JPH0532029B2 - - Google Patents

Description

[Detailed description of the invention]

(Industrial Application Field) This invention produces polysaccharides, particularly peptidoglycans having hypoglycemic effects named panaxane A, B, C, D and E, which can be isolated from medicinal ginseng, by culturing medicinal ginseng callus. Regarding the method. (Prior art) Medicinal ginseng belonging to the Araliaceae family, particularly Panax ginseng (Panax ginseg CAMayer), has been used as a tonic, tonic, diuretic, anti-inflammatory, and anti-diabetic drug since ancient times. ing. Regarding the components contained in this medicinal ginseng, various saponins are known as components soluble in lower aliphatic alcohols. In addition to these saponins, the inventor discovered 1, which is completely different from saponins, as a medicinal ingredient that is contained in the water-soluble fraction and has a hypoglycemic effect.
We have discovered a new group of peptidoglycans,
These were named panaxan (patent application 1983-
29083). Among the ingredients contained in medicinal ginseng, it is already known that saponins can be produced by tissue culture, and some knowledge has been reported regarding the culture conditions. It is completely unknown to do so. Panaxan is not produced from plant callus obtained by conventional tissue culture. (Problems to be Solved by the Invention) The present invention solves the above conventional problems, and its purpose is to provide a method for effectively producing panaxanes by culturing medicinal carrot callus. It is about providing. (Means for Solving the Problems) The present invention has been made through repeated research on a method for producing panaxane by callus culture of medicinal carrots, and as a result, we have succeeded in isolating a strain capable of producing panaxan, and established a cultivation method for it. did. Panaxanes are polysaccharides contained in the water-soluble fraction of medicinal ginseng. The method for producing polysaccharides by callus culture of the present invention includes:
Panax ginseng (Panax)
ginseg CAMayer), and extracting at least one member selected from the group consisting of panaxan A, panaxan B, panaxan C, panaxan D, and panaxan E having the following characteristics from the callus culture. encompasses,
This achieves the above objective. Panaxane A Molecular weight (gel filtration method): 14000 Specific optical rotation: [α] _D +187° (c 0.23, water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ):
3370, 1014 ¹ H nuclear magnetic resonance δ: 4.89 (d, J3Hz); 5.20 (d,
J3Hz) Elemental analysis value (%): C, 40.38; H, 5.97; N,
0.15 Gel chromatography (TSK Gel G
4000W column) elution time: 29.3 minutes DEAE-cellulose chromatography elution time: 3.3 hours Panaxane B Molecular weight (gel filtration method): 4000000 Specific rotation: [α] _D +180° (c 0.12, water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ):
3430, 1012 ¹ H nuclear magnetic resonance δ: 4.88 (d, J3Hz); 5.24 (d,
J3Hz) Elemental analysis value (%): C, 43.41; H, 5.99; N,
0.42 Gel chromatography (TSK Gel G
4000W column) elution time: 18.1 minutes DEAE-cellulose chromatography elution time: 3.3 hours Panaxane C Molecular weight (gel filtration method): 34000 Specific rotation: [α] _D +96.1° (c 0.10, water) Infrared Absorption spectrum (KBr) νmax (cm ^-1 ):
3390, 1021 ¹ H nuclear magnetic resonance δ: 4.88 (d, J3Hz); 5.18 (d,
J3Hz) Elemental analysis value (%): C, 35.37; H, 5.35; N,
1.06 Gel chromatography (TSK Gel G
4000W column) elution time: 28.9 minutes DEAE-cellulose chromatography elution time: 7.3 hours Panaxane D Molecular weight (gel filtration method): 350000 Specific optical rotation: [α] _D +126° (c 1.01, water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ):
3420, 1035 ¹ H nuclear magnetic resonance δ: 4.92 (d, J3Hz); 5.23 (d,
J3Hz) Elemental analysis value (%): C, 39.20; H, 6.02; N,
1.23 Gel chromatography (TSK Gel G
4000W column) elution time: 28.0 minutes DEAE-cellulose chromatography elution time: 7.3 hours Panaxane E Molecular weight (gel filtration method): 1500000 Specific optical rotation: [α] _D +188° (c 0.20, water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ):
3390, 1017 ¹ H nuclear magnetic resonance δ: 4.87 (d, J3Hz); 5.22 (d,
J3Hz) Elemental analysis value (%): C, 37.41; H, 6.25; N,
0.68 Gel chromatography (TSK Gel G
Elution time for DEAE-cellulose chromatography (4000W column): 17.7 minutes Elution time for DEAE-cellulose chromatography: 7.3 hours According to the method of the present invention, first, a part of the medicinal carrot tissue is cultured in a conventional manner to induce callus. Panax ginseng is preferably used as the medicinal ginseng. For example, a part of the tissue (root, stem, etc.) of this Panax ginseng is cut out, the cut end and surface are thoroughly sterilized using an aqueous disinfectant solution, and then washed with sterile water. This is cut into rounds with a thickness of 5 to 10 mm, and statically cultured at 20 to 25°C using a Murashige-Skoog agar medium or the like. The medium used does not need to be special, and the medium used is the Murashige-
In addition to Skoog's medium, White's medium commonly used for plant tissue culture, Linsmaier-Skoog's medium,
Gautheret's medium, Tulecke's medium, Morel's medium, etc. can be used. In order to promote callus induction, plant hormones (auxins and cytokinins) are added to the medium in advance using a conventional method.
is added. For example, auxins include 2.4
- Dichlorophenoxyacetic acid (2.4-D), idoleacetic acid (1AA), and naphthaleneacetic acid (NAA) are added to the medium at a rate of 0.1 to 10 ppm.
Examples of cytokinins include kinetin and benzyladenine, which are added to the medium at a rate of 0.05 to 5 ppm. The callus induced in this way has a white color. This white callus is transferred to a callus culture medium and statically cultured. As the callus culture medium, a known medium (a solid medium such as an agar medium or a liquid medium) such as Murashige-Skoog's medium can be used as in the above-mentioned callus induction medium. In addition, glucose
It is added in a proportion of 0.1 to 5% by weight, preferably 0.1 to 2% by weight. The amount of sucrose contained in the medium may be reduced depending on the amount of glucose added. The added glucose is taken up into the callus and acts as a precursor of panaxan, so that panaxan can be advantageously produced within the callus. If the amount of glucose is too low, the production of panaxan will be low.
Even if it is excessive, it has no further effect. The amount of plant hormone contained in the callus culture medium is adjusted to be smaller than the amount of plant hormone in the callus induction medium. Usually, auxins are contained at a ratio of 5 ppm or less and cytokinins at a ratio of 2 ppm or less.
It may not contain any plant hormones at all.
When the proliferated white callus is subcultured, calli with different colors and shapes appear. From white callus to brown callus (type B strain callus) and differentiated callus with a shape similar to a leaf (L type strain callus) are produced. From the L-type callus, a callus (R-type callus) having a shape closer to a root is generated. Four types of callus are obtained, including a white callus (W-type strain callus) obtained by subculture in a callus culture medium. The W-type callus has almost the same properties as the white callus derived from the tissue of Panax ginseng, but has a higher panaxan production ability than the white callus. W-type strain callus does not have the ability to produce saponins and alkaloids that are normally contained in medicinal ginseng. B-type strain callus, L-type strain callus, and R-type strain callus have the ability to produce not only panaxanes but also saponins and alkaloids. To obtain panaxanes from these calluses,
First, callus is extracted with water or an aqueous organic solvent, either as it is or after drying. Extracting after drying the callus increases the extraction rate of panaxan. This is thought to be because panaxanes are contained in the cell walls of callus, and therefore panaxanes can be extracted advantageously by dying the callus cells. Although water is sufficient for extraction, an aqueous organic solvent may be used to prevent spoilage of the extract and to accelerate extraction. Alternatively, both may be extracted.
As the aqueous organic solvent, a mixed solution of water and a lower alcohol such as methanol or ethanol, or a water-soluble organic solvent such as acetone, acetonitrile, dimethylformamide, or dimethylacetamide is used. The concentration varies depending on the dry state of the callus, but is approximately 1 to 60%, preferably approximately 1 to 20%.
%. Heating is preferable because it accelerates extraction. The above extract is subjected to dialysis or ultrafiltration to obtain a panaxane mixture. When subjecting to dialysis or ultrafiltration, the above extract may be used as it is, or may be concentrated by distilling off the solvent. In addition, after adding a water-soluble organic solvent such as methanol or ethanol to the above extract to precipitate a precipitate containing the desired panaxane,
The obtained precipitate may be diluted by adding an appropriate amount of water or the above-mentioned aqueous organic solvent. The panaxane mixture thus obtained usually contains at least 60% of the total amount of panaxanes A, B, C, D and E. It is desirable that the panaxane content is 80% or more. Such panaxane mixtures can themselves be used as hypoglycemic agents. However, this mixture may be further subjected to the following fractionation treatment to isolate each panaxane component and use it as a hypoglycemic agent. First, the above panaxane mixture is treated with an anion exchange resin such as DEAE Cephadex and DEAE Toyopearl (both trade names) to form a fraction containing panaxane A and B and a fraction containing panaxane C, D and E. Fractionate. The fraction containing panaxane A and B thus obtained is then subjected to molecular weight fractionation. Examples of this molecular weight fractionation method include a gel filtration method using Sepharose, Cephacryl, Sephadex, Agarose beads (all trade names), and a method using an ultrafiltration membrane with a molecular weight cutoff of 20,000 to 100,000. In this way, molecular weight fractionation method,
For example, by column chromatography using Sepharose 6B, it is fractionated into a fraction containing panaxane A and a fraction containing panaxane B. The fraction containing Panaxane B is further purified using Cephacryl S-500 (trade name) or the like to obtain Panaxane B. The previously obtained fractions containing panaxane C, D and E were further purified by molecular weight fractionation, for example column chromatography using Sepharose 6B. It is fractionated into Fractions containing panaxane C and D can be further analyzed by molecular weight fractionation methods such as Cephacryl S-200.
Column chromatography method using (trade name),
The fraction is separated into a fraction containing panaxane C and a fraction containing panaxane D. (Effect) In this way, panaxanes are advantageously produced by culturing medicinal carrot callus. The yield of the obtained panaxanes and the respective content ratios of panaxane A, B, C, D and E are as follows:
Panaxanes can be advantageously extracted from any type of callus, although it differs somewhat depending on type callus, L type callus, and R type callus. isolated panaxane A,
B, C, D, and E all have excellent hypoglycemic effects, and almost no side effects are observed. (Example) The present invention will be described below with reference to Examples. Example (A) Callus induction and culture: A part of the root of Panax ginseng was cut out and immersed in a 70% ethanol solution for 3 minutes. Furthermore, it was immersed in an aqueous sodium hypochlorite solution (Cl content 1.5%) for 20 minutes, and then washed with sterile water. The roots of Panax ginseng thus sterilized were cut into rounds with a thickness of 5 to 10 mm, and placed on a callus induction medium prepared in advance. As a callus induction medium, Murashige-
IAA as auxin in Skoog's medium
A conventional agar medium for callus induction was used, to which 10 ppm and 2 ppm of kinetin as a cytokinin were added. Heat this in the dark at 20-25℃ for 30 minutes.
A white callus was formed after culturing for one day. Separately, the Succhrose content was set to 2% by weight.
Add 0.5 glucose to Murashige-Skoog's medium.
A callus culture medium (agar medium) was prepared by adding 5 ppm of IAA as auxin and 1 ppm of kinetin as cytokinin. The above white callus was transplanted onto this callus culture agar medium and cultured under the same conditions. When the proliferated callus was subcultured, a brown callus (type B strain callus) and a differentiated callus with a leaf shape (L type strain callus) appeared. White callus (W-type callus), B-type callus, and L-type callus were separated and subsequently cultured. The L-type strain was further differentiated into a callus with root morphology (R-type strain). An R-type strain was also isolated, and the respective calli were subsequently cultured. Next, a liquid medium having the same components as the above-mentioned callus culture medium was prepared, and 500 ml of the medium was dispensed into each of four single Erlenmeyer flasks, and sterilized using a conventional method. Approximately 20 g of each of the four types of calli described above were transplanted into this callus culture liquid medium, and cultured with shaking at 25° C. for 4 weeks. The medium was filtered to obtain 120 g of type W strain callus, 105 g of type B strain callus, 110 g of type L strain callus, and 93 g of type R strain callus. (B) Extraction of panaxanes from type W strain callus: (A)
After drying the type W strain callus obtained in Section 1, it was extracted once with 50 ml of 50% ethanol and twice with water. After the extracts were combined and concentrated, dialysis was performed for 2 days using a cellulose tube. When the resulting dialyzed fluid was subjected to ethanol precipitation and its weight was measured, it was found that 592 mg of crude panaxanes were contained per 100 g of callus (in terms of dry matter). This dialyzed solution was applied to a DEAE cellulose column (2 cmφ x 100 cm), and then treated with PH8 Tris-HCl buffer and then with 0.1-0.5M NaCl.
Fraction P-1 and fraction P-2 were obtained by elution with an aqueous solution. The amount of panaxane contained in Fraction P-1 and Fraction P-2 was 99 mg and 234 mg per 100 g of callus (in terms of dry matter), respectively. This fraction P-1 was transferred to a Sepharose 6B column (2 cmφ
95cm; 0.1M NaCl) to obtain fraction P-3 and fraction P-4. Fraction P-3 was concentrated to obtain 63 mg of white powder. When the physicochemical properties of this white powder were investigated, the following measured values were obtained. Molecular weight (gel filtration method): 14000 Specific rotation: [α] _D +187° (c 0.23, water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ):
3370, 1014 ¹ H nuclear magnetic resonance δ: 4.89 (d, J3Hz); 5.20 (d,
J3Hz) Elemental analysis value (%): C, 40.38; H, 5.97; N,
0.15 Glass fiber paper electrophoretic mobility: 19.7cm (glucose 13.0cm) Polyacrylamide electrophoretic mobility: 0cm Gel chromatography (TSK Gel G
4000W column) elution time: 29.3 minutes DEAE-cellulose chromatography elution time: 3.3 hours The white powder from the above data was found to be a single substance, Panaxane A. Fraction P-4 was transferred to a Sephacryl S-500 column (2 cmφ x 95 cm, 0.1M Tris-HCl buffer PH
7.0) to obtain 34 mg of white powder. When the physicochemical properties of this white powder were investigated, the following measured values were obtained. Molecular weight (gel filtration method): 4000000 Specific rotation: [α] _D +180° (c 0.12, water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ):
3430, 1012 ¹ H nuclear magnetic resonance δ: 4.88 (d, J3Hz); 5.24 (d,
J3Hz) Elemental analysis value (%): C, 43.41; H, 5.99; N,
0.42 Glass fiber paper electrophoretic mobility: 19.0cm (glucose 13.0cm) Polyacrylamide electrophoretic mobility: 0cm Gel chromatography (TSK Gel G
Elution time for DEAE-cellulose chromatography (4000W column): 18.1 minutes Elution time for DEAE-cellulose chromatography: 3.3 hours The above data revealed that this white powder was Panaxane B. Next, the fraction P
-2 to a Cearose 6B column (2 cmφ x 95 cm,
0.1M NaCl) to obtain fractions P-5 and P-6. Fraction P-
5 is Sephacryl S-200 column (2cmφ×95
cm, 0.1M Tris-HCl buffer PH7.0, 0.5M
86 mg of white powder () and
93 mg of white powder () was obtained. When the physicochemical properties of each were investigated, the following measured values were obtained. Physicochemical properties of () Molecular weight (gel filtration method): 34000 Specific rotation: [α] _D +96.1° (c 0.10, water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ):
3390, 1021 ¹ H nuclear magnetic resonance δ: 4.88 (d, J3Hz); 5.18 (d,
J3Hz) Elemental analysis value (%): C, 35.37; H, 5.35; N,
1.06 Glass fiber filter paper electrophoretic mobility: 19.0 cm (glucose 13.0 cm) Polyacrylamide electrophoretic mobility: 0 cm Gel chromatography (TSK Gel G
4000W column) elution time: 28.9 minutes DEAE-cellulose chromatography elution time: 7.3 hours () Physicochemical properties Molecular weight (gel filtration method): 350000 Specific rotation: [α] _D +126° (c 1.01, Water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ): 3420, 1035 ¹ H nuclear magnetic resonance δ: 4.92 (d, J3Hz); 5.23 (d,
J3Hz) Elemental analysis value (%): C, 39.20; H, 6.02; N,
1.23 Glass fiber paper electrophoretic mobility: 19.0 cm (glucose 13.0 cm) Polyacrylamide electrophoretic mobility: 0 cm Gel chromatography (TSK Gel G
Elution time for DEAE-cellulose chromatography (4000W column): 28.0 minutes Elution time for DEAE-cellulose chromatography: 7.3 hours From the above data, these white powders () and () were found to be Panaxane C and Panaxane D, respectively. Fraction P-6 was concentrated to yield 23 mg of white powder. When the physicochemical properties of this white powder were investigated, the following measured values were obtained. Molecular weight (gel filtration method): 1500000 Specific rotation: [α] _D +188° (c 0.20, water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ):
3390, 1017 ¹ H nuclear magnetic resonance δ: 4.87 (d, J3Hz); 5.22 (d,
J3Hz) Elemental analysis value (%): C, 37.41; H, 6.25; N,
0.68 Glass fiber paper electrophoretic mobility: 19.3 cm (glucose 13.0 cm) Polyacrylamide electrophoretic mobility: 0 cm Gel chromatography (TSK Gel G
4000W column) elution time: 17.7 minutes DEAE-cellulose chromatography elution time: 7.3 hours From the above data, this white powder is Panaxane E.
It turned out to be. When the decomposition properties of Panaxane A, B, C, D and E to acids were investigated, the following results were obtained. Acid hydrolysis 2N sulfuric acid was added to each of Panaxane A, B, C, D, and E, and the mixture was heated at 100° C. for 6 hours to perform hydrolysis. Thin layer chromatography (developing solvent:
When a mixture of n-butanol-pyridine-water (6:4:3) was used and color was developed with a p-anisidine hydrochloric acid solution, a single spot was observed at Rf0.29. This spot matched the control glucose spot. Further, 6N hydrochloric acid was added to each of Panaxane A to E, and the mixture was heated at 100° C. for 2 hours to perform hydrolysis. Thin layer chromatography (developing solvent: n
-butanol-acetic acid-water (12:3:5) mixture)
After color development with ninhydrin, several spots were observed. Among the above data, molecular weight, glass fiber filtration electrophoresis, polyacrylamide electrophoresis, gel chromatography elution time,
The elution time of DEAE-cellulose chromatography was measured as follows. The component other than C, H, and N in the elemental analysis value is O. (1) Molecular weight Each panaxane is Cephacryl S-200,
Gel filtration was performed using 300 or 500 to determine each retention capacity, and the molecular weight was calculated from a standard curve prepared using Dextran T series. Molecular weight used for measurement Cefacrylpanaxane A 14000 S-200 Panaxan B 4000000 S-500 Panaxan C 34000 S-200 Panaxan D 350000 S-300 Panaxan E 1500000 S-500 (2) Glass fiber filter paper Electrophoretic mobility Glass fiber Filter paper (Watsutman GF/
C.15 x 40 cm) was used to measure the distance traveled.
(Conditions: alkaline borate buffer PH9.450V,
(2 hours) (3) Polyacrylamide electrophoresis The migration distance was measured using a 30% polyacrylamide gel column (inner diameter 10 cm). (Conditions: borate buffer PH9.3, 2 mA, 2 hours,
Color development using thymol sulfate method) (4) Elution time of gel chromatography TSK gel G4000W column (inner diameter 0.75 cm,
Elution was performed with a 0.1M aqueous sodium chloride solution (0.5ml/min.) using a 60cm long tube. (5) Elution time of DEAE-cellulose chromatography DEAE-cellulose column (inner diameter 2 cm,
Elution was performed using a 0.05M Tris-HCl buffer (PH8.0) for 5 hours, and further elution was performed using a buffer prepared by adding sodium chloride to the above buffer. (0~0.5M
NaCl, 20 hours, elution rate 1 ml/min.). (C) Extraction of panaxanes from type B strain callus: After drying the type B strain callus obtained in section (A),
Using this, extraction was performed in the same manner as in section (B). Dialysis fluid after dialysis treatment, fraction P-
The amounts of panaxanes contained in Fraction No. 1 and Fraction P-2 were 729 mg, 68 mg, and 370 mg per 100 g (dry matter equivalent) of callus, respectively. The dialyzed fluid was treated in the same manner as in section (B), and the white powders obtained were confirmed to be panaxane A, B, C, D, and E, respectively. The yield was 28 mg of panaxan A, 22 mg of panaxan B, 86 mg of panaxan C, 93 mg of panaxan D, and 45 mg of panaxan E. (D) Extraction of panaxanes from L-type strain callus: After drying the L-type strain callus obtained in section (A),
Using this, extraction was performed in the same manner as in section (B). Dialysis fluid after dialysis treatment, fraction P-
The amounts of panaxane contained in Fraction No. 1 and Fraction P-2 were 524 mg, 67 mg, and 382 mg per 100 g (dry matter equivalent) of callus, respectively. The dialyzed fluid was treated in the same manner as in section (B), and the white powders obtained were confirmed to be panaxane A, B, C, D, and E, respectively. The yield was 22 mg of panaxan A, 18 mg of panaxan B, 128 mg of panaxan C, 86 mg of panaxan D, and 55 mg of panaxan E. (E) Extraction of panaxanes from R type strain callus: After drying the R type strain callus obtained in section (A),
Using this, extraction was performed in the same manner as in section (B). Dialysis fluid after dialysis treatment, fraction P-
The amounts of panaxane contained in Fraction No. 1 and Fraction P-2 were 268 mg, 105 mg, and 133 mg per 100 g (dry matter equivalent) of callus, respectively. The dialysis fluid was treated in the same manner as in section (B), and the white powders obtained were panaxane A, B, C, and D, respectively.
and E. The yield was 36 mg of panaxan A, 21 mg of panaxan B, 130 mg of panaxan C, 172 mg of panaxan D, and 46 mg of panaxan E. Comparative example: 100g of dried roots of Panax ginseng are crushed and 50%
Extraction was performed once with 200 ml of methanol aqueous solution and twice with 300 ml of water. The extracts were combined and concentrated under reduced pressure to obtain 38 g of extract. Add this extract to 50g of cellulose powder (Watsuman GF11; trade name).
mixed with. The above mixture was layered on the cellulose layer of a column previously filled with cellulose powder of the same quality, and methanol was injected to wash the column. After further washing with a 50% methanol aqueous solution, elution was performed with water. The obtained fraction was concentrated to obtain 3.8 g of a water-soluble fraction. This water-soluble fraction was collected in a cellulose tube (manufactured by Visking Co.).
Dialysis was performed for 4 days using The resulting dialyzed fluid contained 1082 mg of panaxane per 100 g of dried Panax ginseng root. Dialysis fluid
Molecular weight fractionation was performed using a DEAE cellulose column (2 cmφ x 100 cm) (PH8.0 Tris-HCl buffer, 0.1-0.5M NaCl) to obtain fractions P-1 and P-2. Fraction P-
Fraction P-2 contained panaxanes in a total amount of 619 mg per 100 g of dried Panax ginseng root, and Fraction P-2 contained a total of 412 mg of panaxanes per 100 g of dried Panax ginseng root. The contents of panaxanes contained in the dialysis fluid, fraction P-1, and fraction P-2 in Examples and Comparative Examples are summarized in the table below.

[Table] (Effects of the Invention) According to the present invention, callus having a high production capacity for polysaccharides and panaxanes contained in medicinal ginseng can be effectively tissue cultured in a medium containing glucose. Therefore, a large amount of panaxane can be produced in a short period of time without having to spend a long period of time cultivating medicinal ginseng as in the past. Since tissue culture is used, it is not affected by natural conditions such as weather. The obtained panaxanes have a hypoglycemic effect and can be used as various forms of drugs.

Claims (1)

[Scope of Claims] 1. Panax ginseng (Panax), which belongs to the family Araliaceae.
Panaxan A, Panaxan B, Panaxan C,
A method for producing a polysaccharide having a hypoglycemic effect, which comprises extracting at least one member selected from the group consisting of panaxan D and panaxan E. Panaxane A Molecular weight (gel filtration method): 14000 Specific optical rotation: [α] _D +187° (c 0.23, water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ):
3370, 1014 ¹ H nuclear magnetic resonance δ: 4.89 (d, J3Hz); 5.20 (d,
J3Hz) Elemental analysis value (%): C, 40.38; H, 5.97; N,
0.15 Gel chromatography (TSK Gel G
4000W column) elution time: 29.3 minutes DEAE-cellulose chromatography elution time: 3.3 hours Panaxane B Molecular weight (gel filtration method): 4000000 Specific rotation: [α] _D +180° (c 0.12, water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ):
3430, 1012 ¹ H nuclear magnetic resonance δ: 4.88 (d, J3Hz); 5.24 (d,
J3Hz) Elemental analysis value (%): C, 43.41; H, 5.99; N,
0.42 Gel chromatography (TSK Gel G
4000W column) elution time: 18.1 minutes DEAE-cellulose chromatography elution time: 3.3 hours Panaxane C Molecular weight (gel filtration method): 34000 Specific rotation: [α] _D +96.1° (c 0.10, water) Infrared Absorption spectrum (KBr) νmax (cm ^-1 ):
3390, 1021 ¹ H nuclear magnetic resonance δ: 4.88 (d, J3Hz); 5.18 (d,
J3Hz) Elemental analysis value (%): C, 35.37; H, 5.35; N,
1.06 Gel chromatography (TSK Gel G
4000W column) elution time: 28.9 minutes DEAE-cellulose chromatography elution time: 7.3 hours Panaxane D Molecular weight (gel filtration method): 350000 Specific optical rotation: [α] _D +126° (c 1.01, water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ):
3420, 1035 ¹ H nuclear magnetic resonance δ: 4.92 (d, J3Hz); 5.23 (d,
J3Hz) Elemental analysis value (%): C, 39.20; H, 6.02; N,
1.23 Gel chromatography (TSK Gel G
4000W column) elution time: 28.0 minutes DEAE-cellulose chromatography elution time: 7.3 hours Panaxane E Molecular weight (gel filtration method): 1500000 Specific optical rotation: [α] _D +188° (c 0.20, water) Infrared absorption spectrum (KBr) νmax (cm ^-1 ):
3390, 1017 ¹ H nuclear magnetic resonance δ: 4.87 (d, J3Hz); 5.22 (d,
J3Hz) Elemental analysis value (%): C, 37.41; H, 6.25; N,
0.68 Gel chromatography (TSK Gel G
4000W column) elution time: 17.7 minutes DEAE-cellulose chromatography elution time: 7.3 hours 2. Claim 1, wherein the medium used for the callus culture contains 0.1 to 5.0% by weight of glucose.
Manufacturing method described in Section.