The euphorbia lunulate bge extractant and production method and use thereof

Abstract

The invention relates to the method of extracting active compounds from medicinal plant Euphorbia Lunulate Bge, consists of the following steps: The Acetone extractants of Euphorbia Lunulate Bge are abstracted and crude extracted by using organic solvents; the organic solvents extractants are subjected to inhibit production of NO activity test, and further isolate the extractants which have high production of NO inhibiting activity and no toxicity; then the isolated compositions are subjected to inhibit production of NO activity test again, and further refine the compositions which have high production of NO inhibiting activity; the achieved single compound is subjected to inhibit production of NO activity test and physical and chemical identification so as to definition its structure. The achieved compound has the activity of inhibiting production of NO, in which the structure of compound 1-3 are following formulas.

Description

technical field

[0001] The invention relates to the separation and extraction of anti-inflammatory active substances from the medicinal plant Euphorbia Lluaulata Bge and its preparation method and application. Background technique

[0002] Daily inflammatory diseases that plague people include joint inflammation such as osteoarthritis, rheumatoid arthritis, rheumatoid spondylitis, gouty arthritis, etc.; inflammatory skin diseases such as eczema, psoriasis, dermatitis, etc.; Uveitis, conjunctivitis, etc.; lung diseases such as asthma, bronchitis, acute respiratory distress syndrome, etc.; bacteremia, that toxemia, aphthous ulcers, gingivitis, pancreatitis, etc.; Ileitis, atrophic gastritis, ulcerative colitis, celiac disease, peptic ulcer, irritable bowel syndrome such as mucosal inflammation caused by Helicobacter pylori infection or gastrointestinal tract caused by non-steroidal anti-inflammatory drugs illness and so on.

[0003] Regarding the mechanism of various inflamm...

Examples

Embodiment 1

[0028] Example 1 .The preparation method of cat's eye grass extract

[0029] (1) The crude extraction method and product of cat's eye grass:

[0030] Take 9.0 kg of cat's eye grass (Euphorbia lunulata Bge), extract three times with 70% acetone (27 L), concentrate the extract, add water to the obtained extract to make it a suspension, and extract with chloroform, ethyl acetate, and n-butanol in sequence. The NO formation inhibition test and MTT test were carried out on these three extracts respectively, and the sample concentration was 100 μg / ml. The results are shown in Table 1:

[0031] Table 1. Yield, NO production inhibition rate and MTT toxicity of each extract

[0032] Extracts

Yield (g)

NO production inhibition rate

MTT toxicity

Chloroform extract

65.25

89.8％

have

Ethyl acetate extract

52.43

82.1％

none

n-butanol extract

86.51

33.1％

none

[0033] Select the ethyl acetate extrac...

Embodiment 2

[0044] Example 2 .Physical and chemical properties of main compounds

[0045] The physical and chemical parameters of the three compounds obtained by separation and purification were analyzed, and the main data are shown in Tables 3, 4 and 5.

[0046] Table 3. Basic physical and chemical properties of compounds 1-3:

[0047] Compound 1

Compound 2

Compound 3

character

yellow needle powder

yellow needle crystal

White powder

HR-MS(m / z)

Negative FAB

767.1087

767.1095

PositiveFAB

617.1096

617.1089

Negative FAB

787

molecular weight

768

616

788

molecular formula

C 35 h 27 o 20

C 28 h 24 o 16

C 38 h 43 o 14

UV lambda nm max (lgε)

269(4.17)

356(3.87)

267(4.20)

350(3.82)

230(4.21)

IR v cm-1 max

3391(OH)

1714 (C=O)

1654

1507(C=C)

3401(OH)

1712 (C=O)

1650(C=C)

3509(OH) ...

Embodiment 3

[0054] Example 3 .NO production inhibitory activity of cat's eye extract

[0055] Materials used:

[0056] RAW264.7 cells (macrophages, transformed with Abelson leukemia virus) were obtained from ATCC (American Type Culture Collection), USA. Ham's F12 medium containing 10% FBS in 5% CO 2 , cultured at 37°C.

[0057] IFN-γ: Genzyme / Techne Company; LPS: Sigma Company; Sulphonamide: Wako Company; N-1-Naphthethylene-diamine Dihyrochloride: Wako Company Griess reagent: (1) N- 1-naphthaleneethylenediamine hydrochloride was dissolved in 5ml of water for injection;

[0058] (2) Add 250 μl of phosphoric acid to 5 ml of water for injection and 50 mg of sulfonamide.

[0059] Instrument: Microtiter plate reader (Microtiter Plate Reader Model 3550, BIO-RAD).

[0060] Specific methods and results:

[0061] Take 50ml of RAW264.7 cells and put them in a Falcon tube for centrifugation (1000rpm, 3min, 4°C) to pellet the cells, remove the supernatant with a pipette, add 10ml of new mediu...

technical field

[0001] The invention relates to the separation and extraction of anti-inflammatory active substances from the medicinal plant Euphorbia Lluaulata Bge and its preparation method and application. Background technique

[0002] Daily inflammatory diseases that plague people include joint inflammation such as osteoarthritis, rheumatoid arthritis, rheumatoid spondylitis, gouty arthritis, etc.; inflammatory skin diseases such as eczema, psoriasis, dermatitis, etc.; Uveitis, conjunctivitis, etc.; lung diseases such as asthma, bronchitis, acute respiratory distress syndrome, etc.; bacteremia, that toxemia, aphthous ulcers, gingivitis, pancreatitis, etc.; Ileitis, atrophic gastritis, ulcerative colitis, celiac disease, peptic ulcer, irritable bowel syndrome such as mucosal inflammation caused by Helicobacter pylori infection or gastrointestinal tract caused by non-steroidal anti-inflammatory drugs illness and so on.

[0003] Regarding the mechanism of various inflamm...

Examples

Embodiment 1

[0028] Example 1 .The preparation method of cat's eye grass extract

[0029] (1) The crude extraction method and product of cat's eye grass:

[0030] Take 9.0 kg of cat's eye grass (Euphorbia lunulata Bge), extract three times with 70% acetone (27 L), concentrate the extract, add water to the obtained extract to make it a suspension, and extract with chloroform, ethyl acetate, and n-butanol in sequence. The NO formation inhibition test and MTT test were carried out on these three extracts respectively, and the sample concentration was 100 μg / ml. The results are shown in Table 1:

[0031] Table 1. Yield, NO production inhibition rate and MTT toxicity of each extract

[0032] Extracts

Yield (g)

NO production inhibition rate

MTT toxicity

Chloroform extract

65.25

89.8％

have

Ethyl acetate extract

52.43

82.1％

none

n-butanol extract

86.51

33.1％

none

[0033] Select the ethyl acetate extrac...

Embodiment 2

[0044] Example 2 .Physical and chemical properties of main compounds

[0045] The physical and chemical parameters of the three compounds obtained by separation and purification were analyzed, and the main data are shown in Tables 3, 4 and 5.

[0046] Table 3. Basic physical and chemical properties of compounds 1-3:

[0047] Compound 1

Compound 2

Compound 3

character

yellow needle powder

yellow needle crystal

White powder

HR-MS(m / z)

Negative FAB

767.1087

767.1095

PositiveFAB

617.1096

617.1089

Negative FAB

787

molecular weight

768

616

788

molecular formula

C 35 h 27 o 20

C 28 h 24 o 16

C 38 h 43 o 14

UV lambda nm max (lgε)

269(4.17)

356(3.87)

267(4.20)

350(3.82)

230(4.21)

IR v cm-1 max

3391(OH)

1714 (C=O)

1654

1507(C=C)

3401(OH)

1712 (C=O)

1650(C=C)

3509(OH) ...

Embodiment 3

[0054] Example 3 .NO production inhibitory activity of cat's eye extract

[0055] Materials used:

[0056] RAW264.7 cells (macrophages, transformed with Abelson leukemia virus) were obtained from ATCC (American Type Culture Collection), USA. Ham's F12 medium containing 10% FBS in 5% CO 2 , cultured at 37°C.

[0057] IFN-γ: Genzyme / Techne Company; LPS: Sigma Company; Sulphonamide: Wako Company; N-1-Naphthethylene-diamine Dihyrochloride: Wako Company Griess reagent: (1) N- 1-naphthaleneethylenediamine hydrochloride was dissolved in 5ml of water for injection;

[0058] (2) Add 250 μl of phosphoric acid to 5 ml of water for injection and 50 mg of sulfonamide.

[0059] Instrument: Microtiter plate reader (Microtiter Plate Reader Model 3550, BIO-RAD).

[0060] Specific methods and results:

[0061] Take 50ml of RAW264.7 cells and put them in a Falcon tube for centrifugation (1000rpm, 3min, 4°C) to pellet the cells, remove the supernatant with a pipette, add 10ml of new mediu...

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