Study On Green Extraction And Enrichment Of Main Saponins From Panax Notoginseng Roots

Panax notoginseng（Burk.）F.H.Chen,a well-known and valuable Chinese medicinal plant,is commonly the dried root of Panax notoginseng.Research shows that Panax notoginseng has significant pharmacological activities,such as hemostasis,invigorating blood,anti-tumor and anti-inflammatory.The major bioactive compounds of Panax notoginseng are widely recognized as sapoins,which possess extensive bioactivities and are also investigated as quality evaluation index of Panax notoginseng.In the present study,formulated surfactant,a green and novel extraction solvent,was combined with ultrasonic assisted extraction technology for the extraciton and enrichment of five main sapoin compounds(notoginsenoside R₁,ginsenosides Rg₁,Re,Rb₁ and Rd)from Panax notoginseng.In addition,the influential parameters of extraciton and enrichment processes were optimized.And then macroporous resin was used for separation and purification of five sapoins in Panax notoginseng.In this paper,we successfully synthetized a novel Gemini surfactant,and developed an efficient,rapid and reliable technical route for analysis,extraction and enrichment of notoginsenoside R₁,ginsenosides Rg₁,Re,Rb₁ and Rd in Panax notoginseng.Our study provided scientific guidance for reasonable development and efficient utilization of Panax notoginseng.The main findings were as follows:1.A HPLC method was established for rapid determination of notoginsenoside R₁,ginsenosides Rg₁,Re,Rb₁ and Rd in Panax notoginseng.The detail conditions were as follows:Chromatographic column:HIQ sil C18W column（250 mm×4.6 mm i.d.,5μm）;Flow rate:1.5 mL/min;Column temperature:30 ^oC;Injection volume:5μL;Detection wavelength:203 nm;Mobile phase:acetonitrile（A）and pure water（B）;Gradient elution:0-20 min,21%A;20-45 min,21-46%A.2.The Gemini surfactant of 16-5-16 was synthesiezd,and the optimal reaction conditions were as follows:Reaction solvent:ethanolReaction raion:C₁₆H₃₃N（CH₃）₂:Br（CH₂）₅Br=2.5:1Reaciton temperature:80 ^oCUnder the optimal synthesis conditions,the average yield of Gemini 16-5-16 was 94.6%.Besides,the product was indentified by IR,¹H-NMR and ¹³C-NMR.3.The influential parameters of formulated surfactant-ultrasonic assisted extraction processes were optimized.The optimal extraction and enrichment conditions were as follows:Surfactant type and concentration:2%Triton/0.03%Gemini 16-5-16Extraction pH:pH=8Liquid/solid ratio:30:1 mL/gUltrasonic power:250 WUltrasonic time:40 minElectrolyte salt concentration:20%（NH₄）₂SO₄Enrichment factor:11.5Under the optimal extraction conditions,the average extraction yields of notoginsenoside R₁,ginsenosides Rg₁,Re,Rb₁ and Rd were 28.66 mg/g,46.52 mg/g,12.91 mg/g,29.31 mg/g and 9.26 mg/g,respectively.Under the optimal enrichment conditions,the average recovery yields of notoginsenoside R₁,ginsenosides Rg₁,Re,Rb₁ and Rd were 86.65%,88.28%,84.94%,81.45%and 89.38%,respectively.And the concentration of five target compounds changed from 0.96 mg/mL,1.55 mg/mL,0.43 mg/mL,0.98 mg/mL and 0.31 mg/mL to 9.55 mg/mL,15.80 mg/mL,4.22 mg/mL,9.18 mg/mL and 3.24 mg/mL after enrichment by cloud-point phenomenon.Besides,the extraction efficiency by formulated surfactants was 1.13-1.52 fold better than that by conventional organic reagents.4.Macroporous resin was used for separation and purification of five sapoins in Panax notoginseng.The experimental results were as follows:Resin type:AB-8Sample concentration:notoginsenoside R₁ 1.11 mg/mL,ginsenosides Rg₁ 1.27 mg/mL,ginsenosides Re 0.44 mg/mL,ginsenosides Rb₁ 0.82 mg/mL and ginsenosides Rd 0.25 mg/mLProcessing volume:8/3 BVAdsorption flow rate:1 BV/hDesorption solution concentration for impurities:30%ethanolDesorption solution volume for impurities:3 BVDesorption solution concentration for target compounds:70%ethanolDesorption solution volume for target compounds:4.5 BVAfter the treatment by AB-8 under the optimal conditions,the contents of notoginsenoside R₁,ginsenosides Rg₁,Re,Rb₁ and Rd were 12.04%,16.62%,4.31%,12.35%and 3.61%,increased 4.57-fold,5.24-fold,4.89-fold,6.02-fold and 5.78-fold,respectively.In addition,the recoveries of notoginsenoside R₁,ginsenosides Rg₁,Re,Rb₁ and Rd were 68.53%、78.54%、73.45%、90.33%and 86.64%,and the product yield after enrichment is 3.9%,respectively.In this paper,the novel Gemini surfactant was synthesiezd,and firstly mixed with nonionic surfactant,then combined with ultrasonic assisted technology for extraction of notoginsenoside R₁,ginsenosides Rg₁,Re,Rb₁ and Rd in Panax notoginseng.Meanwhile,macroporous resin was used for enrichment of five target compounds.The procedure of formulated surfactant/ultrasonic assisted extraction followed by macroporous resin enrichment is a kind of environmental friendly,efficient and economic technique for extraction and enrichment of natural products in the Chinese herbal medicine.Compared with traditional methods,the proposed method uses green extraction solvent and simple extraction technique,which could lead to high extraction yields and recoveries.This study provides a scientific basis for the comprehensive development and utilization of Panax notoginseng.Moreover,it possesses important theoretical value and economic benefit for green and efficient extraction and enrichment of natural products.