RCS-8   中文名称: (4-甲氧基苯基)(1-戊基-1H-吲哚-3-基)甲酮

Absorption, Distribution and Excretion
Synthetic cannabinoids (SCs) have become an increasingly important issue in forensic toxicology. Currently, there is a lack of controlled human studies evaluating the pharmacokinetic data of SCs, and published animal studies are also scarce. Therefore, interpreting analytical results from poisoned or intoxicated individuals is extremely difficult. To address this, this study investigated two selected SCs in pigs: 4-ethylnaphth-1-yl-(1-pentylindol-3-yl)methyl ketone (JWH-210) and 2-(4-methoxyphenyl)-1-(1-pentylindol-3-yl)methyl ketone (RCS-4), as well as Δ9-tetrahydrocannabinol (THC) as a reference. Each pig (n=6 for each drug) received a single intravenous injection of 200 μg/kg body weight of JWH-210, RCS-4, or THC. Six hours after administration, animals were bled, and samples of relevant organs, vital fluids (such as bile), tissues (such as muscle and adipose tissue), and atrophic meninges and vitreous fluid were collected. After hydrolysis and solid-phase extraction, analysis was performed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). To overcome the matrix effect of LC-MS/MS analysis, a standard addition method was used for quantification. The parent compound was detected in all analyzed samples except for THC, which was not detected in the dura mater and vitreous fluid. Moderate concentrations of the parent compound were present in brain tissue (the site of biological effect). Metabolite concentrations were highest in tissues involved in metabolism and/or clearance. Brain tissue, adipose tissue, and muscle tissue can be used as alternative sample sources, in addition to kidneys and lungs, which are routinely analyzed in post-mortem toxicology, especially when other samples are unavailable. Bile is the optimal sample for detecting synthetic cannabinoids (SCs) and tetrahydrocannabinol (THC) metabolites.

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Metabolisms/Metabolites
3-(4-methoxybenzoyl)-1-pentylindole (RCS-4) is a synthetic indole cannabinoid analogue first reported to the European Centre for Drug and Addiction Monitoring by Hungarian authorities through an early warning system in 2010, and subsequently discovered during trial purchases in a smoking paraphernalia shop in Ireland. We used gas chromatography-mass spectrometry to identify a series of RCS-4 metabolites in urine samples from patients hospitalized with symptoms of drug intoxication. These metabolites were initially identified as products of the following reactions: (i) aromatic monohydroxylation; (ii) dihydroxylation; (iii) aromatic hydroxylation/oxidation of the N-pentyl chain to ketone; (iv) O-demethylation; (v) O-demethylation/monohydroxylation of the N-pentyl chain; (vi) O-demethylation/oxidation of the N-pentyl chain to ketone; (vii) O-demethylation/aromatic hydroxylation/oxidation of the N-pentyl chain to ketone; (viii) N-depentylation/aromatic monohydroxylation; and (ix) N- and O-dealkylation. No parent compound was detected. O-demethylated metabolites are considered the most useful metabolic markers for identifying RCS-4 intake.
Understanding the pharmacokinetic (PK) properties of synthetic cannabinoids (SCs) is crucial for interpreting analytical results, such as those found in poisoned individuals. Due to the lack of human data from controlled studies, animal models to elucidate the pharmacokinetics of SCs must be established. Pigs, which provide a large number of biological fluid samples, were used for testing to predict human pharmacokinetic data. In this context, in addition to Δ9-tetrahydrocannabinol (THC), the metabolic pathways of two model synthetic cannabinoids (SCs) – 4-ethylnaphth-1-yl-(1-pentylindol-3-yl)methyl ketone (JWH-210) and 2-(4-methoxyphenyl)-1-(1-pentylindol-3-yl)methyl ketone (RCS-4) – were elucidated. Pig urine was collected hourly after intravenous injection of the compounds and analyzed using liquid chromatography-high resolution mass spectrometry. The following metabolic pathways were observed: JWH-210 involved hydroxylation of the ethyl or pentyl side chain, or combinations thereof, followed by glucuronidation; RCS-4 involved hydroxylation of the methoxyphenyl moiety or pentyl side chain, followed by glucuronidation, and O-demethylation, followed by glucuronidation or sulfation. The metabolic pathway of THC involves THC glucuronidation, 11-hydroxylation, followed by carboxylation and glucuronidation. Unlike THC, the parent compounds of both synthetic cannabinoids (SCs) were not detected in urine. These results are consistent with findings from human hepatocyte and/or human case studies. The urinary biomarker for JWH-210 is glucuronide of the N-hydroxypentyl metabolite (detectable for 3–4 hours), while the urinary biomarkers for RCS-4 are glucuronides of the N-hydroxypentyl, hydroxymethoxyphenyl (detectable for at least 6 hours), and O-demethylhydroxy metabolites (detectable for 4 hours). Since 2009, legislation regulating synthetic cannabinoids has spurred the emergence of new compounds to circumvent legal restrictions. 2-(4-methoxyphenyl)-1-(1-pentyl-indol-3-yl)methyl ketone (RCS-4) is a potent cannabinoid receptor agonist commonly used in herbal tobacco blends. The absence of maternal synthetic cannabinoids in urine underscores the critical role of metabolite identification in detecting RCS-4 intake in clinical and forensic investigations. We identified 18 RCS-4 metabolites, many of which are previously unreported, using 1-hour human hepatocyte incubation and time-of-flight high-resolution mass spectrometry (TOF-HRMS). Most metabolites underwent hydroxylation (with or without demethylation), carboxylation, and dealkylation, followed by glucuronidation. Additionally, a sulfated metabolite was observed. O-demethylation was the most common biotransformation and yielded the major metabolite. …We present a metabolic profile of RCS-4 obtained from human hepatocytes, including phase I and II metabolites. Metabolite structural information and associated high-resolution mass spectrometry data can be used to develop urine screening methods for RCS-4 in clinical and forensic laboratories.

Toxicity Summary
Identification and Uses: RCS-4 is a Class I controlled substance used in the manufacture of synthetic cannabis. Human Studies: Significant dose-dependent effects were observed in in vitro single-cell gel electrophoresis (SCGE) assays using human lymphocytes and oral and lung-derived human cell lines (TR-146 and A-549). These experiments, based on the measurement of DNA migration in an electric field, were able to detect single- and double-strand breaks and depurination sites. Furthermore, studies found that both drugs can induce micronucleus formation, which is caused by chromosomal aberrations. Animal Studies: RCS-4 has a high affinity for the CB1 receptor, ranging from nanomolar to nanomolar (0.59 to 22.5 nM). Compared to the CB1 receptor complete agonist CP 55940, RCS-4 exhibits nanomolar potency and can function as a complete agonist. RCS-4 reduced the motor activity of mice to less than 50% of the vector control group, and the inhibitory effect lasted for more than 1.5 hours. RCS-4 completely replaces the discriminative stimulus effect of Δ9-THC (Δ9-THC response is less than 80%). No induction of gene mutations was detected in bacterial (Salmonella/microsomal) assays.

1-Pentyl-3-[(4-methoxy)benzoyl]indole is a Category 1 controlled substance under the U.S. Drug Enforcement Administration (DEA). Category 1 controlled substances currently have no recognized medical use in the United States, lack a recognized safety profile for use under medical supervision, and have a high potential for abuse. It is a cannabinoid analogue.

C21H23NO2

321.41282

375.219

1345966-78-0

56841530

Crystalline solid

5

31.2

0

2

7

24

403

0

OZCYJKDWRUIFFE-UHFFFAOYSA-N

InChI=1S/C21H23NO2/c1-3-4-7-14-22-15-19(18-8-5-6-9-20(18)22)21(23)16-10-12-17(24-2)13-11-16/h5-6,8-13,15H,3-4,7,14H2,1-2H3

(4-methoxyphenyl)-(1-pentylindol-3-yl)methanone

BTM-8; SR-18; RCS-8

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

注意: 如下所列的是一些常用的体内动物实验溶解配方，主要用于溶解难溶或不溶于水的产品（水溶度<1 mg/mL）。 建议您先取少量样品进行尝试，如该配方可行，再根据实验需求增加样品量。

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注射用配方1: DMSO : Tween 80： Saline = 10 : 5 : 85 (如： 100 μL DMSO → 50 μL Tween 80 → 850 μL Saline)
*生理盐水/Saline的制备：将0.9g氯化钠/NaCl溶解在100 mL ddH ₂ O中，得到澄清溶液。
注射用配方 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (如： 100 μL DMSO → 400 μL PEG300 → 50 μL Tween 80 → 450 μL Saline)
注射用配方 3: DMSO : Corn oil = 10 : 90 (如： 100 μL DMSO → 900 μL Corn oil)
示例: 以注射用配方 3 (DMSO : Corn oil = 10 : 90) 为例说明, 如果要配制 1 mL 2.5 mg/mL的工作液, 您可以取 100 μL 25 mg/mL 澄清的 DMSO 储备液，加到 900 μL Corn oil/玉米油中, 混合均匀。

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注射用配方 4: DMSO : 20% SBE-β-CD in Saline = 10 : 90 [如：100 μL DMSO → 900 μL (20% SBE-β-CD in Saline)]
*20% SBE-β-CD in Saline的制备（4°C，储存1周）：将2g SBE-β-CD (磺丁基-β-环糊精) 溶解于10mL生理盐水中，得到澄清溶液。
注射用配方 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (如： 500 μL 2-Hydroxypropyl-β-cyclodextrin (羟丙基环胡精)→ 500 μL Saline)
注射用配方 6: DMSO : PEG300 : Castor oil : Saline = 5 : 10 : 20 : 65 (如： 50 μL DMSO → 100 μL PEG300 → 200 μL Castor oil → 650 μL Saline)
注射用配方 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (如： 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
注射用配方 8: 溶解于Cremophor/Ethanol (50 : 50), 然后用生理盐水稀释。
注射用配方 9: EtOH : Corn oil = 10 : 90 (如： 100 μL EtOH → 900 μL Corn oil)
注射用配方 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (如： 100 μL EtOH → 400 μL PEG300 → 50 μL Tween 80 → 450 μL Saline)

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口服配方 1: 悬浮于0.5% CMC Na (羧甲基纤维素钠)
口服配方 2: 悬浮于0.5% Carboxymethyl cellulose (羧甲基纤维素)
示例: 以口服配方 1 (悬浮于 0.5% CMC Na)为例说明, 如果要配制 100 mL 2.5 mg/mL 的工作液, 您可以先取0.5g CMC Na并将其溶解于100mL ddH2O中，得到0.5%CMC-Na澄清溶液；然后将250 mg待测化合物加到100 mL前述 0.5%CMC Na溶液中，得到悬浮液。

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口服配方 3: 溶解于 PEG400 (聚乙二醇400)
口服配方 4: 悬浮于0.2% Carboxymethyl cellulose (羧甲基纤维素)
口服配方 5: 溶解于0.25% Tween 80 and 0.5% Carboxymethyl cellulose (羧甲基纤维素)
口服配方 6: 做成粉末与食物混合

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注意: 以上为较为常见方法，仅供参考， InvivoChem并未独立验证这些配方的准确性。具体溶剂的选择首先应参照文献已报道溶解方法、配方或剂型，对于某些尚未有文献报道溶解方法的化合物，需通过前期实验来确定（建议先取少量样品进行尝试），包括产品的溶解情况、梯度设置、动物的耐受性等。

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请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
2、取适量母液，按从左到右的顺序依次添加助溶剂，澄清后再加入下一助溶剂。以 下列配方为例说明 (注意此配方只用于说明，并不一定代表此产品 的实际溶解配方):
10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
3、溶剂前显示的百分比是指该溶剂在最终溶液/工作液中的体积所占比例;
4、 如产品在配制过程中出现沉淀/析出，可通过加热(≤50℃)或超声的方式助溶;
5、为保证最佳实验结果，工作液请现配现用！
6、如不确定怎么将母液配置成体内动物实验的工作液，请查看说明书或联系我们;
7、 以上所有助溶剂都可在 Invivochem.cn网站购买。