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Clinical tolerability and pharmacokinetics of Erigerontis hydroxybenzene injection: results of a randomized phase I study in healthy Chinese volunteers.

ABSTRACT

Multiple phenolic compounds in the extract of Erigeron breviscapus synergistically contribute to the neurovascular protective effects. We conducted a phase I and pharmacokinetic study with the phenolic compound-enriched product extracted from Erigeron breviscapus, Erigerontis hydroxybenzenes injection (EHI), in healthy Chinese volunteers.

A randomized, open-label, single-center, double-arm, dose-escalation study of EHI was conducted. The tolerability of intravenously EHI administrated in single- or multiple-dose (once daily for 7 days) was studied in 40 healthy Chinese volunteers and the pharmacokinetics of EHI was studied in additional 10 volunteers.

The tolerated dose of intravenous infusion of EHI in healthy Chinese volunteers was 6 vials (equivalent to 90 mg bioactive phenolic compounds). The main limitations to dose escalation of EHI were transit changes in electrocardiogram and mild, transit increase in alanine aminotransferase. After intravenous administration of EHI, the average systemic clearance of multiple phenolic compounds of scutellarin, 1,3-dicaffeoylquinic acid, 3,5-dicaffeoylquinic acid, and 3,4-dicaffeoylquinic acid were 131,29, 262,112 L/h for male volunteers and 202,28,252,117 L/h for female volunteers.

The intervention of intravenous infusion of EHI in healthy Chinese volunteers was generally tolerated. The findings from this study provide data on the tolerability and pharmacokinetics of the extract from Erigeron breviscapus and support further trials.

Keywords:

Erigeron breviscapus

Tolerability

Pharmacokinetics

Introduction

The extract from Erigeron breviscapus (Vant.) Hand.-Mazz. (Deng-zhan-xi-xin) has been approved by China Food and Drug administration (CFDA, www.sfda.gov.cn) to be used in clinical setting for the treatment of neurovascular diseases (cerebral embolism, cerebral thrombosis, coronary heart disease, and angina pectoris etc.). Its reported clinical indications include the treatment of visual field defects, blood stasis and stagnation, apoplectic hemiplegia, extremity numbness, speech dysphasia, chest stuffiness and pain, as well as similar syndromes caused by ischemic stroke, coronary heart disease and angina pectoris (Flan et al. 2012; Zhong et al. 2010). It has been suggested that multiple bioactive phenolic compounds in Erigeron breviscapus synergistically contributed to these pharmacological effects (Chai et al. 2013; Sun and Zhao 2009). These phenolic compounds have been recognized and identified (Li et al. 2012, 2013; Liu et al. 2008; Zhang et al. 2007), including scutellarin, 1,3-dicaffeoylquinic acid (1,3-DCQA), 1,4-dicaffeoylquinic acid (1,4-DCQA), 3,5-dicaffeoylquinic acid (3,5DCQA), 4,5-dicaffeoylquinic acid (4,5-DCQA), 3,4-dicaffeoylquinic acid (3,4-DCQA), 4-caffeoylquinic acid (4-CQA), 9-caffeoyl-2,7anhydro-2- octulosonic acid (9-COA), quercetin-3-O-glucuronide, eriodictyol-7-O-glucuronide, erigoster A and B, erigeside 1,3-caffeoyl2,7- anhydro-3-deoxy-2-octulopyranosonic acid (3-CDOA).

Scutellarin is a bioactive flavonoid in controlling vascular disease (Wang et al. 2011). Based on various in vitro studies, scutellarin has been shown to inhibit the coagulation (Zhou et al. 1992), inhibit high glucose-mediated vascular inflammation (Luo et al. 2008), and promote angiogenesis in human umbilical vein endothelial cells (Gao et al. 2010). While, based on in vitro systems, dicaffeoylquinic acid compounds have been indicated to have potent superoxide anion radical scavenging activity (Heilmannet al. 1995; Kim and Lee 2005); 3.5- DCQA was suggested to be a potential therapeutic agent for treating or preventing neurodegenerative diseases implicated with oxidative stress (Kim et al. 2005); in addition, 3,4-DCQA has been shown to strongly inhibit human platelet aggregation (Chang and Hsu 1992).

Erigerontis hydroxybenzenes injection (EHI) is a phenolic compound-enriched product extracted from Erigeron breviscapus. To maintain the stability of phenolic compounds and prolong the shelf life of the product, EHI product is developed into lyophilized powder. In the extraction rate of phenolic compounds from the herb of Erigeron breviscapus, EHI product can achieve 1.2% compared to the traditional extraction rate of 0.2-0.6%. Additionally, the content of scutellarin, 3.5- DCQA, 1,5-DCQA, 4,5-DCQA, and erigoster B in EHI product were controlled to be [greater than or equal to] 80% (Sun and Zhao 2009).

The extract of Erigeron breviscapus has been safely used in Chinese medicine for centuries. Although preclinical studies in animals have offered scientific basis of the effectiveness and tolerability of EHI, no data was available concerning the tolerability and pharmacokinetics in humans. In the present study, we for the first time evaluated clinical tolerability and pharmacokinetics of intravenous EHI in healthy Chinese volunteers.

Methods

Study volunteers

The study protocol was approved by the Institutional Review Board (1RB# 2003L04455) of the China Food and Drug Administration (Beijing, China). After clinical assessment of health status, 50 adult healthy Chinese volunteers over 21 years of age (half male/half female, age: 24 [+ or -] 3 years) were selected for this study. Exclusion criteria included; (1) clinical laboratory test abnormalities, (2) clinically significant electrocardiogram (ECG) abnormalities, (3) receiving enzyme-inducing or enzyme-inhibiting drugs, (4) a history of migraine. All the volunteers were informed of the aim and risk involved in the study and written informed consent were obtained. From September to November, 2004, this study was performed in the care units of the Affiliated Hospital of Nanjing University of Chinese Medicine (Nanjing, China). During the study period, all volunteers took standard meal and were recorded adverse drug reactions that occurred. During the study, all volunteers avoided strenuous exercise and drinking of tea, coffee and alcohol drinks.

Drug treatment and study design

This was a randomized, open-label, single-center, double-arm, dose-escalation trial. Forty volunteers were studied for the tolerability of the product of EHI administered in a single dose or multiple doses once daily for 7 days (Table 2) and additionally 10 volunteers were studied for the pharmacokinetics of EHI. This study aimed to test the tolerability, and aimed to assess its pharmacokinetics in humans. Intravenous EHI infusion was given to each volunteer with an infusion time of 1 h after dissolution of appropriate dose of EHI product into 250 mL 0.9% sodium chloride solution. Tested EHI product (per vial containing a total of 15 mg bioactive phenolic compounds based on its label information, lot no. 20040702) in the pre-marketing phase was manufactured by Spirin Biotechnology Co. (Yunnan, China). The quality of EHI product is controlled due to the fact that the content of phenolic compounds in the herb were reported to be correlated with environmental factors (Li et al. 2013). The quality control standard is that the total content of scutellarin, 3,5-dicaffeoylquinic acid (3,5-DCQA), 1.5- dicaffeoylquinic acid (1,5-DCQA), 4,5-dicaffeoylquinic acid (4,5-DCQA), and erigoster B in EHI product were defined to be [greater than or equal to] 80%. Using HPLC chemical quantification in our laboratory, per vial tested EHI product contained 6.20 mg scutellarin, 0.21 mg 1,3-DCQA, 2.74 mg 3,5-DCQA, and 3.50 mg 3,4-DCQA.

To investigate the clinical tolerability of EHI, 28 volunteers (half male/half female) received a single dose of EHI through intravenous infusion at 5 escalating levels: level 1 (2 vials, equivalent to 30 mg bioactive ingredients, n = 4), level 2 (4 vials, equivalent to 60 mg bioactive ingredients, n = 6), level 3 (6 vials, equivalent to 90 mg bioactive ingredients, n = 6), level 4 (9 vials, equivalent to 135 mg bioactive ingredients, n = 6), level 5 (12 vials, equivalent to 180 mg bioactive ingredients, n = 6); another 12 volunteers (half male/half female) received multiple doses of EHI (once daily for 1 week) at 2 levels: level 4 (9 vials, equivalent to 135 mg bioactive ingredients, n = 6) and level 5 (12 vials, equivalent to 180 mg bioactive ingredients, n = 6). The randomization code was generated by an independent staff member and was kept in sealed opaque envelopes until the study was completed. The starting dose of EHI (level 1,30 mg bioactive ingredients) in humans was estimated based on the preclinical data of no observed adverse effect level (NOAEL) of EHI in animal species (Sun and Zhao 2009) according to FDA Guidance for Industry - Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutics in Adult Healthy Volunteers (FDA 2005). In preclinical studies, it has been shown that beagles were more sensitive to the treatment of EHI than the species of mice or rats (Sun and Zhao 2009). Once a starting dose level has been determined, dose increments of EHI were determined based on a modified Fibonacci dose-escalation method with the incremental ratio of 2.00, 1.50, and then 1.33 for subsequent dose-levels (Penel and Kramar 2012). Dose selection in multiple doses of EHI was wide due to that the multiple lower doses of EHI (levels 1 (30 mg), 2 (60 mg), and 3 (90 mg), respectively) in our preceding small-scale results was safe and well-tolerated. The principle for dose escalation was to avoid exposing too many subjects to subtherapeutic doses while preserving safety.

To investigate the clinical pharmacokinetics of EHI, 10 volunteers (half male/half female, age: 24.1 [+ or -] 3.0 years, body weight: 58.0 [+ or -] 7.1 kg) received a single dose of EHI through intravenous infusion: 6 vials (equivalent to 90 mg bioactive ingredients).

Tolerability and safety assessments

Tolerability and safety were assessed according to the Common Terminology Criteria for Adverse Events (CTCAE) by National Institutes of Health and National Cancer Institute, on the basis of (1) physical status measured before dose and 24 h after each dose, including blood pressure (blood pressure measurements were made in the same arm every time), pulse, body temperature, and physical examinations; (2) subjective and objective adverse events assessed continuously before dose and after each dose; (3) clinical laboratory tests performed by the Department of Laboratory Testing & Pathology at the Affiliated Hospital of Nanjing University of Chinese Medicine, including hematology (red blood cell count, hemoglobin, white blood cell count, granulocytes, lymphocytes, platelet, prothrombin time, international normalized ratio, activated partial thromboplastin time), biochemistry (blood urea nitrogen, serum creatinine, aspartate aminotransferase (AST), alanine transaminase (ALT), albumin, globulin, alkaline phosphatase, total bilirubin and direct bilirubin), and urinalysis (urine protein, urine glucose); laboratory tests for single-dose tolerability were done at baseline and 24 h after dose; laboratory tests for multipledose tolerability were done at baseline and 24 h after the last dose; (4) electrocardiogram (ECG) recorded continuously upon 2 h after each dose and evaluated by a clinical cardiologist; (5) follow-up measurements up to 1 week after the last dose. All adverse events that occurred during the study were recorded, together with the duration, severity, and any measures. The investigator then judged, from the tolerability and safety profile, whether the study could proceed to the next step.

Blood sampling and bio-analysis of drug plasma concentrations

To investigate the pharmacokinetics of intravenous EHI, blood samples were collected into heparinized tubes (green tops) from forearm vein before the administration and from the other arm at 0.17, 0.5, 1.0, 1.1, 1.2, 1.3, 1.7, 2.0, 2.5, 3.0, 4.0, 5.0 h after the start of intravenous infusion of EHI. Plasma was separated following centrifugation of blood samples and stored at -75[degrees]C until use. The concentrations of scutellarin, 1,3-DCQA, 3,5-DCQA, and 3,4-DCQA in human plasma samples were determined using ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) (Quattro Premier, Waters, Milford, MA, USA). During the bio-analysis, the reference standard of scutellarein (HPLC purity = 96.2%) were purchased from ChromaDex Corporation (Irvine, CA, USA); the reference standards of 1,3-DCQA (HPLC purity = 98.0%), 3,5-DCQA (HPLC purity = 98.0%), and 3,4-DCQA (HPLC purity = 98.0%) were purchased from the National Institute for the Control of Pharmaceutical and Biological Products (Beijing, China). In brief, 250 [micro]L plasma was mixed with 10 [micro]L internal standard (rutin, IS, 1.0 [micro]g/mL rutin). Then the mixture was acidified with 25 [micro]L [H.sub.3]P[0.sub.4] (20%, v/v) and extracted with 1.2 mL ethyl acetate. After centrifugation, 1 mL of supernatant was transferred into another tube and evaporated to dryness under a gentle stream of nitrogen. The residue was reconstituted into 150 [micro]L 90% methanol, vortexed, and then centrifuged. Exactly 3.5 [micro]L supernatant was injected onto the UPLC-MS/MS system.

Chromatographic separation of analytes was performed onto an Acquity UPLC BEH C18 column (1.7 [micro]m, 2.1 x 50 mm; Waters). Mobile phase A was water containing 0.1% formic acid. Mobile phase B was acetonitrile-isopropanol (1:1, v/v) containing 0.1% formic acid. The analytes were eluted from the column with a linear gradient from 5% B to 95% B in 6.5 min, then maintained at 95% B for 0.5 min before returning to initial condition. The flow rate was 0.4 mL/min. The retention times were 3.7, 1.8, 4.2, 5.9, and 3.9 min for scutellarin, 1,3-DCQA, 3,5-DCQA, 3,4-DCQA, and rutin (IS), respectively. A tandem mass spectrometer equipped with electro spray ionization source operated in positive ionization mode was used as detector. Multiple reaction monitoring (MRM) using the precursor to product ion pairs for quantification of scutellarin, DCQA, and rutin (IS) was m/z 463 [right arrow] 287, 517 [right arrow] 163, and 611 [right arrow] 303. The MS parameters were optimized for individual analyte. The chromatographic separations of scutellarin, 1,3-DCQA, 3,5-DCQA, and 3,4-DCQA are shown in Fig. 1.

Calibration curves for the quantification of analyte concentrations in plasma samples were constructed using weighted (1/[X.sup.2]) linear regressions of the ratio of peak area of each analyte to that of IS (Y) against the corresponding nominal concentrations (X) of scutellarin (2.6, 5.3, 13, 26, 52, 132, 264, and 528 ng/mL), 1,3-DCQA (1.5, 3.1, 6.1, 15, 31, and 61 ng/mL), 3,5-DCQA (1.3, 3.2, 6.4, 13, 32, 64, and 128 ng/mL), 3,4-DCQA (1.8,3.6,8.9,18,36,89,178, and 355 ng/mL) in blank plasma. The LLOQs of scutellarin, 1,3-DCQA 3,5-DCQA, and 3,4-DCQA were 2.6,1.5,1.3, and 1.8 ng/mL, respectively. The bio-analytical method was fully validated (Table 1) according to FDA Bioanalytical Method Validation guidance (FDA 2013).

[FIGURE 1 OMITTED]

Pharmacokinetic data analysis and statistical analysis

Plasma concentrations of scutellarin and DCQAs were used for pharmacokinetic data analysis with Phoenix software (Pharsight Corp, Mountain View, CA, USA). The following pharmacokinetic parameters for scutellarin and DCQAs were determined, peak plasma concentration ([C.sub.max]) was read directly from the data, area under the curve (AUC) was determined using the trapezoidal rule, first order elimination rate constant (X) as calculated by least-square regression analysis of terminal ln-linear portions of the plasma concentrationtime profile during post-infusion phase, and terminal phase half-life ([t.sub.1/2]) was calculated by 0.693 divided by K.

The equations required for computation of pharmacokinetic parameters were:

during infusion phase, when t < 1 h, [C.sub.o-t] = Q/VK x (1 - [e.sup.-Kt]) (1)

during post - infusion phase,

when t > 1 h, [C.sub.1h-t] = [Q/VK x (1 - [e.sup.-K])] x [e.sup.-K(t-1)] (2)

where C is the plasma concentration at time t, Q is the zero order input rate, and V is the volume of distribution.

In the above equations, the assumption was one compartment model with zero order input (constant infusion rate) and first order elimination rate.

Results

Study subject characteristics

Table 2 summarizes the demographics and baseline characteristics of study volunteers recruited in clinical tolerability study. Forty healthy volunteers were enrolled in the single-dose and multiple doses of intravenous infusion of EHI for clinical tolerability study, another 10 volunteers received a single dose of EHI for pharmacokinetic study. No subject withdrew during the study.

Tolerability and tolerability studies

Table 3 lists the adverse drug reactions stratified by dose levels in the single-dose tolerability study of EHI. No adverse drug reactions were observed in any volunteers when given intravenous EHI up to the dose level 3. A total of 3 (10.7%) of the 28 volunteers at EHI dose levels 4 and 5 experienced a total of 3 adverse drug reactions (1 adverse events and 2 ECG abnormalities). One female volunteer at EHI dose level 5 experienced a mild dizziness, and the symptoms disappeared soon. One female volunteer at EHI dose level 4 experienced a transit change in ECG (minor T wave flattening) and one male volunteer at EHI dose level 5 experienced a transit change in ECG (sinus tachycardia). Both ECG changes recovered the next day after dose. The severity of the above adverse events was of grade 1 according to CTCAE.

Table 4 lists the adverse drug reactions stratified by dose levels in multiple dose tolerability study of EHI. A total of 7 (58.3%) of the 12 volunteers experienced a total of 12 adverse drug reactions (7 adverse events, 3 clinical laboratory test abnormalities, and 2 ECG abnormalities). Most of the adverse events, including dizziness, headache, nausea, diarrhea, nasal obstruction, were mild and disappeared during the study. One male volunteer at dose level 4 experienced a mild increase in ALT, a mild increase in urine protein, and a change in ECG (minor T wave flattening); one female volunteer at dose level 5 experienced a mild increase in urine glucose; and one female individual at dose level 5 also experienced a change in ECG (minor T wave flattening). The severity of the above adverse events was of grade 1 according to CTCAE. All above changes recovered within 7-day follow up.

Pharmacokinetic studies

Fig. 2 displays the mean plasma concentration-time profiles for scutellarin, 1,3-DCQA, 3,5-DCQA and 3,4-DCQA after administration of a single-dose of EHI (6 vials). The plasma concentrations of scutellarin, 1,3-DCQA, 3,5-DCQA and 3,4-DCQA decreased significantly after EHI infusion stopped. Peak plasma concentrations of scutellarin in male volunteers were higher than those in female volunteers. Table 5 demonstrates the main pharmacokinetic parameters for scutellarin, 1,3-DCQA, 3,5-DCQA and 3,4-DCQA. For scutellarin, there were significance gender differences on the peak plasma concentrations ([C.sub.max]) (P < 0.05) and on the area under the plasma concentration-time curve (AUC, [AUC.sub.0[right arrow]1h] x [AUC.sub.0[right arrow]5h], [AUC.sub.0[right arrow][infinity]] (P < 0.001); while for DCQAs, there was no significant gender difference (P > 0.05) in values of [C.sub.max] or AUC, by two sample t-test. There was no gender difference on the terminal phase half-life for either scutellarin or DCQAs, by two sample t-test.

Discussion

This is the first study to our knowledge evaluating the clinical tolerability and pharmacokinetics of the extract from Erigeron breviscapus in healthy Chinese volunteers. Historically and still currently, herbal supplements are monitored under the Dietary Supplement Health and Education Act of 1994 (DSHEA). Because of this classification, before being marketed, they usually do not undergo clinical safety testing required by FDA authority. While, the current study with the extract from Erigeron breviscapus is to provide proactive insights about the clinical safety testing for herbal supplements.

[FIGURE 2 OMITTED]

We have shown that the extract from Erigeron breviscapus, EH1 given by intravenous infusion exhibited an acceptable tolerability profile in 40 Chinese volunteers. Based on our conservative approach in assessing toxicides, we determined intravenous infusion of EHI at 6 vials (equivalent to 90 mg bioactive phenolic compounds) might be a clinically safe and tolerated dose. After intravenous infusion of EHI at higher dose, only mild drug reactions (mild adverse events, mild increase in ALT, mild increase in urine protein and glucose, and mild change in ECG) were observed (Tables 3 and 4). In the multiple doses of EHI administration in these volunteers, higher incidence of adverse drug reactions after given at 9 vials was observed than that at 12 vials (Table 4). Interpretation of these adverse drug reactions and ascribing them to the drug treatment rather than to an alternative cause seemed to be not straightforward. Given the small sample size in each dose group, it was difficult to differentiate predictable and dose-related adverse drug reactions and unpredictable and doseindependent adverse drug reactions (Tohkin et al. 2010). However, all these mild drug reactions disappeared at follow up. In addition, out of a total of 40 volunteers, 10 (5 female/5 male, 25%) volunteers experienced adverse drug reactions. There was no significant gender difference in the incidence of adverse drug reactions (P > 0.05) after administration of EHI.

Additionally, pharmacokinetics of the four active phenolic compounds of scutellarin, 1,3-DCQA, 3,5-DCQA and 3,4-DCQA after intravenous administration of EHI were studied in 10 Chinese volunteers. It was observed that there were large inter-individual variations in the plasma pharmacokinetic profile of scutellarin, 1,3-DCQA, 3,5-DCQA and 3,4-DCQA in 10 Chinese volunteers (Fig. 2). Pharmacokinetics of scutellarin in humans has only been reported by Chen et al. (2006). In their study, they observed similar large inter-individual variations in the plasma concentrations of scutellarin (<5 ng/mL) and its major metabolite scutellarein 6-0-[beta]- d -glucuronide (38-128 ng/mL) after oral administration of 60 mg of scutellarin in 20 male Chinese healthy volunteers. The pharmacokinetics of 1,3-DCQA, 3,5-DCQA, 3,4-DCQA in humans could be differential, even for structural isomers. Pharmacokinetics of 3,5-DCQA and 3,4-DCQA in humans have been reported after consumption of coffee (Monteiro et al. 2007). Likewise, they observed large inter-individual variation in the pharmacokinetic profile of all DCQA compounds.

The plasma concentration of each compound at a given time (C) was influenced by the infusion rate (Q), the volume of distribution (V), and the first order elimination rate constant (K), according to the Eqs. (1) and (2). Therefore, the drug concentration during the phase of drug infusion (t < 1 h) was determined by drug infusion rate (Q) and systemic clearance (CL = V x K)\ the drug concentration during the phase of drug clearance (t > 1 h) was determined by drug plasma concentration achieved at 1 h and the elimination rate constant (K). In the current study, during the phase of drug infusion, there was gender difference in pharmacokinetics of scutellarin after intravenous administration of EHI in Chinese volunteers (Fig. 2, Table 5). In our study, the peak plasma concentrations of scutellarin in male volunteers were higher than those in female volunteers (P < 0.05). This gender difference in drug concentrations of scutellarin might not be induced by body weight that might influence drug distribution, due to the fact that the body weight within this group of volunteers (59 [+ or -] 5 kg in male vs 60 [+ or -] 4 kg in female) did not differ significantly (P > 0.05). Rather, this gender difference in scutellarin concentrations might be attributed to the gender difference in systemic clearance of scutellarin (P < 0.001) (Table 5). In the current study, it was also observed that the plasma concentrations of 3,5-DCQA were only about 25% of those of 3,4-DCQA (Fig. 2), despite similar amounts of 3,5-DCQA and 3,4-DCQA in EH1 products. Significant differences in the volume of distribution (from 56 to 1672 L) and the systemic clearance values (from 28 to 262 L/h) of 1,3-DCQA, 3,5-DCQA and 3,4-DCQA in these Chinese volunteers were observed. Their biotransformation and tissue uptake for these compounds in humans need further investigation.

Larger-scale studies are needed to establish a relationship between drug concentrations and potential clinical efficacy/toxicity of EHI. In the current study, linear pharmacokinetics was not determined given that only single dose pharmacokinetics for intravenous EHI has been studied here. The plasma concentrations of metabolites derived from either DCQAs or CQAs from EHI should be determined to elucidate the potential toxicity of EHI. It has been speculated that CQA molecules may undergo metabolism by cytochrome P450 (P450) enzymes, horseradish peroxidase/[H.sub.2][O.sub.2], and tyrosinase/[O.sub.2] to form cytotoxic quinone intermediate metabolites and cause adverse effects to human health, due to a catechol and an [alpha], [beta]-unsaturated carbonyl group in the chemical structures (Xie et al. 2012). In addition, due to the fact that healthy volunteers were used in this study, caution is advised in extrapolating tolerability data as well as pharmacokinetic data from this study to actual patient populations who might have abnormal liver and kidney function.

In conclusion, intravenous EHI showed acceptable tolerability in healthy Chinese volunteers and the pharmacokinetics after single dose of intravenous EHI was characterized in this study.

Conflict of interest

The authors declared no conflict of interest.

ARTICLE INFO

Article history:

Received 21 May 2014

Revised 9 November 2014

Accepted 25 November 2014

Acknowledgments

This project was support by Leading Talents of Scientific Research in Traditional Chinese Medicine of Jiangsu Province, China (No. LJ200906). We are thankful to Xiang Long from Spirin Biotechnology Co. Ltd (Yunnan, China) for the help and Ms. Kate Yu from Waters (Milford, MA, USA) for guidance in the bio-analysis.

References

Chai, L., Guo, H., Li, H., Wang, S., Wang, Y.L, Shi, F., Hu, LM., Liu, Y., Adah, D., 2013. Scutellarin and caffeic acid ester fraction, active components of Dengzhanxixin injection, upregulate neurotrophins synthesis and release in hypoxia/reoxygenation rat astrocytes. J. Ethnopharmacol. 150,100-107.

Chang, W.C., Hsu, F.L., 1992. Inhibition of platelet activation and endothelial cell injury by poiyphenolic compounds isolated from Lonicerajaponica Thunb. Prostaglandins Leukot Essent Fatty Acids 45,307-312.

Chen, X., Cui, L., Duan, X., Ma, B., Zhong, D., 2006. Pharmacokinetics and metabolism of the flavonoid scutellarin in humans after a single oral administration. Drug Metab. Dispos. 34, 1345-1352.

FDA, 2005. Guidance for Industry Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutics in Adult Healthy Volunteers. U.S. Department of Health and Human Services, Food and Drug Administration, Center for Drug Evaluation and Research (CDER). http://www.fda.gov/downloads/ Drugs/Guidances/UCM078932.pdf.

FDA, 2013. Guidance for industry Bioanalytical Method Validation. US Department of Health and Human Services, Food and Drug Administration, http://www.fda.gov/ downloads/Drugs/GuidanceComplianceRegulatorylnformation/Guidances/ UCM368107.pdf.

Gao, Z.X., Huang, D.Y., Li, H.X., Zhang, LN., Lv, Y.H., Cui, H.D., Zheng, J.H., 2010. Scutellarin promotes in vitro angiogenesis in human umbilical vein endothelial cells. Biochem. Biophys. Res. Commun. 400,151-156.

Han, Y.L, Li, D., Ren, B., Jing, C.P., Meng, X.L., Zhou, Z.Y., Yu, Q., Li, Y., Wan, LL., Guo, C., 2012. Evaluation of impact of Herba Erigerontis injection, a Chinese herbal prescription, on rat hepatic cytochrome P450 enzymes by cocktail probe drugs. J. Ethnopharmacol. 139,104-109.

Heilmann, J., Merfort, L, Weiss, M., 1995. Radical scavenger activity of different 3',4dihydroxyflavonols and 1,5-dicaffeoylquinic acid studied by inhibition of chemiluminescence. Planta Med. 61,435-438.

Kim, H.J., Lee, Y.S., 2005. Identification of new dicaffeoylquinic acids from Chrysanthemum morifolium and their antioxidant activities. Planta Med. 71,871-876.

Kim, S.S., Park, R.Y.,Jeon, H.J., Kwon, Y.S., Chun, W., 2005. Neuroprotective effects of 3,5dicaffeoylquinic acid on hydrogen peroxide-induced cell death in SH-SY5Y cells. Phytother. Res. 19,243-245.

Li, F., Zhang, L.D., Li, B.C., Yang, J., Yu, H., Wan, J.B., Wang, Y.T., Li, P., 2012. Screening of free radical scavengers from Erigeron breviscapus using on-line HPLC-ABTS/DPPH based assay and mass spectrometer detection. Free Radic. Res. 46, 286-294.

Li, X., Peng, LY., Zhang, S.D., Zhao, Q.S., Yi, T.S., 2013. The relationships between chemical and genetic differentiation and environmental factors across the distribution of Erigeron breviscapus (Asteraceae). PLoS One 8, e74490.

Liu, C.Z., Gao, M., Guo, B., 2008. Plant regeneration of Erigeron breviscapus (vant.) Hand. Mazz. and its chromatographic fingerprint analysis for quality control. Plant Cell Rep. 27,39-45.

Luo. P., Tan, Z.H., Zhang, Z.F., Zhang, H., Liu, X.F., Mo, Z.J., 2008. Scutellarin isolated from Erigeron multiradiatus inhibits high glucose-mediated vascular inflammation. Yakugaku Zasshi 128,1293-1299.

Monteiro, M., Farah, A., Perrone, D., Trugo, LC., Donangelo, C., 2007. Chlorogenic acid compounds from coffee are differentially absorbed and metabolized in humans. J. Nutr. 137,2196-2201.

Penel, N., Kramar, A, 2012. What does a modified-Fibonacci dose-escalation actually correspond to? BMC Med. Res. Methodol. 12,103

Sun, H.D., Zhao, Q.S., 2009. A drug for treating cardio-cerebrovascular diseases phenolic compounds of Erigeron breviscapus. Prog. Chem. 21,77-83.

Tohkin, M., Ishiguro, A., Kaniwa, N., Saito, Y., Kurose, K., Hasegawa, R., 2010. Prediction of severe adverse drug reactions using pharmacogenetic biomarkers. Drug Metab. Pharmacokinet. 25,122-133.

Wang, Y., Ao, H., Qian, Z., Zheng, Y., 2011. Intestinal transport of scutellarein and scutellarin and first-pass metabolism by UDP-glucuronosyltransferase-mediated glucuronidation of scutellarein and hydrolysis of scutellarin. Xenobiotica 41, 538-548.

Xie, C., Zhong, D., Chen, X., 2012. Identification of the ortho-benzoquinone intermediate of 5-O-caffeoylquinic acid in vitro and in vivo: comparison of bioactivation under normal and pathological situations. Drug Metab. Dispos. 40,1628-1640.

Zhang, Y., Shi, P., Qu, H., Cheng, Y., 2007. Characterization of phenolic compounds in Erigeron breviscapus by liquid chromatography coupled to electrospray ionization mass spectrometry. Rapid Commun. Mass Spectrom. 21, 2971-2984.

Zhong, Y., Xiang, M., Ye, W., Cheng, Y., Jiang, Y., 2010. Visual field protective effect of Erigeron breviscapus (vant.) Hand. Mazz. extract on glaucoma with controlled intraocular pressure: a randomized, double-blind, clinical trial. Drugs R D 10, 75-82.

Zhou, Q.S., Zhao, Y.M., Bai, X., Li, P.X., Ruan, C.G., 1992. Effect of new- breviscapine on fibrinolysis and anticoagulation of human vascular endothelial cells. Zhongguo Yao LiXue Bao 13,239-242.

http://dx.doi.org/10.1016/j.phymed.2014.11.014

Wen-Zheng Ju (a,1), Yang Zhao (b,1) *, Fang Liu (a), Ting Wua, Jun Zhang (a), Shi-Jia Liu (a), Ling Zhou (c), Guo-Liang Dai (c), Ning-Ning Xiong (a), Zhu-Yuan Fang (a,d)

(a) Department of Clinical Pharmacology, The Affiliated Hospital of Nanjing University of Chinese Medicine, Nanjing, China

(b) Department of Pharmaceutical Sciences, University of Pittsburgh, PA, USA

(c) Nanjing University of Chinese Medicine, Nanjing, China

(d) Department of Cardiovascular Sciences, The Affiliated Hospital of Nanjing University of Chinese Medicine, Nanjing, China

* Corresponding author at: Department of Pharmaceutical Sciences, University of Pittsburgh, 3501 Terrace St. 716 salk Pittsburgh, Pennsylvania 15213, USA. Tel.: +14126247913; fax: +14126487671.

E-mail addresses: wzju333@hotmail.com (W.-Z. Ju), zhaoyang4110@gmail.com (Y. Zhao).

(1) The first two authors contributed equally to the work.
Table 1
Bioanalytical method validation for the analytes of scutellarin,
1,3-DCQA, 3,5-DCQA, and 3,4-DCQA in human plasma samples.

Bioanalytical method     Scutellarin             1,3-DCQA
validation term

Retention time, min      3.7                     1.8
Mass transition, m/z     463 [right arrow] 287   517 [right arrow] 163
Extraction recovery, %   44-53                   71-73
Matrix effects, %        101-104                 105-113
Sensitivity, ng/mL       2.6                     1.5
Accuracy, %              86-115                  93-113
Precision                2-7                     2-7
  (coefficient of
  variation), %
Stability after          109-113                 95-109
  storage in
  autosampler at 4
 [degrees]C for 24
  h, %
Stability after          104-113                 89-114
  storage at -20
 [degrees]C for 14
  days, %
Stability after three    106-115                 98-107
  freeze/thaw cycles
  at -20[degrees]C, %

Bioanalytical method     3,5-DCQA                3,4-DCQA
validation term

Retention time, min      4.2                     5.9
Mass transition, m/z     517 [right arrow] 163   517 [right arrow] 163
Extraction recovery, %   77-86                   77-87
Matrix effects, %        102-112                 98-108
Sensitivity, ng/mL       1.3                     1.8
Accuracy, %              86-112                  87-114
Precision                5-9                     5-7
  (coefficient of
  variation), %
Stability after          85-94                   85-101
  storage in
  autosampler at 4
 [degrees]C for 24
  h, %
Stability after          89-112                  94-106
  storage at -20
 [degrees]C for 14
  days, %
Stability after three    86-95                   86-99
  freeze/thaw cycles
  at -20[degrees]C, %

Note: Quality control samples at three plasma concentrations for
scutellarin (4.4,35, and 352 ng/mL), 1,3-DCQA (4.6,15, and 46 ng/mL),
3,5-DCQA (6.4, 38, and 90 ng/mL), and 3,4-DCQA (4.7, 24, and 237
ng/mL) were assayed to obtain the bioanalytical method validation
parameters of accuracy, precision, and recovery. The peak area of
the stability samples under testing conditions were compared with
that of control samples.

Table 2
Baseline characteristics of the Chinese volunteers used in
tolerability study.

Characteristics                 Single-dose trial

                                Level 1            Level 2

No. of volunteers               4                  6
Dose, vials intravenously       2                  4
Age, mean [+ or -] SD, years     24 [+ or -] 2.6    24 [+ or -] 1.9
Weight, mean [+ or -] SD, kg     60 [+ or -] 4      54 [+ or -] 7
Height, mean [+ or -] SD, cm    167 [+ or -] 4     161 [+ or -] 8
SBP, mean [+ or -] SD, mmHg     115 [+ or -] 7     102 [+ or -] 8
DBP, mean [+ or -] SD, mmHg      75 [+ or -] 6      68 [+ or -] 6
Pulse, mean [+ or -] SD,         83 [+ or -] 7      78 [+ or -] 5
  beats/min

Characteristics                 Single-dose trial

                                Level 3            Level 4

No. of volunteers               6                  6
Dose, vials intravenously       6                  9
Age, mean [+ or -] SD, years     25 [+ or -] 3.7    24 [+ or -] 2.4
Weight, mean [+ or -] SD, kg     57 [+ or -] 9      53 [+ or -] 6
Height, mean [+ or -] SD, cm    163 [+ or -] 9     159 [+ or -] 5
SBP, mean [+ or -] SD, mmHg     115 [+ or -] 6     105 [+ or -] 10
DBP, mean [+ or -] SD, mmHg      74 [+ or -] 5      63 [+ or -] 4
Pulse, mean [+ or -] SD,         79 [+ or -] 10     73 [+ or -] 11
  beats/min

Characteristics                 Single-dose trial

                                Level 5

No. of volunteers               6
Dose, vials intravenously       12
Age, mean [+ or -] SD, years     23 [+ or -] 2.4
Weight, mean [+ or -] SD, kg     61 [+ or -] 9
Height, mean [+ or -] SD, cm    169 [+ or -] 8
SBP, mean [+ or -] SD, mmHg     116 [+ or -] 4
DBP, mean [+ or -] SD, mmHg      68 [+ or -] 3
Pulse, mean [+ or -] SD,         79 [+ or -] 11
  beats/min

Characteristics                 Multiple-dose trial

                                Level 4            Level 5

No. of volunteers               6                  6
Dose, vials intravenously       9                  12
Age, mean [+ or -] SD, years     23 [+ or -] 0.4    26 [+ or -] 3.4
Weight, mean [+ or -] SD, kg     58 [+ or -] 9      57 [+ or -] 5
Height, mean [+ or -] SD, cm    162 [+ or -] 9     166 [+ or -] 5
SBP, mean [+ or -] SD, mmHg     112 [+ or -] 11    103 [+ or -] 8
DBP, mean [+ or -] SD, mmHg      75 [+ or -] 11     68 [+ or -] 8
Pulse, mean [+ or -] SD,         83 [+ or -] 8      77 [+ or -] 7
  beats/min

Note: SBP: systolic blood pressure. DBP: diastolic blood pressure.

Table 3
Summary of the incidence of adverse drug reactions reported after
single dose of EH1 administration in healthy Chinese volunteers.

Adverse events/laboratory   Single-dose trial, no. of incidence,
tests                       gender

                            Level 4               Level 5

Dose, vials intravenously   9                     12
Dizziness                   0                     1 (grade 1), female
Headache                    0                     0
Nausea                      0                     0
Nasal obstruction           0                     0
Perspiration                0                     0
Increase in ALT             0                     0
Increase in urine protein   0                     0
Increase in urine glucose   0                     0
Change in ECG               1 (grade 1), female   1 (grade 1), male

Note: ALT: alanine aminotransferase, ECG: electrocardiogram.

Table 4
Summary of the incidence of adverse drug reactions reported after
multiple doses of EHI administration in healthy Chinese volunteers.

Adverse events/      Multiple-dose trial, no. of incidence, gender
laboratory tests
                     Level 4                      Level 5

Dose, vials          9                            12
  intravenously
Dizziness            3 (grade 1), 2 female/male   0
Headache             1 (grade 1), female          0
Nausea               1 (grade 1), female          0
Nasal obstruction    1 (grade 1). male            0
Perspiration         1 (grade 1), male            0
Increase in ALT      1 (grade 1), male            0
Increase in urine    1 (grade 1), male            0
  protein
Increase in urine    0                            1 (grade 1), female
  glucose
Change in ECG        1 (grade 1), male            1 (grade 1), female

Note: ALT: alanine aminotransferase, ECG: electrocardiogram.

Table 5
Pharmacokinetic parameters of scutellarin, 1,3-DCQA, 3,5-DCQA, and
3,4-DCQA after EH I administration in healthy volunteers.

Pharmacokinetic      Gender   Scutellarin           1,3-DCQA
parameters

[C.sub.max], ng/mL   Male      309 [+ or -] 41      25.4 [+ or -] 8.2
                     Female    219 [+ or -] 25 *    23.5 [+ or -] 2.7
[AUC.sub.0 [right    Male      232 [+ or -] 19        17 [+ or -] 4
  arrow] 1h], ng     Female    151 [+ or -] 14 **     17 [+ or -] 3
  h/mL
[AUC.sub.0 [right    Male      283 [+ or -] 24        41 [+ or -] 7
  arrow] 5h], ng     Female    183 [+ or -] 15 **     42 [+ or -] 5
  h/mL
[AUC.sub.0 [right    Male      286 [+ or -] 24        44 [+ or -] 8
  arrow]             Female    185 [+ or -] 14 **     46 [+ or -] 6
  [infinity]], ng
  h/mL
V,L                  Male      229 [+ or -] 32        56 [+ or -] 12
                     Female    404 [+ or -] 246       57 [+ or -] 12
CL, L/h              Male      131 [+ or -] 11        29 [+ or -] 6
                     Female    202 [+ or -] 15 **     28 [+ or -] 4
K, 1/h               Male     0.58 [+ or -] 0.07    0.53 [+ or -] 0.11
                     Female   0.62 [+ or -] 0.27    0.51 [+ or -] 0.11
ti/2. h              Male      1.2 [+ or -] 0.2      1.3 [+ or -] 0.3
                     Female    1.4 [+ or -] 0.8      1.4 [+ or -] 0.3

Pharmacokinetic      Gender   3,5-DCQA             3,4-DCQA
parameters

[C.sub.max], ng/mL   Male     65.9 [+ or -] 18      190 [+ or -] 62
                     Female   63.3 [+ or -] 21      195 [+ or -] 67
[AUC.sub.0 [right    Male       47 [+ or -] 10      135 [+ or -] 34
  arrow] 1h], ng     Female     44 [+ or -] 12      136 [+ or -] 38
  h/mL
[AUC.sub.0 [right    Male       61 [+ or -] 12      185 [+ or -] 43
  arrow] 5h], ng     Female     57 [+ or -] 14      180 [+ or -] 46
  h/mL
[AUC.sub.0 [right    Male       65 [+ or -] 14      195 [+ or -] 40
  arrow]             Female     70 [+ or -] 20      188 [+ or -] 48
  [infinity]], ng
  h/mL
V,L                  Male      993 [+ or -] 414     361 [+ or -] 147
                     Female   1672 [+ or -] 1481    279 [+ or -] 72
CL, L/h              Male      262 [+ or -] 64      112 [+ or -] 25
                     Female    252 [+ or -] 75      117 [+ or -] 27
K, 1/h               Male     0.31 [+ or -] 0.15   0.33 [+ or -] 0.06
                     Female   0.25 [+ or -] 0.18   0.42 [+ or -] 0.07
ti/2. h              Male      2.7 [+ or -] 1.3     2.2 [+ or -] 0.4
                     Female    5.6 [+ or -] 6.4     1.7 [+ or -] 0.2

Note: [C.sub.max]: peak plasma concentration, AUC: area under the
plasma concentration-time curve, V: volume of distribution, CL:
systemic clearance, K: elimination rate constant, [t.sub.1/2]:
terminal phase half-life, DCQA: dicaffeoylquinic acid, *: P < 0.05
between male and female, P < 0.001 between male and female.
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Article Details
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Author:Ju, Wen-Zheng; Zhao, Yang; Liu, Fang; Wu, Ting; Zhang, Jun; Liu, Shi-Jia; Zhou, Ling; Dai, Guo-Liang
Publication:Phytomedicine: International Journal of Phytotherapy & Phytopharmacology
Article Type:Report
Geographic Code:9CHIN
Date:Feb 15, 2015
Words:6257
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