Solid-state NMR studies and DFT calculations of flavonoids: baicalein, baicalin and wogonoside

Three flavonoids of pharmaceutical importance-baicalein, baicalin, and wogonoside-were isolated from a Chinese medicinal plant Scutellaria baicalensis Georgi and studied by 13C NMR in solution and solid state. Two-dimensional (2D) NMR spectroscopy in the liquid phase and dipolar dephasing (DD) experiments in magic-angle spinning (MAS) spectra enabled the assignment of 13C resonances. The cross-polarization (CP) time constants T(CH) and relaxation times T(H) (1rho) were obtained from the variable-contact time experiments. The principal elements of the 13C chemical shift tensor were determined in the spectra recorded under slow sample spinning (2 kHz) using phase-adjusted spinning sideband (PASS)-2D NMR technique, and were verified by density functional theory gauge-independent atomic orbital (DFT GIAO) calculations of shielding constants. Analysis of the 13C delta(ii) and comparison with shielding parameters calculated for different conformers of compounds 1-3 enabled the selection of the most reliable geometry in the solid phase. In all three compounds, an intramolecular hydrogen bond C5--OH...=C4 is formed; the existence of baicalein and baicalin with 'anticlockwise' orientation of OH groups is more probable.     

Screening of radical scavengers in Scutellaria baicalensis using HPLC with diode array and chemiluminescence detection

Scutellaria baicalensis Georgi is a well-known medical plant widely used as a famous traditional Chinese medicine. It has been reported that S. baicalensis can protect against oxidative stress and possess anti-inflammatory effect. In the present paper, an HPLC-diode array-chemiluminescence detection method for on-line detection was successfully developed to screen antioxidants in complex S. baicalensis extracts. Using the proposed approach, eight compounds in the S. baicalensis extracts were found to possess a potential antioxidant activity. Furthermore, the effects of purified compounds on protecting RAW 264.7 cells from hydrogen peroxide injury were also investigated in vitro to confirm the established method, which were consistent with the results of HPLC-diode array-chemiluminescence detection method. These results demonstrated that this method was a useful technique for rapidly screening and identifying bioactive components from complex herbal medicines.     

A New Flavanone and a New Flavanone Glycoside from Scutellaria baicalensis

A new flavanone (2S)-5, 7, 2',5'-tetrahydroxy-flavanone 1 together with a new flavanone glycoside (2S)-5, 7, 2',5'-tetrahydroxy-flavanone 7-O-β-D-glucopyranoside 2, were isolated from the dry roots of Scutellaria baicalensis. Their structures were elucidated on the basis of spectroscopic data.     

Synthesis of Ring A‐Modified Baicalein Derivatives

Baicalein, an important active constituent of the traditional Chinese herb Scutellaria baicalensis, exhibited antitumor activity and inhibitory activity against P‐gp 170. The syntheses of 25 baicalein derivatives, 2 – 26 (Table), are described here (Scheme 1). These compounds were systematically modified with O‐alkylation and O‐acylation at HO? C(5), HO? C(6), and HO? C(7), singly or in combination, on the ring A of baicalein in order to evaluate the effects of such modifications on their inhibitory activities against multidrug‐resistant tumor cell lines and P‐gp 170. Highly selective and efficient alkylations at HO? C(7) of peracetylated baicalein were the key to the distinction between HO? C(6) and HO? C(7) of baicalein.     

Simple preparation of baicalin from Scutellariae Radix

A simple, rapid and reproducible method was developed to isolate high-purity baicalin from Scutellariae Radix, the dried roots of Scutellaria baicalensis Georgi. The method involves partition/recrystallization steps without repeated column chromatography or special instruments. Isolated baicalin was characterized by comparisons of TLC, HPLC, IR, MS, and NMR data with an authentic sample. Moreover, beta-glucuronidase hydrolysis of baicalin sequentially yielded glucuronic acid and baicalein as confirmed by co-TLC with authentic samples. The purity of baicalin was more than 97% with yield ca. 8.7% (w/w). The method presented here appears suitable for commercial application.     

Investigation of flavonoid profile of Scutellaria bacalensis Georgi by high performance liquid chromatography with diode array detection and electrospray ion trap mass spectrometry

Scutellaria baicalensis Georgi is a well-known medicinal plant widely used in China and other East Asian countries. High performance liquid chromatography combined with diode array detection and electrospray ion trap mass spectrometry was used to determine the flavonoid profile of S. baicalensis. Under the optimized experiment conditions, 32 flavonoids were clearly detected. Eighteen main ones were doubtless identified by comparing their retention time, UV and MS (MSⁿ) data with isolated or commercial standards. The UV characteristics of these 18 known standards were studied in detail. The rules summarized provided valuable indications for the subsequent on-line identification processes. By interpreting both the MS and the UV data in detail, other 13 minor flavonoids in S. baicalensis were on-line identified successfully.     

Microfluidic Chip-Based Induced Phase Separation Extraction as a Fast and Efficient Miniaturized Sample Preparation Method

Induced phase separation extraction (IPSE) is an efficient sample clean-up technique that can replace liquid-liquid extraction (LLE). The purpose of this study was to miniaturize IPSE by carrying it out in a microfluidic chip. An IPSE chip was designed and evaluated for its ability to separate and purify samples on a microscale. The 5 × 2 cm chip was fed with a solution of polar to non-polar model compounds in acetonitrile-water (1:1). In the 100 µm wide and 40 µm deep microchannels, the sample solution was efficiently separated into two immiscible phases by adding a hydrophobic solvent as inducer. Analytes present in the sample solution each migrated to their own favorable phase upon phase separation. After optimization, extraction and fractionation were easily and efficiently achieved. The behavior of analytes with a pH-dependent partitioning could be influenced by adjusting the pH of the sample solution. Scutellaria baicalensis extract, used in Traditional Chinese Medicine (TCM), was successfully separated in aglycones and glycosides. In this microscale system, the sample and solvent consumption is reduced to microliters, while the time needed for the sample pretreatment is less than one minute. Additionally, the extraction efficiency can reach up to 98.8%, and emulsion formation is avoided.     

黄芩有效成分的紫外-可见吸收光谱表征

利用紫外-可见光谱的方法,对从中药黄芩中分离得到的3种黄芩有效成分的分子结构进行了研究。通过分析黄芩有效成分在甲醇溶液中及分别加入甲醇钠(NaOMe)、醋酸钠(NaOAc)、醋酸钠/硼酸(NaOAc/H₃BO₃)等5种诊断试剂后的紫外-可见光谱特征,讨论了利用紫外-可见吸收光谱推断黄酮类化合物母核结构及取代基位置的方法,为天然产物有效成分的结构鉴定提供了参考依据。     

Topical application of Scutellaria baicalensis suppresses 2,4-dinitrochlorobenzene-induced contact dermatitis

Allergic contact dermatitis (ACD) is a prototypic T-cell-mediated cutaneous inflammatory response. In the present study we describe the anti-allergic effect of topically applied Scutellaria bacalensis aqueous extract (WSBE) in suppressing 2,4-dinitrochlorobenzene (DNCB)-induced ACD in BALB/c mice. Topically applied WSBE attenuated the epidermal thickness and mast cell infiltration into the skin in DNCB-induced contact dermatitis. Furthermore, WSBE suppressed DNCB-induced production of serum IgE as well as IL-4, IFN-γ, and TNF-α in the skin. Topical application of WSBE also ameliorated the significant decrease in dermal glutathione and superoxide dismutase levels. Moreover, present results demonstrated that the baicalin, bioactive compound of WSBE, was able to penetrate into the skin following topical application, which was confirmed by the HPLC analysis using rat model. Taken together, topical application of WSBE exerts beneficial effects in contact dermatitis model, suggesting that WSBE might be a candidate for the treatment of contact dermatitis.     

Comparative chemical fingerprinting of Oroxylum indicum and Scutellaria baicalensis using liquid chromatography and mass spectrometry

Introduction: Identification of Oroxylum indicum and Scutellaria baicalensis provides an interesting challenge in selection of biomarker compound to be used in routine analysis. Both plants have similar phytochemical profile and are rich sources of flavones and flavone glycosides. The objective of this study was to prepare the chemical fingerprinting of O. indicum bark and S. baicalensis roots using the liquid chromatography and mass spectroscopy in single chromatographic method. Materials and methods: Extracts prepared using various solvent systems (methanol, aqueous methanol, chloroform, hexane, and water) of both plants were analyzed using C18 reverse phase column with solvent system containing acetonitrile and 0.1% formic acid. Major flavonoids were identified based on mass spectra, fragmentation pattern, and UV spectra. Results: In this article, well-resolved high-performance liquid chromatographic (HPLC) separation in both plant extracts was obtained and chemical fingerprints for both plant extracts were established and flavonoids present (baicalin, oroxylin A-7-O-glucuronide, chrysin-7-O-glucuronide, baicalein, chrysin, oroxylin-A, wogonin, skullcap flavone II) were identified as possible biomarkers. Conclusion: Mass spectrometry coupled with HPLC can be a tool for fingerprinting of various natural products used in dietary supplement industry. The fingerprint developed in the article can be used for quality evaluation as well as identifying possible adulteration of extracts of both the plants.