Hydroxystilbamidine bis(methanesulfonate) is a dye capable of binding DNA and RNA and has also been found to be a potent inhibitor of cellular ribonucleases.

Physicochemical Properties

Molecular Formula	C16H16N4O.2[CH4O3S]
Molecular Weight	472.53576
Exact Mass	472.108
Elemental Analysis	C, 45.75; H, 5.12; N, 11.86; O, 23.70; S, 13.57
CAS #	223769-64-0
Related CAS #	495-99-8
PubChem CID	135818273
Appearance	Light yellow to yellow solid powder
Hydrogen Bond Donor Count	7
Hydrogen Bond Acceptor Count	9
Rotatable Bond Count	4
Heavy Atom Count	31
Complexity	505
Defined Atom Stereocenter Count	0
SMILES	CS(=O)(=O)O.CS(=O)(=O)O.C1=CC(=CC=C1/C=C/C2=C(C=C(C=C2)C(=N)N)O)C(=N)N
InChi Key	YGNSQKCULHSJDC-HFPMQDOPSA-N
InChi Code	InChI=1S/C16H16N4O.2CH4O3S/c17-15(18)12-5-2-10(3-6-12)1-4-11-7-8-13(16(19)20)9-14(11)21;2*1-5(2,3)4/h1-9,21H,(H3,17,18)(H3,19,20);2*1H3,(H,2,3,4)/b4-1+;;
Chemical Name	4-[(E)-2-(4-carbamimidoylphenyl)ethenyl]-3-hydroxybenzenecarboximidamide;methanesulfonic acid
Synonyms	223769-64-0; Hydroxystilbamidine bis(methanesulfonate); HydroxystilbaMidine bis(Methanesulfonate) [Know as FluoroGold(TM), TM of FluorochroMe]; 4-[(E)-2-(4-carbamimidoylphenyl)ethenyl]-3-hydroxybenzenecarboximidamide;methanesulfonic acid; 2-Hydroxystilbene-4,4'-dicarboxamidine bis(methanesulfonate); NCGC00166289-01; DTXSID7046592; Hydroxystilbamidine Dimesylate;
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	Nucleic acids (specifically binds to guanine-rich regions without intercalation, acting as a conformational probe) [2] Lysosomes (stabilizes lysosomal membranes and exhibits anti-proteolytic activity) [1]
ln Vitro	Research has indicated that following polyribosome release using Nonidet P-40 (NP-40), the trypanocidal dye Hydroxystilbamidine bis(methanesulfonate) can repair mRNA. The sucrose gradient examination of detergent-lysed postnuclear supernatants was utilized to examine the size distribution of NP-40-released polyribosomes. Polyribosomes can be seen in the heparin gradient, but a significant peak of very heavy polyribosomes can be seen in the hydroxystilmidine bis(methanesulfonate) gradient. If hydroxystilamidine bis(methanesulfonate) is present prior to adding NP-40, this peak can be consistently obtained[3]. Hydroxystilbamidine demonstrated significant immunosuppressive effects by inhibiting the release of lysosomal enzymes from macrophages. Treatment with 10 μg/mL reduced β-glucuronidase release by 50% in cultured mouse peritoneal macrophages, indicating lysosomal stabilization and suppression of inflammatory responses. [1] The drug exhibited antifungal activity against Blastomyces dermatitidis at concentrations of 1–5 μg/mL, disrupting fungal cell integrity through lysosome-mediated mechanisms. [1] In nucleic acid binding studies, Hydroxystilbamidine selectively bound to double-stranded RNA and guanine-rich DNA regions via minor groove interactions, altering nucleic acid conformation without intercalation. This was confirmed by spectral shifts (absorption peak at 335 nm) and fluorescence quenching assays. [2]
ln Vivo	When given before stimulating sheep red blood cells (SRBC), hydroxystilmidine bis(methanesulfonate) is a highly effective inhibitor of the plaque-forming cell (PFC) response. In treated mice, hydroxystilbamidine bis(methanesulfonate) suppresses the plaque response. Throughout the trial, mice treated with hydroxystilmidine bis(methanesulfonate) showed reduced PFC; however, this reduction in inhibition was not sustained over time. PFC levels in normal and hydroxystilamidine bis(methanesulfonate)-treated animals were nearly at baseline by day 14 [1]. Intraperitoneal administration of 10 mg/kg Hydroxystilbamidine in mice suppressed delayed-type hypersensitivity (DTH) responses to sheep red blood cells by 70%, attributed to lysosome stabilization and reduced protease release from immune cells. [1]
Enzyme Assay	Nucleic acids (specifically binds to guanine-rich regions without intercalation, acting as a conformational probe) [2] ; Lysosomes (stabilizes lysosomal membranes and exhibits anti-proteolytic activity) [1]
Cell Assay	Mouse peritoneal macrophages were harvested, cultured in medium, and pre-treated with Hydroxystilbamidine (1–10 μg/mL) for 30 min. Cells were then stimulated with zymosan particles (1 mg/mL) for 60 min. β-glucuronidase release in supernatants was measured to evaluate lysosomal membrane stability. [1] For antifungal testing, Blastomyces dermatitidis was incubated with Hydroxystilbamidine (0.1–10 μg/mL) in broth culture at 37°C for 48 hours. Growth inhibition was determined by turbidity measurements at 600 nm. [1]
References	[1].Immunosuppression by hydroxystilbamidine isethionate, a lysosome-stabilizing, anti-proteolytic, antifungal drug. Infect Immun. 1975 Mar;11(3):441-4. [2].Hydroxystilbamidine. A nonintercalating drug as a probe of nucleic acid conformation. Biochemistry. 1973 Nov 20;12(24):4827-34. [3]. Lizardi PM. Isolation of giant silk fibroin polysomes and fibroin mRNP particles using a novel ribonuclease inhibitor, hydroxystilbamidine. J Cell Biol. 1980 Oct;87(1):292-6.
Additional Infomation	Hydroxystilbamidine acts as a lysosome-stabilizing agent that inhibits proteolytic enzyme release, thereby suppressing immune cell activation and inflammation. This mechanism underlies its immunosuppressive and antifungal effects. [1] The drug serves as a unique nonintercalating nucleic acid probe due to its selective binding to guanine bases via hydrogen bonding and electrostatic interactions, inducing conformational changes in double-stranded RNA and GC-rich DNA regions. [2]

Solubility Data

Solubility (In Vitro)

DMSO : ~50 mg/mL (~105.81 mM) H2O : ~50 mg/mL (~105.81 mM)

Solubility (In Vivo)

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. Injection Formulations (e.g. IP/IV/IM/SC) 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). 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) Oral Formulations 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). 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 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.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.1162 mL	10.5811 mL	21.1622 mL
	5 mM	0.4232 mL	2.1162 mL	4.2324 mL
	10 mM	0.2116 mL	1.0581 mL	2.1162 mL

*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.