An Efficient Chemical Synthesis of Scutellarein: An in Vivo Metabolite of Scutellarin

Scutellarein (2), which is an important in vivo metabolite of scutellarin (1), was synthesized from 3,4,5-trimethoxyphenol (3) in high yield in four steps. This strategy relies on acetylation, aldolization, cyclization and hydrolysis reactions, respectively.


Introduction
As a frequently-occurring disease, ischemic cerebrovascular is a serious threat to human health and it has been one of the leading causes of death and disability around the world [1]. Traditional Chinese medicines are rich sources for drug lead compound discovery as they have been used clinically for thousands years. Scutellarin (1, Figure 1), a main active ingredient extracted from Erigeron breviscapus (Vant.) Hand-Mazz., which mainly grows in Yunnan Province of China, has been wildly used to treat acute cerebral infarction and paralysis induced by cerebrovascular diseases such as hypertension, cerebral thrombosis and cerebral hemorrhage in China since 1984 [2]. Interestingly, scutellarin (1) is mainly hydrolyzed into scutellarein (2, Figure 1) in the intestine [3], and scutellarein (2) was much more easily absorbed than scutellarin (1) after oral administration of both of them in equal doses [4]. In previous studies, our research group has found that scutellarein (2) had better protective effects than scutellarin (1) against neuronal injury in a rat cerebral ischemia model [5,6].

Introduction
As a frequently-occurring disease, ischemic cerebrovascular is a serious threat to human health and it has been one of the leading causes of death and disability around the world [1]. Traditional Chinese medicines are rich sources for drug lead compound discovery as they have been used clinically for thousands years. Scutellarin (1, Figure 1), a main active ingredient extracted from Erigeron breviscapus (Vant.) Hand-Mazz., which mainly grows in Yunnan Province of China, has been wildly used to treat acute cerebral infarction and paralysis induced by cerebrovascular diseases such as hypertension, cerebral thrombosis and cerebral hemorrhage in China since 1984 [2]. Interestingly, scutellarin (1) is mainly hydrolyzed into scutellarein (2, Figure 1) in the intestine [3], and scutellarein (2) was much more easily absorbed than scutellarin (1) after oral administration of both of them in equal doses [4]. In previous studies, our research group has found that scutellarein (2) had better protective effects than scutellarin (1) against neuronal injury in a rat cerebral ischemia model [5,6].  Unfortunately, scutellarein (2) is not readily available commercially, so the chemical synthesis of this metabolite has become important in recent years. We have previously synthesized scutellarein (2) from scutellarin (1) by hydrolysis with 6N HCl in 90% ethanol under reflux, however, the yield was very low (only 17%) [7]. Cui et al. [8] completed one synthetic route to scutellarein (2) from 2-hydroxy-4,5,6-trimethoxyacetophenone (4) and 4-methoxybenzoyl chloride in three steps, unfortunately, there was a by-product in this acetylation and the yield was low. Yen et al. [9] synthesized scutellarein (2) in 47% yield in five steps starting from 3,4,5-trimethoxyphenol (3) and acetic anhydride, then the obtained 3,4,5-trimethoxyphenol acetate was transformed into 1-(6-hydroxy-2,3,4-trimethoxyphenyl)ethanone (4) after Fries rearrangement.
In this paper, we report an efficient chemical synthesis of scutellarein (2) from 3,4,5-trimethoxyphenol (3) and acetic acid in only four steps and with high total yield (58%).
In this paper, we report an efficient chemical synthesis of scutellarein (2) from 3,4,5-trimethoxyphenol (3) and acetic acid in only four steps and with high total yield (58%).

Results and Discussion
As shown in Scheme 1, the starting material 3,4,5-trimethoxyphenol (3, 200 mg, 1.09 mmol) was first reacted with acetic acid (1 mL) in boron trifluoride diethyl etherate (5 mL) under a N2 atmosphere at 85 °C, to afford a high yield (92%) of the desired Friedel Crafts acylation reaction product 4. Next, compound 5 was synthesized by a base-catalyzed Claisen-Schmidt condensation reaction of 4 and 4-methoxybenzaldehyde. Fortunately, the cyclization of 5 produced the desired compound 6 in 82% yield, using iodine as the catalyst in dimethyl sulfoxide solution at 100 °C for 2 h. Finally, the demethylation of 6 with 40% HBr in the refluxing CH3COOH led to scutellarein (2) in 90% yield, for a total overall yield of 58%.

General Information
Reagents and solvents were purchased from commercial sources and used without further purification unless otherwise specified. Air-and moisture-sensitive liquids and solutions were transferred via syringe or stainless steel cannula. Organic solutions were concentrated by rotary evaporation (BuChi R-3, Surat, India) below 45 °C at approximately 20 mm Hg. All non-aqueous reactions were carried out under anhydrous conditions using flame-dried glassware in an argon atmosphere in dry, freshly distilled solvents, unless otherwise noted. Yields refer to chromatographically and spectroscopically ( 1 H-NMR) homogeneous materials, unless otherwise stated. Reactions were monitored by thin-layer chromatography (TLC) carried out on 0.15-0.20 mm silica gel plates (RSGF 254, Yantai, China) using UV light as the visualizing agent. The melting points (m.p.) were measured on a WRS-1B apparatus (Hangzhou, China) and are not corrected. 1

General Information
Reagents and solvents were purchased from commercial sources and used without further purification unless otherwise specified. Air-and moisture-sensitive liquids and solutions were transferred via syringe or stainless steel cannula. Organic solutions were concentrated by rotary evaporation (BuChi R-3, Surat, India) below 45˝C at approximately 20 mm Hg. All non-aqueous reactions were carried out under anhydrous conditions using flame-dried glassware in an argon atmosphere in dry, freshly distilled solvents, unless otherwise noted. Yields refer to chromatographically and spectroscopically ( 1 H-NMR) homogeneous materials, unless otherwise stated. Reactions were monitored by thin-layer chromatography (TLC) carried out on 0.15-0.20 mm silica gel plates (RSGF 254, Yantai, China) using UV light as the visualizing agent. The melting points (m.p.) were measured on a WRS-1B apparatus (Hangzhou, China) and are not corrected. 1 H-NMR (300 MHz) and 13 C-NMR spectra (75 MHz) were obtained with a Bruker AV-300 spectrometer (Karlsruhe, Germany). Chemical shifts are recorded in ppm downfield from tetramethylsilane. J values are given in Hz. Abbreviations used are s (singlet), d (doublet), t (triplet), q (quartet), b (broad) and m (multiplet).