Part 1:using the potassium manganese to product the ketone group , then synthesis the quinazolinocarboline alkaloids. part 2:use the electrooxidation reaction to product tropine then synthesis the atropine alkaloids.

Abstract In the present study, the effects of rutaecarpine on ionic currents of NG108-15 neuronal cells were studied. Rutaecarpine (2-100 £gM) suppressed the amplitude of voltage-dependent K+ outward current (IK(DR)) in a concentration-dependent manner. The IC50 value for rutaecarpine-induced inhibition of IK was 11£gM. However, rutaecarpine (20 £gM) had little effect on L-type Ca2+ current. IK(DR) present in these cells is sensitive to the inhibition by quinidine and dendrotoxin, yet not by E-4031. Rutaecarpine enhanced the rate and extent of IK(DR) inactivation, although it had no effect on the initial activation phase of IK(DR). Recovery from block by rutaecarpine (5 £gM) was fitted by a single exponential with a value of 2.87 s. Cell-attached single-channel recordings revealed that rutaecarpine decreased channel activity over the length of the test potential without altering single-channel amplitude. With the aid of binding scheme, a quantitative description of the actions of rutaecarpine on IK(DR) was provided. Under current-clamp configuration, rutaecarpine also prolonged action potential duration in NG108-15 cells without altering other variables of the action potential. The results clearly show that rutaecarpine is a blocker of the KDR channel. The increase in action potential duration induced by rutaecarpine can be explained mainly by its blocking effects on IK(DR).

The aryl hydrocarbon receptor is a ligand-activated transcription factor that is involved in many important functions in the body. To study the role and function of AHR, an abundant amount of in vitro expressed and purified protein is needed. A baculovirus insect expression system is commonly employed to express AHR, however, there are several drawbacks with this method, such as mutation potential and high cost. A better in overexpression system is needed and we hypothesize that Pichia pastoris, a yeast expression system, could stably express AHR and ARNT (aryl hydrocarbon receptor nuclear translocator) in sufficient amount with reasonable cost. Codon optimized human AHR and ARNT genes were separately transformed into the Pichia pastoris genome and expressed. Co-immunoprecipitation, gel-shift assay and western analysis indicate Pichia pastoris was able to stably overexpress functional AHR and ARNT proteins in comparable yield and lower cost compared to baculovirus insect expression system and the expressed proteins were used to develop a new in vitro method to study AHR and ARNT binding. Pharmacokinetic studies were performed to investigate the role of AHR in rutacarpine-caffeine interactions. Oral RUT pretreatment was shown to reduce oral caffeine area under the curve (AUC) in rats, due to an increase in caffeine clearance (CL) and a decrease in oral bioavailability (F). RUT, an AHR ligand, increases caffeine CL by inducing Cyp1A2 enzyme, but the mechanism by which RUT reduces caffeine F is not understood. We hypothesize that it is also mediated via AHR pathway. To test the hypothesis, wild type (WT) and AHR knock out (KO) mice were administered caffeine IV and orally, with and without VEH or RUT pretreatment. As expected, PK data show higher caffeine CL and lower F values in WT, but similar CL and F values in AHR KO mice, upon RUT co-administration. Rats study, in which with pretreatment of vehicle, AHR ligands: RUT, beta-naphthoflavone or indole-3-carbinol before caffeine was dosed orally is consistent with mice study, that all three AHR agonists tested were able to reduce oral caffeine AUCs in rats. RUT reduces caffeine’s oral bioavailability is through AHR signaling pathway, however, However, the mechanism by which AHR mediates the reduced caffeine F is not known.

Benzo[a]pyrene (BaP), is one of the most potent mutagens and carcinogens known. It requires metabolic activation through cytochrome P450 (CYP)1A1 to yield the ultimate carcinogenic metabolite, benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide (BPDE). BPDE can bind to DNA and form predominantly covalent (+) trans adducts at the N2 position of guanine causing DNA damage. Rutaecarpine (RTC) is an herbal medicine that has been used to treat several diseases such as headache, hypertension, gastrointestinal disorders, amenorrhea, and anti-inflammation. It has also been reported as a potent inducer of CYP enzymes, including CYP1A1, and CYP1A2. The mechanisms underlying up-regulation of CYP1A1 by RTC is dependent on aryl hydrocarbon receptors. Meanwhile, RTC can inhibit the activity of CYP1A1, CYP1A2 and CYP1B1. To investigate the effect of RTC on the BaP-induced DNA damage, we analyzed the CYP1A1 enzyme activity and DNA damage level in two cell lines, namely mucoepidermoid pulmonary carcinoma cells (H292) and hepatocellular carcinoma cells (Hep3B). The cells either were treated with only 5 μM BaP or 1.25, 2.5, 5 and 10 μM RTC, respectively; or were co-administrated 5 μM BaP and one of the four concentrations of RTC for 24 hours. Ethoxyresorufin-O-deethylase (EROD) assay was used to detect CYP1A1 enzyme activity. The results showed that both BaP and RTC significantly (p<0.05) induced CYP1A1 enzyme activity when administered separately, with RTC induction exhibiting a concentration-dependent manner. Interestingly, co-administration of RTC with BaP, especially at high concentration (10 μM) of RTC, induced less CYP1A1 enzyme activity compared to either only RTC or BaP administration. MuseTM Multi-Color DNA Damage kit was used to evaluate the DNA damage level in cells. The data showed that the DNA damage induced by BaP alone was about 2-fold higher (p&;lt;0.05) than that by concurrent administration of RTC and BaP. In conclusion, our data showed that although both RTC and BaP are inducers of CYP1A1 enzyme, their co-administration will reduce CYP1A1 enzyme activity compared with BaP administration alone. The DNA damage kit results supported that there is a potential protective effect of RTC against BaP-induced DNA damage in both H292 and Hep3B cells.

Many people like to drink caffeinated beverages, such as coffee or tea, but are sensitive to effects of caffeine. Therefore, they either avoid drinking caffeinated beverages altogether, or they avoid drinking them close to bedtime to prevent caffeine from interfering with their sleep. Ruta Cleanse and Ruta Sleep are natural supplements containing rutaecarpine that are designed to speed up the removal of caffeine from the blood. The recommendation is to take two capsules (equivalent to 100 mg rutaecarpine), as needed, to reduce caffeine level. Customers have reported positive effects, when taken 30 minutes to 2 hours prior. However, there is no scientific data to show how soon Ruta Cleanse and Ruta Sleep need to be taken in order for it to work. Therefore, we tested in rats the effect of single dose after 3, 6, 12, 24 hour and 7 doses (once a day, for seven days) of oral 100 mg/kg rutaecarpine (in suspension) induction on caffeine pharmacokinetics upon 15 mg/kg intravenous bolus and 20 mg/kg oral caffeine doses. The MROD data showed that as early as 3 hours after oral rutaecarpine administration, CYP1A2 activity in the liver tissue is increased by almost 3-fold compared to control rats and highest activity (9-fold compared to control) is found in the liver of rats administered with daily oral dose of rutaecarpine, for seven days. A suspension form of 100 mg/kg orally administered rutaecarpine significantly decreases the oral systemic exposure and mean residence time of caffeine and its metabolites (paraxanthine, theophylline and theobromine), as early as 3 hours before oral caffeine administration. The oral caffeine bioavailability (F) decreases by about 50% for the 3, 6 and 12-hour, 70% for the 24 hour and 80% for the one week daily rutaecarpine treatment groups. Currently we do not know the mechanism by which rutaecarpine significantly decreases the F values of caffeine upon oral administration. The systemic exposure of caffeine and its metabolites are also decreased when caffeine is given intravenously, though the effect is less pronounced compared to when caffeine is given orally. Interestingly, rutaecarpine achieves this effect without achieving detectable plasma level (less than 10 ng/mL). However, since the target organ for rutaecarpine is the liver, rutaecarpine can still induce CYP1A2 enzyme in the liver (as indicated by MROD data), without having to get absorbed into blood circulation.