NHC-diphosphate triammonium is the active intracellular metabolite of NHC and exists in the form of diphosphate. NHC-diphosphate triammonium is a weak substrate surrogate for the viral polymerase and is incorporated into HCV (hepatitis C virus) replicon RNA.

Physicochemical Properties

Molecular Formula	C9H24N6O12P2
Molecular Weight	470.27
Related CAS #	NHC-diphosphate;39023-73-9
Appearance	White to off-white solid powder
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: (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.
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

ln Vitro

HCV replicon cells are treated with 10 μM 3H-labeled NHC in an intracellular metabolism test. After 1, 2, and 8 hours of incubation, intracellular nucleotide levels are measured. NHC is quickly transformed into the mono-, di-, and triphosphate forms; within 8 hours, NHC-TP can reach 71.12 pM[1]. In the absence of NHC-triphosphate triammonium (NHC-TP) (3-5 μM), full-length polymerization products result, and it may be a poor substitute substrate. Furthermore, in cell-free HCV NS5B polymerization events, an evident electrophoretic shift is seen, and the molecular weight of the polymerization product increases by 16 (one extra oxygen) upon insertion of NHC-TP rather than CTP[1]. NHC (10–50 μM; 4 h) or a McGuigan phosphoramidate prodrug of NHC are cultured in Huh-7 cells. LC-MS/MS is used to evaluate the intracellular levels of the primary chemicals and phosphorylated metabolites. While NHC-triphosphate triammonium continues to be the most prevalent metabolite, trace levels of NHC-monophosphate (MP) and NHC-diphosphate (DP) are also visible[2]. The metabolite NHC-triphosphate triammonium (NHC-TP) may act as a nonobligate chain terminator and directly target the viral polymerase. Through chain termination or mutagenesis, it inhibits early negative-strand RNA synthesis in a significant way. This can potentially obstruct the proper assembly of replicase complexes.

References

[1]. Ribonucleoside analogue that blocks replication of bovine viral diarrhea and hepatitis C viruses in culture.Antimicrob Agents Chemother. 2003 Jan;47(1):244-54. [2]. Characterization of β-d- N4-Hydroxycytidine as a Novel Inhibitor of Chikungunya Virus.

Solubility Data

Solubility (In Vitro)	H2O :~160 mg/mL (~340.23 mM)
Solubility (In Vivo)	Solubility in Formulation 1: 50 mg/mL (106.32 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.1264 mL	10.6322 mL	21.2644 mL
	5 mM	0.4253 mL	2.1264 mL	4.2529 mL
	10 mM	0.2126 mL	1.0632 mL	2.1264 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.