CN101245087A - Neoflavone glycoside compound in sweetening chrysanthemum and separation identification method - Google Patents
Neoflavone glycoside compound in sweetening chrysanthemum and separation identification method Download PDFInfo
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Abstract
The invention discloses a new flavonoid glycoside in stevia rebaudiana bertoni and the separation and preparation method. The chemical structural formula of the new flavonoid glycoside compound quercetin-3-O-(4-O-reverse-coffee acyl-Alpha-L-rhamnose-(1 to 6)-Beta-D-galactosidase) in the stevia rebaudiana bertoni is shown as the figure.
Description
Technical field
The present invention relates to neoflavone glycoside compound and isolation and identification method thereof in a kind of sweet Stevia.
Background technology
Sweet Stevia (Stevia Rebaudiana Bertoni) is a composite family Stevia per nnial herb.Mainly contain stevia rebaudianum glycoside and flavones ingredient, wherein flavones ingredient has biological activitys such as good hypoglycemic, step-down, lipopenicillinase.By literature search, from sweet Stevia, separate the compound Quercetin-3-O-[4-O-obtain trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside] be a neoflavone glycosides compound.
Summary of the invention
The object of the present invention is to provide neoflavone glycosides compound in a kind of sweet Stevia; Another object of the present invention is to provide a kind of separation method of new compound; The 3rd purpose of the present invention is to provide a kind of All new compounds authentication method.
The present invention is achieved through the following technical solutions:
Neoflavone glycoside compound Quercetin-3-O-[4-O-in a kind of sweet Stevia is trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside], its chemical structural formula is as follows:
The compounds of this invention can also be prepared into compound and salt or the derivative that contains the said structure formula.Wherein the salt of this compound or derivative can be but be not limited to following several:
A. on this compound flavones parent nucleus-on the OH, coffee acyl-OH can methylate respectively or simultaneously, generates the derivative that methylates, the example structure formula is:
B. on the described derivative flavones of this compound and the A parent nucleus-on the OH, coffee acyl-OH respectively or simultaneously and Na
+, K
+In conjunction with generating the metal-salt derivative, example derivant structure formula is Deng metal ion:
C. adjacent two phenolic hydroxyl groups, the 5-hydroxyl of flavones parent nucleus on this compound and A, the described derivative of B, the 4-carbonyl can be distinguished or simultaneously and metal ion Al
3+, Sr
2+, Mg
2+, Zr
2+Generate metal complex Deng combination, example complex structure formula is:
D. the trans alkene hydrogen on this compound and A, the described derivative coffee acyl of B, C can be converted into the cis derivative, and example derivant structure formula is:
E. addition reaction can take place and generates a series of derivatives in the alkene hydrogen on this compound and A, B, C, the described derivative coffee acyl of D, and example derivant structure formula is:
F. intramolecularly polymerization reaction can take place in this compound and A, B, C, the described derivative of D, E, and example derivant structure formula is:
H. molecule inner dewatering reaction can take place in this compound and A, B, C, the described derivative of D, E, and example derivant structure formula is:
The separation method of new compound of the present invention is:
Select commercially available sweet Stevia (Stevia Rebaudiana Bertoni) for use; dry blade for composite family Stevia per nnial herb; water; ethanol; methyl alcohol; acetone; ethyl acetate; propyl carbinol or their at least two kinds of mixtures are solvent extraction; reclaim behind the solvent extract by being selected from solvent extration; macroreticular resin absorbing method; polymeric amide chromatography method; silica gel column chromatography; reversed phase column chromatography method or Toyopearl HW-40; Pharmadex LH-20 column chromatography; one or more modes of recrystallization method are separated; water; ethanol; methyl alcohol; acetone; acetonitrile; chloroform; sherwood oil; ethyl acetate; methylene dichloride; hexanaphthenes etc. or their at least two kinds of mixtures are as the eluent wash-out; adopt the thin layer chromatography inspection to know; merge elutriant, after the drying neoflavone glycoside compound Quercetin-3-O-[4-O-trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside].
According to the present invention, when raw material is extracted, can be under room temperature or heated condition, selecting water, ethanol, methyl alcohol, acetone, ethyl acetate, propyl carbinol or their at least two kinds of mixtures for use is solvent, adopt decoction, reflux, supersound extraction, cold soaking, diacolation, microwave extraction, high pressure extract, extraction time can be once also can be repeatedly.Wherein, preferred: 10～95% aqueous ethanolic solution heating and refluxing extraction 1～3 time, extract 0.5～2h at every turn.
According to the present invention, when adopting macroreticular resin absorbing method to separate, resin can be selected for use and be D101 type macroporous adsorbent resin, D3526 type macroporous adsorbent resin, HPD400 type macroporous adsorbent resin, AB-8 type macroporous adsorbent resin, S-8 type macroporous adsorbent resin, X-5 type macroporous adsorbent resin and other model polystyrene macroporous resin, in eluting solvent water, ethanol, methyl alcohol, the acetone one or more, isocratic elution can be adopted during wash-out, also gradient elution can be adopted.Wherein macroreticular resin absorbing method is preferably: with extracting solution or extract water dispersing and dissolving, supernatant liquor or filtered liquid pass through macroporous adsorbent resin, the aqueous solution of water or 5～20% acetone, ethanol, methyl alcohol or propyl alcohol cleans removes impurity, consumption is 1～7 times of resin volume, use the aqueous solution wash-out of 20～90% acetone, ethanol, methyl alcohol or propyl alcohol again, consumption is 2～8 times of resin volumes, collects elutriant, reclaim solvent, promptly get the position of containing sweet Stevia neoflavone glycosides.
In the used polymeric amide chromatography method of the present invention, can select column chromatography or Static Adsorption filtration method for use, eluent be selected from water, methyl alcohol, ethanol, acetone, chloroform, methyl alcohol, ethyl acetate etc. one or more, can adopt isocratic elution during wash-out, also can adopt gradient elution.Be preferably: will be adsorbed with sample (contain Quercetin-3-O-[4-O-trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside]) polymeric amide be splined on polymeric amide chromatography post; collect step by step with ethyl acetate, ethyl acetate-methyl alcohol, methyl alcohol gradient elution and chloroform, chloroform-methanol, methyl alcohol gradient elution; according to inspection knowledge situation; combined segment stream part; reclaim solvent, get neoflavone glycoside compound in the sweet Stevia.
In the used silica gel column chromatography of the present invention, can select normal pressure or pressurized column chromatography for use, used filler is 40～400 order silica gel, with sherwood oil, chloroform, acetone, ethyl acetate, methyl alcohol, ethanol, propyl alcohol, butanone, methylene dichloride, water etc. or their at least two kinds of admixture solvents, isocratic elution can be adopted during wash-out, also gradient elution can be adopted.
In the used reversed phase column chromatography of the present invention, can select normal pressure or pressurized column chromatography for use, used filler is octadecyl bonding phase or eight alkyl linked phases, eluting solvent be aqueous methanol, aqueous ethanol, aqueous acetone, moisture propyl alcohol one or more, isocratic elution can be adopted during wash-out, also gradient elution can be adopted.
In the column chromatography of the used Toyopearl HW-40 of the present invention, can select normal pressure or pressurized column chromatography for use, eluting solvent be selected from aqueous methanol, aqueous ethanol, aqueous acetone, moisture propyl alcohol one or more, organic phase concentration is 0～100%, substep is collected, according to detection case, and combined segment stream part, isocratic elution can be adopted during wash-out, also gradient elution can be adopted.
In the used Pharmadex LH-20 column chromatography of the present invention, can select normal pressure or pressurized column chromatography for use, eluting solvent be selected from aqueous methanol, aqueous ethanol, aqueous acetone, moisture propyl alcohol one or more, organic phase concentration is 5～100%, substep is collected, according to detection case, and combined segment stream part, isocratic elution can be adopted during wash-out, also gradient elution can be adopted.
In the used recrystallization method of the present invention, that recrystallization solvent is selected from is moisture, a kind of, several or two or more admixture solvent in the methyl alcohol, ethanol, acetone, chloroform, the repeated multiple times recrystallization.
New compound Quercetin-3-O-[4-O-of the present invention is trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside] the structure authentication method be:
Yellow amorphous powder, mp:240 ℃ (decomposition), hydrochloric acid-magnesium powder reaction is positive UV λ maxMeOH:255.4,351.8; Show that its structure parent nucleus is a flavones.It is C that high resolution FAB-MS provides its molecular formula
36H
36O
19(measured value: 795.177399; Calculated value: 795.174849).
By
1H-NMR as can be known, exist on one group of phenyl ring in this compound structure between position coupling hydrogen signal, promptly δ 6.33 (1H, d, J=2.0Hz), δ 6.18 (1H, d, J=2.0Hz).Also contain ABX coupled system on two groups of phenyl ring, promptly δ 7.52 (1H, d, J=2.0Hz), δ 7.66 (1H, dd, J=2.0Hz, 8.0Hz), δ 6.81 (1H, d, J=8.0Hz), δ 7.03 (1H, d, J=2.0Hz), δ 6.98 (1H, dd, J=2.0Hz, 8.0Hz), δ 6.74 (1H, d J=8.0Hz), shows three replacements that contain in the structure on two phenyl ring.(1H, signal s) are the signal that 5 OH of A ring go up hydrogen, illustrate that last 6,8 of the A ring of flavones parent nucleus has position coupled hydrogen between two, on the B ring ABX system are arranged at δ 12.57.
By
1H-
1HCOSY can distinguish 6 hydrogen signals of two groups of ABX coupled systems, is respectively: δ 7.52 (1H, d, J=2.0Hz), δ 7.66 (1H, dd, J=2.0Hz, 8.0Hz), δ 6.81 (1H, d is J=8.0Hz) with δ 7.03 (1H, d, J=2.0Hz), δ 6.98 (1H, dd, J=2.0Hz, 8.0Hz), δ 6.74 (1H, d, J=8.0Hz).
1H-NMR and
1H-
1(J=16.0Hz), (1H, d J=16.0Hz) are two trans coupling hydrogen on the ethylene linkage to δ 6.20 to δ 7.44 among the HCOSY for 1H, d; HMQC composes demonstration, and δ 7.44 hydrogen (hydrogen on the ethylene linkage) are connected with δ 145.88 carbon; The HMBC spectrum shows δ 145.88 carbon and δ 7.03 hydrogen and δ 6.98 hydrogen, and δ 7.44 hydrogen are relevant with the carbonyl carbon of δ 166.99, contain coffee acyl in the above description architecture.
Table 1 new compound, methyl-β-D-galactoside and methyl-alpha-L-rhamnoside
13The C-NMR data
(
13C-NMR?data?for?New?Compound、Methyl-β-D-galacoside?andMethyl-α-L-rhamoside)
1δ 4.50 among the H-NMR (1H, br s), (1H, d are that sugar is gone up terminal hydrogen J=7.6Hz) to δ 5.31.By
1(3H, d J=2.0Hz) reach δ 0.84 among the H-NMR
13Among the C-NMR signal of δ 17.494 as can be known one of them sugar be rhamnosyl, and δ 4.50 (1H s) is its terminal hydrogen.The signal of sugar in the carbon of this compound spectrum relatively (is shown 2-2) with 1 methide of standard sugar, and as can be known, 4 of rhamnosyl are substituted, and contain beta galactose in the structure, and its 6 are substituted.In the HMBC spectrum, 1 hydrogen δ 4.50 of rhamnosyl and 6 carbon δ 66.6 reference points of semi-lactosi have α-L-rhamnosyl-(1 → 6)-β-D-semi-lactosi segment in the description architecture.
In the HMBC spectrum, 1 hydrogen δ 5.31 of semi-lactosi and 3 carbon δ 134.12 of flavones illustrate that 3 carbon of semi-lactosi C-1 and flavones link.
The carbonyl carbon δ 166.99 of coffee acyl has reference point with 4 carbon of rhamnosyl, and coffee acyl links to each other with the C-4 of rhamnosyl.
To sum up, the structure of this compound is Quercetin-3-O-[4-O-trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside] (Quercetin-3-O-(4-O-trans-caffeoyl-α-L-rhamnopyranosyl-(1 → 6)-β-D-galacopyranoside)).
Following experimental example and embodiment are used to further specify but are not limited to the present invention.
By 2DNMR (HMBC, HMQC, DEPT,
1H-
1HCOSY, TOCSY) determined each hydrocarbon ownership on this compound.The results are shown in Table 2.
The NMR data of table 2 new compound (400MHz, in DMSO)
(Table1-2?NMR?data?of?New?Compound(400MHz，in?DMSO))
A is because shielding effect poor TV signal clear (Signals pattern are unclear due to overlapping.)
Following embodiment all can realize the effect of above-mentioned experimental example
Embodiment
Embodiment 1:
Compound Quercetin-3-O-[4-O-is trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside], its structural formula is as follows:
Its separation method is: select the dry leave 7000g of composite family Stevia per nnial herb sweet Stevia (SteviaRebaudiana Bertoni) for use, with 50～95% extraction using alcohols of 14 times of parts by volume 3 times, each 1.0 hours, filter.Merging filtrate and be evaporated to 10L after, be diluted with water to 56L, by AB-8 type macroporous adsorptive resins, use 30～70% ethanol elutions again behind 3 column volumes of water wash-out, be washed till ethanol eluate near colourless after, merge above-mentioned ethanol eluate, 50～70 ℃ of dryings in the rearmounted vacuum drying oven of concentrating under reduced pressure obtain the heavy 917g of dry extract.
Above-mentioned dry extract 917g is separated (chromatographic separation I) with polyamide column chromatography, water-ethanol mixing system gradient elution, portioning is collected, and every 100ml collects a, collect altogether that I washing part, II 30% alcohol are washed, III50% alcohol is washed part, IV70% alcohol is washed part of V 90% alcohol and is washed five streams of part part.The IV70% alcohol that chromatographic separation I obtains is washed part through polymeric amide chromatographic separation (chromatographic separation II), with ethyl acetate-10: 1 → 6: 1 → 4: 1 → 2: 1 → 1: 1 → 1: 3 → 1: 6 gradient elution of methanol mixed solvent, portioning is collected, every 50ml collects a, collect 50～60 stream parts altogether, back merging same stream part is known in the thin inspection of polymeric amide.39～41 streams part merging that chromatographic separation II obtains, separate (chromatographic separation III) through polyamide column chromatography, with 6: 1 → 4: 1 → 2: 1 → 1: 1 → 1: 2 → 1: 4 gradient elution of chloroform-methanol mixed solvent, portioning is collected, every 50ml collects a, collect 15～20 stream parts altogether, back merging same stream part is known in the thin inspection of polymeric amide.10～14 streams that chromatographic separation III obtains part merge, behind acetone recrystallization, obtain Quercetin-3-O-[4-O-trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside] 0.020g.
Embodiment 2:
Compound Quercetin-3-O-[4-O-is trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside], its structural formula is as follows:
Its separation method is: select the dry leave 5000g of composite family Stevia per nnial herb sweet Stevia (SteviaRebaudiana Bertoni) for use, with 50～95% extraction using alcohols of 14 times of parts by volume 2 times, each 2.0 hours, filter.Merging filtrate and be evaporated to 10L after, be diluted with water to 50L, by AB-8 type macroporous adsorptive resins, use 30～70% ethanol elutions again behind 5 column volumes of water wash-out, be washed till ethanol eluate near colourless after, merge above-mentioned ethanol eluate, 50～70 ℃ of dryings in the rearmounted vacuum drying oven of concentrating under reduced pressure obtain the heavy 615g of dry extract.
Above-mentioned dry extract 615g is separated (chromatographic separation I) with polyamide column chromatography, water-ethanol mixing system gradient elution, portioning is collected, and every 100ml collects a, collect altogether that I washing part, II30% alcohol are washed, III50% alcohol is washed part, IV70% alcohol is washed part of V 90% alcohol and is washed five streams of part part.The IV70% alcohol that chromatographic separation I obtains is washed part through polymeric amide chromatographic separation (chromatographic separation II), with ethyl acetate-10: 1 → 6: 1 → 4: 1 → 2: 1 → 1: 1 → 1: 3 → 1: 6 gradient elution of methanol mixed solvent, portioning is collected, every 50ml collects a, collect 50～60 stream parts altogether, back merging same stream part is known in the thin inspection of polymeric amide.39～41 streams part merging that chromatographic separation II obtains, separate (chromatographic separation III) through polyamide column chromatography, with 6: 1 → 4: 1 → 2: 1 → 1: 1 → 1: 2 → 1: 4 gradient elution of chloroform-methanol mixed solvent, portioning is collected, every 50ml collects a, collect 15～20 stream parts altogether, back merging same stream part is known in the thin inspection of polymeric amide.10～14 streams that chromatographic separation III obtains part merge, behind acetone recrystallization, obtain Quercetin-3-O-[4-O-trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside] 0.015g.
Embodiment 3:
Compound Quercetin-3-O-[4-O-is trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside], its structural formula is as follows:
Its separation method is: select the dry leave 10000g of composite family Stevia per nnial herb sweet Stevia (SteviaRebaudiana Bertoni) for use, with 50～95% extraction using alcohols of 16 times of parts by volume 3 times, each 2.0 hours, filter.Merging filtrate and be evaporated to 10L after, be diluted with water to 80L, by AB-8 type macroporous adsorptive resins, use 30～70% ethanol elutions again behind 4 column volumes of water wash-out, be washed till ethanol eluate near colourless after, merge above-mentioned ethanol eluate, 50～70 ℃ of dryings in the rearmounted vacuum drying oven of concentrating under reduced pressure obtain dry extract 1310g.
Above-mentioned dry extract 1310g is separated (chromatographic separation I) with polyamide column chromatography, water-ethanol mixing system gradient elution, portioning is collected, and every 100ml collects a, collect altogether that I washing part, II30% alcohol are washed, III50% alcohol is washed part, IV70% alcohol is washed part of V 90% alcohol and is washed five streams of part part.The IV70% alcohol that chromatographic separation I obtains is washed part through polymeric amide chromatographic separation (chromatographic separation II), with ethyl acetate-10: 1 → 6: 1 → 4: 1 → 2: 1 → 1: 1 → 1: 3 → 1: 6 gradient elution of methanol mixed solvent, portioning is collected, every 50ml collects a, collect 50～60 stream parts altogether, back merging same stream part is known in the thin inspection of polymeric amide.39～41 streams part merging that chromatographic separation II obtains, separate (chromatographic separation III) through polyamide column chromatography, with 6: 1 → 4: 1 → 2: 1 → 1: 1 → 1: 2 → 1: 4 gradient elution of chloroform-methanol mixed solvent, portioning is collected, every 50ml collects a, collect 15～20 stream parts altogether, back merging same stream part is known in the thin inspection of polymeric amide.10～14 streams that chromatographic separation III obtains part merge, behind acetone recrystallization, obtain Quercetin-3-O-[4-O-trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside] 0.025g.
Embodiment 4:
Compound Quercetin-3-O-[4-O-is trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside], its structural formula is as follows:
Its separation method is: select the dry leave 3000g of composite family Stevia per nnial herb sweet Stevia (SteviaRebaudiana Bertoni) for use, with 50～95% extraction using alcohols of 12 times of parts by volume 3 times, each 1.5 hours, filter.Merging filtrate and be evaporated to 5L after, be diluted with water to 20L, by AB-8 type macroporous adsorptive resins, use 30～70% ethanol elutions again behind 3 column volumes of water wash-out, be washed till ethanol eluate near colourless after, merge above-mentioned ethanol eluate, 50～70 ℃ of dryings in the rearmounted vacuum drying oven of concentrating under reduced pressure obtain dry extract 400g.
Above-mentioned dry extract 400g is separated (chromatographic separation I) with polyamide column chromatography, water-ethanol mixing system gradient elution, portioning is collected, and every 100ml collects a, collect altogether that I washing part, II30% alcohol are washed, III50% alcohol is washed part, IV70% alcohol is washed part of V 90% alcohol and is washed five streams of part part.The IV70% alcohol that chromatographic separation I obtains is washed part through polymeric amide chromatographic separation (chromatographic separation II), with ethyl acetate-10: 1 → 6: 1 → 4: 1 → 2: 1 → 1: 1 → 1: 3 → 1: 6 gradient elution of methanol mixed solvent, portioning is collected, every 50ml collects a, collect 50～60 stream parts altogether, back merging same stream part is known in the thin inspection of polymeric amide.39～41 streams part merging that chromatographic separation II obtains, separate (chromatographic separation III) through polyamide column chromatography, with 6: 1 → 4: 1 → 2: 1 → 1: 1 → 1: 2 → 1: 4 gradient elution of chloroform-methanol mixed solvent, portioning is collected, every 50ml collects a, collect 15～20 stream parts altogether, back merging same stream part is known in the thin inspection of polymeric amide.10～14 streams that chromatographic separation III obtains part merge, behind acetone recrystallization, obtain Quercetin-3-O-[4-O-trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside] 0.010g.
Embodiment 5:
Compound Quercetin-3-O-[4-O-is trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside], its structural formula is as follows:
Its separation method is: the windproof medicine materical crude slice 4000g that selects for use the dry root of umbelliferae Saposhnikovia divaricata to make, with 50～95% extraction using alcohols of 10 times of parts by volume 3 times, each 0.5 hour, filter.Merging filtrate and be evaporated to 5L after, be diluted with water to 30L, by AB-8 type macroporous adsorptive resins, use 30～70% ethanol elutions again behind 4 column volumes of water wash-out, be washed till ethanol eluate near colourless after, merge above-mentioned ethanol eluate, 50～70 ℃ of dryings in the rearmounted vacuum drying oven of concentrating under reduced pressure obtain dry extract 500g.
Above-mentioned dry extract 400g is separated (chromatographic separation I) with polyamide column chromatography, water-ethanol mixing system gradient elution, portioning is collected, and every 100ml collects a, collect altogether that I washing part, II30% alcohol are washed, III50% alcohol is washed part, IV70% alcohol is washed part of V 90% alcohol and is washed five streams of part part.The IV70% alcohol that chromatographic separation I obtains is washed part through polymeric amide chromatographic separation (chromatographic separation II), with ethyl acetate-10: 1 → 6: 1 → 4: 1 → 2: 1 → 1: 1 → 1: 3 → 1: 6 gradient elution of methanol mixed solvent, portioning is collected, every 50ml collects a, collect 50～60 stream parts altogether, back merging same stream part is known in the thin inspection of polymeric amide.39～41 streams part merging that chromatographic separation II obtains, separate (chromatographic separation III) through polyamide column chromatography, with 6: 1 → 4: 1 → 2: 1 → 1: 1 → 1: 2 → 1: 4 gradient elution of chloroform-methanol mixed solvent, portioning is collected, every 50ml collects a, collect 15～20 stream parts altogether, back merging same stream part is known in the thin inspection of polymeric amide.10～14 streams that chromatographic separation III obtains part merge, behind acetone recrystallization, obtain Quercetin-3-O-[4-O-trans-coffee acyl-α-L-rhamnosyl-(1 → 6)-β-D-galactoside] 0.012g.
Embodiment 6:
Trans alkene hydrogen on this compound coffee acyl can be converted into formally, and example derivant structure formula is:
Embodiment 7:
On this compound flavones parent nucleus-on the OH, coffee acyl-OH can methylate respectively or simultaneously, generates the derivative that methylates, and the example structure formula is:
Embodiment 8:
On this compound flavones parent nucleus-on the OH, coffee acyl-OH is respectively or simultaneously and Na
+, K
+In conjunction with generating the metal-salt derivative, example derivant structure formula is Deng metal ion:
Embodiment 9:
This compound adjacent two phenolic hydroxyl groups, the 5-hydroxyl of flavones parent nucleus, the 4-carbonyl can be distinguished or simultaneously and metal ion Al
3+, Sr
2+, Mg
2+, Zr
2+Generate metal complex Deng combination, example complex structure formula is:
Description of drawings:
Fig. 1 FAB-MS high resolution mass spectrum data
Fig. 2 new compound
1The H-NMR spectrum
Fig. 3 new compound
1H-NMR composes (partial enlarged drawing)
Fig. 4 new compound
1H-
1HCOSY composes (partial enlarged drawing)
The HMQC spectrum (partial enlarged drawing) of Fig. 5 new compound
The HMBC spectrum (partial enlarged drawing) of Fig. 6 new compound
The HMBC spectrum (partial enlarged drawing) of Fig. 7 new compound
The HMBC spectrum (partial enlarged drawing) of Fig. 8 new compound
The HMBC spectrum (partial enlarged drawing) of Fig. 9 new compound
HMBC spectrum (from hydrogen to carbon) (The HMBC correlations observed in new compound (from H to C)) of Figure 10 new compound.
Claims (14)
1. the neoflavone glycosides compound in the sweet Stevia is characterized in that the chemical structural formula of this compound is:
2. medicine is characterized in that this medicine contains compound and the salt or the derivative of following structural formula:
3. the neoflavone glycoside compound separation method in the sweet Stevia as claimed in claim 1 is characterized in that this method is:
Select for use commercially available sweet Stevia (Stevia Rebaudiana Bertoni) to be the dry blade of composite family Stevia per nnial herb sweet Stevia (Stevia Rebaudiana Bertoni), water, ethanol, methyl alcohol, acetone, ethyl acetate, propyl carbinol or their at least two kinds of mixtures are solvent extraction, reclaim behind the solvent extract by being selected from solvent extration, macroreticular resin absorbing method, polymeric amide chromatography method, silica gel column chromatography, reversed phase column chromatography method or Toyopearl HW-40, Pharmadex LH-20, one or more modes of column chromatography are separated, water, ethanol, methyl alcohol, acetone, ethyl acetate, chloroform, methylene dichloride, sherwood oil, hexanaphthenes etc. or their at least two kinds of mixtures are made the eluent wash-out, adopt the thin layer chromatography inspection to know, the combined segment elutriant gets sweet Stevia neoflavone glycoside compound after the drying.
4. as the separation method of sweet Stevia neoflavone glycoside compound as described in the claim 3, it is characterized in that raw material is extracted is under room temperature or heated condition, adopt decoction, reflux, supersound extraction, cold soaking, diacolation, microwave extraction, high pressure extract, extraction time can be once also can be repeatedly.
5. as the separation method of sweet Stevia neoflavone glycoside compound as described in the claim 3, it is characterized in that used solvent extration is to be selected from methyl alcohol, acetone, ethyl acetate, propyl carbinol or their at least two kinds of mixtures as solvent, under room temperature or heated condition, extract, extraction times can be once also can be repeatedly.
6. sweet Stevia neoflavone glycoside compound separation method as claimed in claim 3, it is characterized in that resin used in the used macroreticular resin absorbing method is the polystyrene type polymeric adsorbent, eluting solvent is selected from water, methyl alcohol, ethanol, acetone, propyl alcohol or their at least two kinds of mixtures as eluent, elution process can be an isocratic elution, also can be gradient elution.
7. macroreticular resin absorbing method as claimed in claim 6 is characterized in that used resin is AB-8 type macroporous adsorbent resin, X-5 type macroporous adsorbent resin, D3526 type macroporous adsorbent resin, HPD400 type macroporous adsorbent resin, S-8 type macroporous adsorbent resin, NKA-II type macroporous adsorbent resin.
8. macroreticular resin absorbing method as claimed in claim 6, it is characterized in that described macroreticular resin absorbing method is: with extracting solution or extract water dispersing and dissolving, supernatant liquor or filtered liquid pass through macroporous adsorbent resin, the aqueous solution of water or 5～20% acetone, ethanol, methyl alcohol or propyl alcohol cleans removes impurity, consumption is 1～7 times of resin volume, use the aqueous solution wash-out of 20～90% acetone, ethanol, methyl alcohol or propyl alcohol again, consumption is 2～8 times of resin volumes.
9. sweet Stevia neoflavone glycoside compound separation method as claimed in claim 3, it is characterized in that described polymeric amide chromatography method is column chromatography or Static Adsorption filtration method, eluent is selected from the solvent that water, methyl alcohol, ethanol, acetone, chloroform, methyl alcohol, ethyl acetate etc. are formed, isocratic elution can be adopted during wash-out, also gradient elution can be adopted.
10. sweet Stevia neoflavone glycoside compound separation method as claimed in claim 3, it is characterized in that described silica gel column chromatography is normal pressure or pressurized column chromatography, used filler is 40～400 order silica gel, with sherwood oil, chloroform, methylene dichloride, acetone, ethyl acetate, methyl alcohol, ethanol, propyl alcohol, butanone, water etc. or their at least two kinds of mixtures is eluent, isocratic elution can be adopted during wash-out, also gradient elution can be adopted.
11. sweet Stevia neoflavone glycoside compound separation method as claimed in claim 3, it is characterized in that described reversed phase column chromatography is normal pressure or pressurized column chromatography, used filler is octadecyl bonding phase or eight alkyl linked phases, eluting solvent is selected from one or more in aqueous methanol, aqueous ethanol, aqueous acetone, the moisture propyl alcohol, isocratic elution can be adopted during wash-out, also gradient elution can be adopted.
12. separation method as sweet Stevia neoflavone glycoside compound as described in the claim 3, it is characterized in that described Toyopearl HW-40 column chromatography is normal pressure or pressurized column chromatography, eluting solvent is selected from the eluent of being made up of aqueous methanol, aqueous ethanol, aqueous acetone, moisture propyl alcohol, isocratic elution can be adopted during wash-out, also gradient elution can be adopted.
13. separation method as sweet Stevia neoflavone glycoside compound as described in the claim 3, it is characterized in that described Pharmadex LH-20 column chromatography is normal pressure or pressurized column chromatography, eluting solvent is selected from the eluent of being made up of aqueous methanol, aqueous ethanol, aqueous acetone, moisture propyl alcohol, isocratic elution can be adopted during wash-out, also gradient elution can be adopted.
14. sweet Stevia neoflavone glycoside compound separation method as claimed in claim 3, after it is characterized in that merging elutriant, can adopt acetone, chloroform, methyl alcohol, ethanol, water or their at least two kinds of admixture solvent recrystallizations, obtain sweet Stevia neoflavone glycoside compound.
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