Method for purifying rebaudioside A by using simulated moving bed

A technology for simulating moving bed and mobile phase, which is applied in chemical instruments and methods, organic chemistry, preparation of sugar derivatives, etc. It can solve the problems of unfeasible process and high energy consumption, and achieve remarkable purification effect

CN104086610AActive Publication Date: 2014-10-08LIAONING QIANQIAN BIOLOGICAL TECH
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Patent Information

Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
Publication Date
2014-10-08

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Abstract

The invention discloses a method for purifying rebaudioside A by using a simulated moving bed. The method comprises the following steps: carrying out chromatograph on stevioside coarse extract by using the simulated moving bed, wherein the conditions are follows: the fixed phase is preparation-level filler purchased from German MERCKLiChroprepNH2 and the moving phase is 80.7% ethanol water; and separating to obtain the purified rebaudioside A. Under laboratory conditions, the following conditions shall be satisfied: eight tubular columns are stainless steel tubular columns of which the diameter is 10mm and the length is 15cm; the flow rate is as follows: QD=5.0mL / min, QF=0.1mL / min, QE=3.0mL / min and QR=2.1mL / min; the switching time is 8.0-8.25 minutes. The method for purifying rebaudioside A by using the simulated moving bed can improve the purity of rebaudioside A from 54% (the purity of rebaudioside A in the stevioside coarse extract) to 97% and the recovery rate can further is over 80%, and the purification effect is remarkable.
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Description

technical field

[0001] The invention relates to a method for purifying rebaudioside A by using a simulated moving bed. Background technique

[0002] Simulated Moving Bed (SMB) is a type of continuous chromatography. Since the stationary phase and the mobile phase are in continuous counterflow and continuous contact, the use efficiency of the stationary phase is improved, and the purpose of continuous feeding to increase production can also be achieved. . In operation, the position of the inlet and outlet of the SMB is continuously switched to simulate the phenomenon of reverse flow of the stationary phase and the mobile phase. figure 1 , figure 2 It is a schematic diagram of the SMB system. The system is composed of 8 packed columns connected in series. figure 1 There are 4 inlets and outlets in total, which are the inlets of feed and mobile phase (Desorbent), and the outlets of extract and raffinate. When the flow rate of the four inlets and outlets and the flow rate o...

Examples

Embodiment 1

[0025] Example 1 Quantitative analysis method of stevioside and rebaudioside A

[0026] Chemicals used in this embodiment:

[0027] Acetonitrile (AcN): HPLC analytical grade (ChromAR), MACRON CHEMICALS.

[0028] Stevioside: HPLC analytical grade (≧96%), Chengdu Master Biotechnology Co., Ltd.

[0029] Rebaudioside A: HPLC analytical grade (≧96%), SIGMA.

[0030] The present invention uses HPLC to carry out the quantitative analysis of rebaudioside A. The chromatographic column used is Supelcosil LC-NH2 (250 × 4.6 mm, 5 um) column. The HPLC system is composed of HITACHI L-2130 liquid pump, 20 microliter sampling loop, HITACHI L-2455 Diode Array Detector, and the UV wavelength used for area integration is 210 nm. The composition of the mobile phase is AcN / DIW=75 / 25 (V / V). In this study, the calibration curves of RA and SS were produced at the same time:

[0031] (3)

[0032] image 3 It is the chromatogram of stevia jaggery used in this experiment. image 3 The peak w...

Embodiment 2

[0033] Example 2 Purification of Rebaudioside A by Simulated Moving Bed

[0034] A commercially available from MERCK LiChroprep NH 2The preparative-grade filler, with a particle size of 40-60 microns, is filled in a stainless steel column with a diameter of 10 mm and a length of 15 cm by dry method. Using 80.7% ethanol aqueous solution (volume percentage) with a flow rate of 3.0 mL / min as the mobile phase, the chromatographic analysis of stevioside and rebaudioside A at 800 and 1000 mg / L was carried out respectively, such as Figure 4 . Figure 4 The chromatograms of stevioside and rebaudioside A are shown in .

[0035] Figure 4 The chromatogram shows that the signal peaks of the two components are quite symmetrical, so the adsorption behavior of stevioside and rebaudioside A between the selected filler and 80.7% ethanol aqueous solution is close to a linear adsorption behavior. According to chromatography theory, the Henry adsorption constant of a linearly adsorbed solut...

Embodiment 3

[0038] Example 3 Purification of Rebaudioside A by Simulated Moving Bed

[0039] The present invention takes figure 2 The SMB used to purify rebaudioside A in stevia raw sugar (purchased from Taiwan market) was the same as that in Example 2.

[0040] In order to illustrate the impact of the operating conditions of SMB on the content of purified rebaudioside A, the purity of rebaudioside A defined at the extraction end by the present invention is as follows:

[0041] (5)

[0042] Wherein A represents the area of ​​the chromatogram, the superscript E represents the sample at the extraction end, and the subscripts RA and imp represent rebaudioside A and all other solute components not containing rebaudioside A, respectively. The present invention only uses the area ratio to define the purity, but does not directly use the calibration curve of the formula (3) to define the purity. The calibration line of formula (3) shows that the response of stevioside (2778.2) is higher t...