Biochemical reagent; natural alkaloid

Oxysophoridine (OSR) is a natural alkaloid extracted from Sophora alopecuroides L and has shown various pharmacological activities. A previous study demonstrated that OSR has various pharmacological actions, including antiarrhythmic, protection of cardiac muscle, antiviral, antineoplastic effects. Furthermore, OSR has antiviral pharmacological actions, which is similar to sophoridine. OSR exhibits anti-inflammatory action and inhibits the biosynthesis of leukotriene B4. The present study hypothesizes that the anti-inflammatory effect of OSR rescues SCI via anti-inflammatory, anti-oxidative stress and anti-apoptosis effects [1].

Oxysophoridine (OSR) is an alkaloid extracted from Sophora alopecuroides L and has various pharmacological activities. The present study aimed to investigate the protective effects and underlying mechanisms of OSR on spinal cord injury (SCI), a clinically common serious trauma, in a rat model. The results of the present study demonstrated that the anti‑inflammatory effect of OSR improved Basso, Beatie and Bresnahan Locomotor Rating Scale scores and reduced spinal cord tissue water contents in an SCI rat model. Inflammatory activation was measured by ELISA, and Prostaglandin E2 (PGE2), intercellular adhesion molecule‑1 (ICAM‑1), cyclooxygenase‑2 (COX‑2), nuclear factor‑κB (NF‑κB) and B‑cell lymphoma 2 (Bcl‑2)/Bcl‑2‑associated X (Bax) protein expression levels using western blotting. The results revealed that treatment with OSR reduced tumor necrosis factor‑α, interleukin (IL)‑1β, IL‑6, IL‑8 and malondialdehyde, and increased superoxide dismutase and glutathione peroxidase levels in the serum of an SCI rat model. OSR significantly reduced the protein expression of inflammation‑associated proteins PGE2, ICAM‑1, COX‑2, NF‑κB and Bcl‑2/Bax ratio in the spinal cord tissue of an SCI rat model. Furthermore, the results of the current study demonstrate that OSR ameliorates SCI via anti‑inflammatory, anti‑oxidative stress and anti‑apoptosis effects.[1]

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Oxysophoridine (OSR) is a bioactive alkaloid extracted from the Sophora alopecuroides Linn. Our aim is to explore the potential anti-inflammation mechanism of OSR in cerebral ischemic injury. Mice were intraperitoneally pretreated with OSR (62.5, 125, and 250 mg/kg) or nimodipine (Nim) (6 mg/kg) for 7 days followed by cerebral ischemia. The inflammatory-related cytokines in cerebral ischemic hemisphere tissue were determined by immunohistochemistry staining, Western blot and enzyme-like immunosorbent assay (ELISA). OSR-treated groups observably suppressed the nuclear factor kappa B (NF-κB), intercellular adhesion molecule-1 (ICAM-1), inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2). OSR-treated group (250 mg/kg) markedly reduced the inflammatory-related protein prostaglandin E2 (PGE2), tumor necrosis factor alpha (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), and interleukin-8 (IL-8). Meanwhile, it dramatically increased the interleukin-10 (IL-10). Our study revealed that OSR protected neurons from ischemia-induced injury in mice by downregulating the proinflammatory cytokines and blocking the NF-κB pathway[2].

Inflammatory activation as measured by ELISA [1]
Whole blood (500 µl) was centrifuged at 2,000 × g for 10 min at 4°C and serum was collected in every rat to determine the levels of tumor necrosis factor-α (TNF-α; H052), interleukin (IL)-1β (H002), IL-6 (H007), IL-8 (H008), malondialdehyde (MDA; A003-1), SOD (A001-1) and GSH-Px (A005) using commercial ELISA kits according to the manufacturer's protocol.

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Western blotting [1]
Spinal cord tissues were isolated from every rat and homogenized in RIPA assay. Protein concentrations were measured using a BCA protein assay kit. Proteins (50–80 µg) were fractionated by 12% SDS-PAGE and transferred to a nitrocellulose membrane. The membranes were blocked in 5% skim milk in TBS-Tween-20 (TBS-T; 0.05%) at room temperature for 1 h on a shaker and incubated with the following primary antibodies.

Animals [1]
Female adult Sprague-Dawley rats (weight, 200–230 g, n=50) were maintained in standard cages (22–24°C and 55–60% humidity) with water and food ad libitum and a 12-h light/dark cycle. All rats were randomly assigned into five groups; sham-operation group, SCI model group, 60 mg/kg Oxysophoridine (OSR) group, 120 mg/kg Oxysophoridine (OSR) group and 180 mg/kg Oxysophoridine (OSR) group. Anesthetized Sprague-Dawley rats received a midline 150 kdyne contusion injury in spinal level T10 using an Infinite Horizon impactor device, which was considered to be the SCI model. The establishment of the SCI model was confirmed by analysis of the Basso, Beatie and Bresnahan (BBB) Locomotor Rating Scale and spinal cord tissue water content. In the 60, 120 and 180 mg/kg OSR groups, SCI rats were administered intragastrically with 60, 120 and 180 mg/kg OSR once per day for 10 days. OSR was purchased from Jinghua Pharmaceutical Group Co., Ltd. (Yanchi, China). In sham-operation group and SCI model group, rats were administered normal saline intragastrically.

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Behavioral assessments [1]
Functional recovery was assessed following treatment with Oxysophoridine (OSR) using the BBB Locomotor Rating Scale to ensure consistency of the lesion. Following 10 days treatment with OSR, the rats were narcotized with 35 mg/kg of pentobarbital and then sacrificed using decollation. Subsequently, abdomen of rats was cut open, spinal level T10 was peeled and spinal cord tissues were collected and washed with PBS. Spinal cord tissues were weighed as wet weight and heated at 80°C for 48 h, and subsequently weighed as dry weight. Spinal cord tissue water content was calculated by (wet weight/dry weight) ×100.

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Experimental Design [2]
Male Institute of Cancer Research (ICR) mice weighing 25–30 g were housed in cages for 6 days at room temperature under a controlled 12 h light/dark cycle and allowed access to pellet food and water ad libitum. Mice were randomly divided into six groups. The first was the sham-treated group. The second was the vehicle-treated group, that is, ischemia was induced for 2 h of middle cerebral artery occlusion (MCAO) followed by reperfusion for 24 h. The Oxysophoridine (OSR)-treated groups were separated into low dosage group (OSR 62.5 mg/kg), moderate dosage group (OSR 125 mg/kg), and high dosage group (OSR 250 mg/kg). The sixth was the Nim-treated group (6 mg/kg). Before ischemia/reperfusion (I/R), all groups were intraperitoneally pretreated with drug or reagent (0.1 ml/10 g) for 7 consecutive days.

Interactions
The combination of Radix Angelicae sinensis (Oliv.) Diels and Radix Sophora flavescens Ait. was extensively used in traditional Chinese medicine to treat inflammatory diseases, such as acne, heart disease, and hepatitis. Sodium ferulate (SF) and oxymatrine (OMT) were effective component of Radix Angelicae sinensis (Oliv.) Diels and Radix Sophora flavescens Ait., respectively. In this study, /the authors/ investigated the synergistic anti-inflammatory effect of the combination of SF and OMT, and its modulation on inflammation-associated mediators in RAW 264.7 cells. In vivo, the anti-inflammatory effects of the combination of SF and OMT were evaluated with the xylene-induced mouse ear edema model and the carrageenan-induced rat paw edema model. In vitro, chemokines and cytokines mRNA expressions in lipopolysaccharide (LPS)-activated RAW 264.7 cells were determined by real-time PCR (RT-PCR) microarray analysis. The levels of interleukin-11 (IL-11), C-reactive protein (CRP) and interferon-gamma (INF-gamma) in the supernatant of LPS-stimulated RAW 264.7 cells were measured by enzyme-linked immune-sorbent assay (ELISA). The combination of SF and OMT could significantly inhibit the edema in the xylene-induced mouse ear edema and carrageenan-induced rat paw edema, but no effect was found when each drug was used alone according to above doses. The combination exhibited a better effect in down-regulating mRNA expressions of inflammation-associated mediators in LPS-stimulated RAW 264.7 cells than SF or OMT alone. The ELISA results showed that the combination synergistically inhibited LPS-induced IL-11, CRP and INF-gamma production in a dose-dependent manner. The combination of SF and OMT showed synergistic anti-inflammatory effect, and the activity was probably related to its modulation on inflammation-associated mediators, especially IL-11, CRP and INF-gamma.
Sodium ferulate (SF) and Oxymatrine (OMT) were compounds extracted from Chinese herbs, and have been used in clinical treatment of heart and hepatic diseases, respectively, in China for many years. The objective of this study was to examine the analgesic effect and the mechanism of the combined treatment of SF and OMT. Using the animal pain models by applying Acetic Acid Writhing Test and Formalin Test, the combination of SF and OMT showed significant analgesic effect in dose-dependent manner. In vitro, the combined treatment inhibited the increase in intracellular calcium concentration evoked by capsaicin in the dorsal root ganglion neurons. Importantly, a synergistic inhibitory effect of SF and OMT on the capsaicin-induced currents was demonstrated by whole-cell patch-clamp. Our results suggest that SF and OMT cause significant analgesic effect which may be related to the synergistic inhibition of transient receptor potential vanilloid-1.
/The aim of this was/ to study the effect of oxymatrine-baicalin combination (OB) against HBV replication in 2.2.15 cells and alpha smooth muscle actin (alpha SMA) expression, type I, collagen synthesis in HSC-T6 cells. The 2.2.15 cells and HSC-T6 cells were cultured and treated respectively. HBsAg and HBeAg in the culture supernatants were detected by ELISA and HBV DNA levels were determined by fluorescence quantitative PCR. Total RNA was extracted from HSC-T6 cells and reverse transcribed into cDNA. The cDNAs were amplified by PCR and the quantities were expressed in proportion to beta actin. The total cellular proteins extracted from HSC-T6 cells were separated by electrophoresis. Resolved proteins were electrophoretically transferred to nitrocellulose membrane. Protein bands were revealed and the quantities were corrected by beta actin. In the 2.2.15 cell culture system, the inhibitory rate against secretion of HBsAg and HBeAg in the OB group was significantly stronger than that in the oxymatrine group (HBsAg, P = 0.043; HBeAg, P = 0.026; respectively); HBV DNA level in the OB group was significantly lower than that in the oxymatrine group (P = 0.041). In HSC-T6 cells the mRNA and protein expression levels of alpha SMA in the OB group were significantly lower as compared with those in the oxymatrine group (mRNA, P = 0.013; protein, P = 0.042; respectively); The mRNA and protein expression levels of type I collagen in the OB group were significantly lower as compared with those in the oxymatrine group (mRNA, P < 0.01; protein, P < 0.01; respectively). /The authors concluded that/ OB combination has a better effect against HBV replication in 2.2.15 cells and is more effective against alpha SMA expression and type I collagen synthesis in HSC-T6 cells than oxymatrine in vitro.
Oxymatrine is proven to protect ischemic and reperfusion injury in liver, intestine and heart, this effect is via anti-inflammation and anti-apoptosis. Whether this protective effect applies to ischemic injury in brain, /the authors/ therefore investigate the potential neuroprotective role of oxymatrine and the underlying mechanisms. Male, Sprague-Dawley rats were randomly assigned to four groups: permanent middle cerebral artery occlusion (pMCAO), high dose (pMCAO+oxymatrine 120 mg/kg), low dose (pMCAO+oxymatrine 60 mg/kg) and sham operated group. /The authors/ used a permanent middle cerebral artery occlusion model and administered oxymatrine intraperitoneally immediately after cerebral ischemia and once daily on the following days. At 24 hr after MCAO, neurological deficit was evaluated using a modified six point scale; brain water content was measured; NF-kappaB expression was measured by immunohistochemistry, Western blotting and RT-PCR. Infarct volume was analyzed with 2, 3, 5-triphenyltetrazolium chloride (TTC) staining at 72 hr. Compared with pMCAO group, neurological deficit in high dose group was improved (P < 0.05), infarct volume was decreased (P < 0.001) and cerebral edema was alleviated (P < 0.05). Consistent with these indices, immunohistochemistry, Western blot and RT-PCR analysis indicated that NF-kappaB expression was significantly decreased in high dose group. Low dose of oxymatrine did not affect NF-kappaB expression in pMCAO rats. Oxymatrine reduced infarct volume induced by pMCAO, this effect may be through the decreasing of NF-kappaB expression.

[1]. Oxysophoridine rescues spinal cord injury via anti inflammatory, anti oxidative stress and anti apoptosis effects. Mol Med Rep. 2018 Feb;17(2):2523-2528.

[2]. Anti-inflammation Effects of Oxysophoridine on Cerebral Ischemia-Reperfusion Injury in Mice. Inflammation. 2015 Dec;38(6):2259-68.

Therapeutic Uses
Anti-Arrhythmia Agents; Antiviral Agents
The aim of this study was/ to evaluate the efficacy and safety of capsule oxymatrine in the treatment of chronic hepatitis B. A randomized double-blind and placebo-controlled multicenter trial was conducted. Injection of oxymatrine was used as positive-control drug. A total of 216 patients with chronic hepatitis B entered the study for 24 weeks, of them 108 received capsule oxymatrine, 36 received injection of oxymatrine, and 72 received placebo. After and before the treatment, clinical symptoms, liver function, serum hepatitis B virus markers, and adverse drug reaction were observed. Among the 216 patients, six were dropped off, and 11 inconsistent with the standard were excluded. Therefore, the efficacy and safety of oxymatrine in patients were analysed. In the capsule treated patients, 76.47% became normal in ALT level, 38.61% and 31.91% became negative both in HBV DNA and in HBeAg. In the injection treated patients, 83.33% became normal in ALT level, 43.33% and 39.29% became negative both in HBV DNA and in HBeAg. In the placebo treated patients, 40.00% became normal in ALT level, 7.46% and 6.45% became negative both in HBV DNA and in HBeAg. The rates of complete response and partial response were 24.51% and 57.84% in the capsule treated patients, and 33.33% and 50.00% in the injection treated patients, and 2.99% and 41.79% in the placebo treated patients, respectively. There was no significance between the two groups of patients, but both were significantly higher than the placebo. The adverse drug reaction rates of the capsule, injection and placebo were 7.77%, 6.67% and 8.82%, respectively. There was no statistically significant difference among them. /It was concluded that/ oxymatrine is an effective and safe agent for the treatment of chronic hepatitis B.

C15H24N2O2

264.36

264.183

54809-74-4

114850

White to off-white solid powder

208 °C

-0.35

0

2

0

19

400

4

C1C[C@@H]2[C@H]3CCC[N+]4([C@H]3[C@@H](CCC4)CN2C(=O)C1)[O-]

XVPBINOPNYFXID-LHDUFFHYSA-N

InChI=1S/C15H24N2O2/c18-14-7-1-6-13-12-5-3-9-17(19)8-2-4-11(15(12)17)10-16(13)14/h11-13,15H,1-10H2/t11-,12+,13+,15-,17?/m0/s1

(1R,2R,9S,17S)-13-oxido-7-aza-13-azoniatetracyclo[7.7.1.02,7.013,17]heptadecan-6-one

Oxymatrine; oxysophoridine; Matrine 1beta-oxide; Oxysophoridine; 54809-74-4; Sophoridine N-oxide; N-Oxysophoridine; (41S,7aS,13aR,13bR)-10-Oxohexadecahydrodipyrido[2,1-f:3',2',1'-ij][1,6]naphthyridine 4-oxide; Matrine N-oxide; Matrine oxide; Ammothamnine

2934.99.9001

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.

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

DMSO : ~25 mg/mL (~94.57 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (9.46 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.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (9.46 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.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (9.46 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.

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Solubility in Formulation 4: 100 mg/mL (378.27 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication.

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