molecules Article Optimal Extraction Study of Gastrodin-Type Components from Gastrodia Elata Tubers by Response Surface Design with Integrated Phytochemical and Bioactivity Evaluation Minhui Hu, Hui Yan, Yuanyuan Fu, Yulan Jiang, Weifeng Yao, Sheng Yu, Li Zhang, Qinan Wu, Anwei Ding and Mingqiu Shan * Jiangsu Collaborative Innovation Center of Chinese Medicinal Resources Industrialization, National and Local Collaborative Engineering Center of Chinese Medicinal Resources Industrialization and Formulae Innovative Medicine, Nanjing University of Chinese Medicine, Nanjing 210023, China; [email protected] (M.H.); [email protected] (H.Y.); [email protected] (Y.F.); [email protected] (Y.J.); [email protected] (W.Y.); [email protected] (S.Y.); [email protected] (L.Z.); [email protected] (Q.W.); [email protected] (A.D.) * Correspondence: [email protected]; Tel.: +86-025-85811519 Received: 9 January 2019; Accepted: 31 January 2019; Published: 2 February 2019 Abstract: Gastrodia elata tuber (GET) is a popular traditional Chinese medicines (TCMs). In this study, response surface methodology (RSM) with a Box–Behnken design (BBD) was performed to optimize the extraction parameters of gastrodin-type components (gastrodin, gastrodigenin, parishin A, parishin B, parishin C and parishin E). Different from the conventional studies that merely focused on the contents of phytochemical, we gave consideration to both quantitative analysis of the above six components by HPLC and representative bioactivities of GET, including antioxidation and protection of human umbilical vein endothelial cells (HUVEC). Four independent variables (ethanol concentration, liquid-material ratio, soaking time and extraction time) were investigated with the integrated evaluation index of phytochemical contents. With the validation experiments, the optimal extraction parameters were as follows: ethanol concentration of 41%, liquid–solid ratio of 28.58 mL/g, soaking time of 23.91 h and extraction time of 46.60 min. Under the optimum conditions, the actual standardized comprehensive score was 1.8134 ± 0.0110, which was in accordance with the predicted score of 1.8100. This firstly established method was proved to be feasible and reliable to optimize the extraction parameters of the bioactive components from GET. Furthermore, it provides some reference for the quality control and extraction optimization of TCMs. Keywords: Gastrodia elata tuber (GET); Response surface methodology; Antioxidation; HUVEC; Gastrodin-type components 1. Introduction Gastrodia elata Bl. is a perennial parasitic herb belonging to the Orchidaceae family. The plant is commonly found in the mountainous areas of Eastern Asia, including China, Korea and Japan [1]. Gastrodia elata tuber (GET), also called “Tianma” in China, was firstly recorded as a premium traditional Chinese medicine (TCM) in “Shen Nong Ben Cao Jing.” As a tonifying herbal medicine, GET has been used to extinguish wind to arrest convulsions, pacify and repress the liver yang, dispel wind and unblock the collaterals in clinic for thousands of years [2]. In modern pharmacological studies, GET has antioxidant, anti-angiogenic, neuroprotective, antidepressant, anxiolytic and sedative activities [3–9]. GET is also a quite favorite food that could improve body function and enhance immunity which Molecules 2019, 24, 547; doi:10.3390/molecules24030547 www.mdpi.com/journal/molecules Molecules 2019, 24, x 2 of 14 Molecules 2019, 24, 547 2 of 14 enhance immunity which frequently appears in the soup or porridge with chicken, pigeon, duck, fishfrequently head and appears so forth, in the in East soup Asia. or porridge Because with of its chicken, high edible pigeon, and duck, health fish value, head this and herbal so forth, medicine in East isAsia. listed Because as oneof ofits the high functional edibleand foods health approved value, by this the herbal Ministry medicine of Health is listed in China as one [10,11]. of the functional foodsFor approved gastrodin by (GD) the Ministry is one of of the Health predominant in China [bioactive10,11]. compounds in GET and gastrodigenin (GG, For4-hydroxybenzyl gastrodin (GD) isalcohol) one of thedue predominant to its aglycone, bioactive which compounds can exert in GET specific andgastrodigenin pharmacological (GG, activities4-hydroxybenzyl [12–15], alcohol) it is listed due toas its one aglycone, of the whichquality can evaluation exert specific indicators pharmacological of GET activitiesin the Chinese [12–15] , Pharmacopoeia.it is listed as one Therefore, of the quality most evaluation studies with indicators respec oft to GET the in processing the Chinese and Pharmacopoeia. extraction of GET Therefore, only focusedmost studies on these with two respect chemical to the processingevaluation and indexe extractions [16–19]. of GET However, only focused there onare thesesome two other chemical ester derivativesevaluation indexesof GD(s) [ 16in –GET19]. However,when combining there are with some a citric other acid, ester such derivatives as parishin of GD(s) A (PA), in GETparishin when B (PB),combining parishin with C (PC) a citric and acid, parishin such asE (PE). parishin The Acont (PA),ents parishin of these B partial (PB), parishincompounds C (PC) were and even parishin higher E than(PE). that The of contents GD [20,21]. of these They partial also compoundsexhibited some were promising even higher pharmacological than that of GD activities, [20,21]. including They also long-termexhibited somepotentiation promising protective, pharmacological antiaging and activities, antipsychotic including effects long-term [22–25], potentiation which were protective,similar to thatantiaging of GD, and GG antipsychotic and GET. The effects cases [22 –might25], which be explained were similar by tosome that researches of GD, GG that and GET.gastrodin The casesand gastrodigeninmight be explained are the by somemajor researchesmetabolites that of gastrodinthe abov ande parishin gastrodigenin derivatives are the [26,27]. major In metabolites this paper, of consideringthe above parishin their similar derivatives chemical [26, 27structures]. In this paper,(see Figure considering 1), these their six similarcomponents chemical are structuresgrouped into (see gastrodin-typeFigure1), these components, six components which are groupedwere reco intognized gastrodin-type as the material components, basis of GET. which were recognized as the material basis of GET. Figure 1. Chemical structures of the gastrodin-type components. Figure 1. Chemical structures of the gastrodin-type components. In addition, as we know, the curative effect is the straightforward index for quality evaluation of herbalIn medicine.addition, GETas we was know, reported the curative as one of effect the earliest is the andstraightforward most fundamental index medicinalfor quality foods evaluation to treat ofvascular herbal diseasesmedicine. including GET was hypertension reported as one and of atherosclerosis. the earliest and The most protection fundamental of this medicinal phytomedicine foods toon treat human vascular umbilical diseases vein endothelial including cellshypertension (HUVEC) and involved atherosclerosis. in these bioactivities The protection [28,29]. of On this the phytomedicineother hand, with on many human polyphenols umbilical includingvein endothelia the gastrodin-typel cells (HUVEC) components involved mentioned in these bioactivities above, GET [28,29].exerted On predominant the other antioxidanthand, withactivity, many polyph which alsoenols played including a crucial the rolegastrodin-type in its efficacies components and was mentionedrelated to many above, pharmacological GET exerted predominant bioactivities, antioxidant such as protection activity, onwhich liver also and played kidney a jury,crucial antiaging, role in itsreducing efficacies cerebral and was ischemia/reperfusion related to many pharmacological injury, antihypertension, bioactivities, neuroprotection such as protection [30 –on36 liver]. Hence, and kidneyHUVEC protectivejury, antiaging, and antioxidant reducing properties cerebral could ischemia/reperfusion also be utilized to reflectinjury, GET antihypertension, quality. neuroprotectionWith multiple [30–36]. quadratic Hence, regression HUVEC protective equation, responseand antioxidant surface properties methodology could (RSM) also be is utilized a novel tostatistical reflect GET approach quality. to solve multi-variable problems. RSM has been widely used to find out the optimumWith multiple parameters quadratic of different regression kinds equation, of process response in TCM, surface such methodology as extraction, (RSM) processing is a novel and statisticalpurification approach [37–42]. to However, solve multi-variable in almost all problems. the relative RSM studies, has been the contents widely ofused major to find compounds out the optimumhave been parameters paid much of attention, different neglecting kinds of process other critical in TCM, bioactivities. such as extraction, In this study, processing taking bothand purificationphytochemical [37–42]. yields However, and bioactivities in almost as all integrated the relative evaluation studies, indicators,the contents we of aim major to optimizecompounds the haveextraction been parameterspaid much of attention, the gastrodin-type neglecting components other critical from bioactivities. GET and to In provide this study, some referencetaking both for phytochemicalnatural products yields extraction and bioactivities from other TCMs.as