Chemical Constituents from Turnip and Their Effects on α-Glucosidase

Brassica rapa var. rapa (turnip) is an important crop in Qinghai-Tibet Plateau (QTP) with anti-hypoxic effect. Turnip is rich in glucosinolates, isothiocyanates and phenolic compounds with diverse biological activities, involving anti-oxidant, anti-tumor, anti-diabetic, anti-inflammatory, anti-microbial, hypolipidemic, cardioprotective, hepatoprotective, nephroprotective and analgesic properties. In this study, the ethyl acetate (EtOAc) and butanol parts of Brassica rapa were first revealed with inhibitory effects on α-glucosidase, whereas the water part was inactive. Subsequent bioassay-guided isolation on the EtOAc and butanol parts yielded 12 compounds, involving three indole derivatives, indole-3acetonitrile (1) 4-methoxyindole-3-acetonitrile (2) and indole-3-aldehyde (3) two flavonoids, liquiritin (4) and licochalcone A (5) two phenylpropanoids, sinapic acid (6) and caffeic acid (7) two phenylethanol glycosides, 2-phenylethyl βglucopyranoside (8) and salidroside (9) and three other compounds, syringic acid (10) adenosine (11) and (3β, 20E)-ergosta-5, 20 (22)-dien-3-ol (12) Licochalcone A (5) and caffeic acid (7) showed α-glucosidase inhibitory activity with IC50 values of 62.4 ± 8.0 μM and 162.6 ± 3.2 μM, comparable to the positive control, acarbose (IC50 = 142 ± 0.02 μM). Docking study suggested that licochalcone A (5) could well align in the active site of α-glucosidase (docking score = -52.88) by forming hydrogen bonds (Gln1372, Asp1420, Gln1372, Arg1510), hydrophobic effects (Tyr1251, Tyr1251, Trp1355, Phe1560, Ile1587, Trp1355, Phe1559, Phe1559) and π-π stacking interaction (Trp1355). This study provides valuable information for turnip as a new resource in searching anti-diabetic candidates.


Introduction
Brassica rapa var. rapa (turnip) is an important crop with medicinal and edible purposes, which is widely consumed for protecting hypoxia in Qinghai-Tibet Plateau (QTP) [1]. Phytochemical investigation on turnip revealed that glucosinolates, isothiocyanates, flavonoids and volatiles were the main constituents [2]. Turnip showed a variety of bioactivities, involving anti-oxidant [3], anti-tumor [4], anti-diabetic [5], anti-inflammatory [6], anti-microbial [7], hypolipidemic, cardioprotective [8], hepatoprotective [9], nephroprotective [10] and analgesic effects [11]. Diabetes is a kind of metabolic disorder characterized by high levels of blood glucose, which affects about 425 million people all over the world [12]. According to the previous investigation, the ethanol extract of turnip showed anti-diabetic potency on type 2 diabetic mice, whereas the active constituents were still unclear [13]. In this study, the ethyl acetate (EtOAc) and butanol parts of turnip were first revealed with inhibitory effects on α-glucosidase, whereas the water part was inactive. As a continuous search for anti-diabetic candidates from natural resources, bioassay-guided fractionation on the EtOAc and butanol parts of turnip yielded 12 compounds. Herein, we reported their isolation and biological effects.

Plant Materials
The tubers of Brassica rapa var. rapa were collected from Huodeng Village, Lajing Township, Lanping County, Nujiang Prefecture, Yunnan Province in March 2016, which were taxonomically identified by Prof. Yong-Ping Yang. A voucher specimen (KTRG-B-43) was deposited in Kunming Botanical Garden, Kunming Institute of Botany, Chinese Academy of Sciences.

Bioassay in Vitro
Inhibitory activity against α-glucosidase was performed in accordance with our previous reports [14,15].

Docking Study
The crystal structure the C-terminal domain of human intestinal α-glucosidase (PDB ID: 3TOP) complexed with acarbose was retrieved from RCSB Protein Data Bank, and was used as molecular target. The docking calculation was performed using Yinfo Cloud Computing Platform (http://cloud.yinfotek.com), a friendly and versatile web server for bio-medicinal, material, and statistical researches. The active site of the enzyme was defined from the co-crystallized ligands from PDB files. The default docking protocols were applied for prediction the binding energies and the interactions between the ligands and protein.

Results
The total extraction of turnip was divided into EtOAc, butanol and water parts, and evaluated for their effects on α-glucosidase. The EtOAc extract showed significant activity (75.0 ± 15.4%), obviously higher than the butanol part (13.0 ± 2.5%). Whereas, the water part was inactive to α-glucosidase at the tested concentration (200 μg/mL). Therefore, the EtOAc and butanol parts were subjected to the following isolation.

Conclusion
In this study, both the EtOAc and butanol extracts of turnip were revealed with anti-diabetic potency in vitro, from which two compounds (5 and 7) with α-glucosidase inhibitory effects were obtained. This investigation provides valuable information for turnip as a new resource in searching anti-diabetic candidates.