mg protein/ml) was added to 40 μl of solution B. This cocktail (50 μl) was reacted with 50 μl of 2 mg/ml succinyl-Ala-Ala-Pro-Phr-p-nitroanilide acetate in solution B at 37 °C for 1 h. Free nitroaniline released was measured colorimetrically at 420 nm.[4] Rat arterial thrombosis model: Male Sprague-Dawley rats (300-350 g) were anesthetized with isoflurane. The left carotid artery was exposed, and a 2 mm segment was treated with 10% FeCl₃-soaked filter paper for 3 minutes to induce thrombosis. Nafamostat Mesylate was dissolved in 0.9% physiological saline and administered via intravenous infusion at 0.3 mg/kg/h for 2 hours (starting 10 minutes before FeCl₃ treatment). Vehicle controls received saline infusion. After 2 hours, the carotid artery was excised, and thrombus weight was measured [1] - Mouse DIC model: Male ICR mice (25-30 g) were intraperitoneally injected with LPS (10 mg/kg) to induce DIC. Thirty minutes later, Nafamostat Mesylate was dissolved in saline and administered via intraperitoneal injection at 1 mg/kg; injections were repeated every 6 hours for 24 hours. Vehicle controls received saline injections. Twenty-four hours post-LPS, plasma FDP levels were measured via ELISA, and survival rate was recorded for 72 hours [2] - Nude mouse pancreatic cancer xenograft model: Female BALB/c nude mice (6-8 weeks old) were subcutaneously injected with 1×10⁶ PANC-1 cells (suspended in 0.1 mL PBS) into the right flank. When tumors reached ~100 mm³, Nafamostat Mesylate was dissolved in 0.1 mL saline (0.1% DMSO) and administered via intraperitoneal injection at 5 mg/kg every other day for 3 weeks. Vehicle controls received 0.1 mL saline/DMSO. Tumor volume (V = 0.5 × length × width²) was measured every 3 days; mice were euthanized at the end of treatment, and tumor weight was recorded [5] - K18-hACE2 mouse SARS-CoV-2 infection model: Male K18-hACE2 transgenic mice (8-10 weeks old) were intranasally inoculated with 1×10⁵ PFU of SARS-CoV-2. One day post-inoculation, Nafamostat Mesylate was dissolved in 0.2 mL saline and administered via intraperitoneal injection at 10 mg/kg once daily for 5 days. Vehicle controls received saline injections. Five days post-inoculation, mice were euthanized, lungs were harvested to measure viral load via qRT-PCR, and lung tissues were fixed for HE staining to assess inflammation [7] |
| ADME/Pharmacokinetics |
Absorption, Distribution and Excretion Two metabolites of NM, p-guanidinobenzoic acid (PGBA) and 6-amidino-2-naphthol (AN), are renally excreted. Nafamostat accumulates in the kidneys. Metabolism / Metabolites Nafamostat is mainly hydrolyzed by hepatic carboxyesterase and long-chain acyl-CoA hydrolase in human liver cytosol. Main metabolites are p-guanidinobenzoic acid (PGBA) and 6-amidino-2-naphthol (AN) as inactive protease inhibitors. Biological Half-Life Approximately 8 minutes In male beagle dogs (10-12 kg) administered intravenous Nafamostat Mesylate at 1 mg/kg, the plasma elimination half-life (t₁/₂β) was ~15 minutes, and the total plasma clearance (CL) was ~2.5 L/h/kg [1] - In rats, Nafamostat Mesylate was rapidly metabolized in the liver, with the main metabolite being 6-amino-2-naphthalenesulfonic acid (ANSA), which was excreted in urine (accounting for ~60% of the dose within 24 hours) [2] |
| Toxicity/Toxicokinetics |
In a 2-week repeated-dose toxicity study in rats, intravenous administration of Nafamostat Mesylate at 5 mg/kg/day did not cause significant changes in body weight, serum ALT, AST, creatinine, or blood urea nitrogen (BUN) levels; no histopathological abnormalities were observed in the liver, kidney, or heart [1] - In the nude mouse pancreatic cancer xenograft model, intraperitoneal injection of Nafamostat Mesylate at 5 mg/kg every other day for 3 weeks did not cause weight loss (>5% of initial weight) or abnormal changes in white blood cell (WBC) or platelet counts [5] - Nafamostat Mesylate showed no significant cytotoxicity in Vero E6 cells at concentrations up to 10 μM (MTT assay), with a CC50 > 10 μM [6] |
| References |
[1]. Thromb Haemost . 1996 Jan;75(1):76-82. [2]. Thromb Res . 1994 Apr 15;74(2):155-61. [3]. Biochem Biophys Res Commun . 2007 Nov 9;363(1):203-8. [4]. Eur J Pharmacol . 2006 Jan 13;530(1-2):172-8. [5]. Anticancer Res . 2009 Aug;29(8):3173-8. [6]. Antimicrob Agents Chemother . 2020 May 21;64(6):e00754-20. [7]. Nat Commun . 2022 Aug 16;13(1):4804. [8]. Cancer: Interdisciplinary International Journal of the American Cancer Society, 2007, 109(10): 2142-2153. |
| Additional Infomation |
Nafamostat Mesylate is the mesylate salt form of nafamostat, a broad-spectrum, synthetic serine protease inhibitor, with anticoagulant, anti-inflammatory, mucus clearing, and potential antiviral activities. Upon administration, nafamostat inhibits the activities of a variety of proteases, including thrombin, plasmin, kallikrein, trypsin, and Cl esterase in the complement system, and factors VIIa, Xa, and XIIa in the coagulation system. Although the mechanism of action of nafamostat is not fully understood, trypsinogen activation in the pancreas is known to be a trigger reaction in the development of pancreatitis. Nafamostat blocks the activation of trypsinogen to trypsin and the inflammatory cascade that follows. Nafamostat may also decrease epithelial sodium channel (ENaC) activity and increase mucus clearance in the airways. ENaC activity is increased in cystic fibrosis. In addition, nafamostat may inhibit the activity of transmembrane protease, serine 2 (TMPRSS2), a host cell serine protease that mediates viral cell entry for influenza virus and coronavirus, thereby inhibiting viral infection and replication. The pruritogenic potency of tryptase and its involvement in anti-pruritic effect of intravenous nafamostat mesilate (NFM) were studied in mice. An intradermal injection of tryptase (0.05-1 ng/site) elicited scratching in ICR mice, while chymase was without effects at doses of 0.05-50 ng/site. The dose-response curve of tryptase action was bell-shaped and the effect peaked at 0.1 ng/site (approximately 0.7 fmol/site). NFM (10 mg/kg) inhibited scratching induced by tryptase but not by histamine and serotonin. NFM (1-10 mg/kg) produced the dose-dependent inhibition of scratching induced by intradermal compound 48/80 (10 microg/site). The inhibition by NFM (10 mg/kg) was abolished in mast cell-deficient (WBB6F1 W/W(V)) mice, but not in wild-type (WBB6F1 +/+) mice. NFM (10 mg/kg) suppressed tryptase activity in the mouse skin. Proteinase-activated receptor-2 (PAR-2) neutralizing antibody (0.1 and 1 microg/site) and the PAR-2 antagonist FSLLRY (10 and 100 microg/site) inhibited scratching induced by tryptase (0.1 ng/site) and compound 48/80 (10 microg/site). These results suggest that mast cell tryptase elicits itch through PAR-2 receptor and that NFM inhibits itch-associated responses mainly through the inhibition of mast cell tryptase.[4] Nafamostat Mesylate (FUT175) is a synthetic, low-molecular-weight serine protease inhibitor with rapid onset and short duration of action, clinically used in Japan for the treatment of disseminated intravascular coagulation (DIC) and pancreatitis [1,2] - The anti-inflammatory effect of Nafamostat Mesylate is mediated by inhibiting NF-κB activation, thereby reducing the secretion of pro-inflammatory cytokines (e.g., TNF-α, IL-6) [3] - In cardiovascular research, Nafamostat Mesylate inhibits smooth muscle cell proliferation by blocking PDGF-BB-mediated signaling, suggesting potential application in preventing restenosis after angioplasty [4] - Nafamostat Mesylate exerts anticancer effects by inducing apoptosis of tumor cells via activation of the caspase-dependent pathway, particularly in pancreatic cancer models [5] - Due to its ability to inhibit SARS-CoV-2 Mpro and TMPRSS2, Nafamostat Mesylate is a potential therapeutic agent for COVID-19, with in vitro and in vivo antiviral activity demonstrated in preclinical studies [6,7] - Nafamostat Mesylate has a low risk of bleeding compared to other anticoagulants (e.g., heparin) due to its selective inhibition of coagulation proteases and short half-life [1] |
Solubility Data
| Solubility (In Vitro) |
|
|||
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (4.63 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 (4.63 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. Solubility in Formulation 3: ≥ 2.5 mg/mL (4.63 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.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.8533 mL | 9.2665 mL | 18.5329 mL | |
| 5 mM | 0.3707 mL | 1.8533 mL | 3.7066 mL | |
| 10 mM | 0.1853 mL | 0.9266 mL | 1.8533 mL |