Xiangyang radix ophiopogonis endophytic penicillium oxalicum and application in steroid saponin preparation

Abstract

The invention discloses Xiangyang radix ophiopogonis endophytic penicillium oxalicum and application in steroid saponin preparation, and relates to the technical field of microorganisms. The problemsthat Xiangyang radix ophiopogonis steroid saponin is mostly directly from Xiangyang radix ophiopogonis extractives, and the Xiangyang radix ophiopogonis steroid saponin is limited in market delivery quantity due to the fact that Xiangyang radix ophiopogonis is long in earthnut production period, low in steroid saponin content and high in saponin extraction cost are solved. Penicillium oxalicum isseparated from Xiangyang radix ophiopogonis (Liriope spicata var.proliferaY.T.Ma) living earthnuts, it is determined as penicillium oxalicum through 18S rDNA sequence authentication, the penicillium oxalicum is preserved in the China Center for Type Culture Collection, the preservation date is 7th December, 2016, and the preservation number is CCTCC No:M2016722. By means of Xiangyang radix ophiopogonis produced in a Xiangyang radix ophiopogonis GAP base-the source area of Xiangyang radix ophiopogonis in Oumiao county, Xiangyang city, Hubei province, due to fermentation of Xiangyang radix ophiopogonis endophytic fungi strain liquid, the Xiangyang radix ophiopogonis steroid saponin is produced. The endophytic fungi are the important microorganism for searching for new resources of the Xiangyang radix ophiopogonis steroid saponin.

Description

technical field

[0001] The invention relates to the technical field of microorganisms, in particular to an endophytic fungus of Ophiopogon japonicus. The endophytic fungus is an endophytic Penicillium oxalicum capable of producing steroidal saponins from Ophiopogon japonicus. Background technique

[0002] Xiangmaidong steroidal saponins are national geographical indication products, authentic Chinese medicinal plants in Hubei——Xiangmaidong Liriope spicatavar. prolifera The main active ingredient of Y. T. Ma tuber root, its saponin basic skeleton belongs to the derivative of Spirostane (Spirostane, which has 27 carbon atoms in total), according to the configuration of C25 and the ring closure state of the ring in the Spirostane structure, it can be It is divided into spirostanols (C25 is S configuration), isospirostanols (C25 is R configuration), furostanols (F ring is an open-chain derivative), deformed spirostanols Alcohols (F ring is a five-membered tetrahydrofuran rin...

Examples

Embodiment 1

[0048] The Xiangmaidong living tuber used for the isolation of the Xiangmaidong endophytic fungus EF036 strain of the present invention comes from Xiangmaidong GAP base, the place of origin of Xiangmaidong in Oumiao Town, Xiangyang City, Hubei Province.

[0049] The endophytic fungus of the present invention is separated and obtained according to the following steps:

[0050] (1) Use a large amount of tap water to fully clean the soil on the surface of the root tuber, then put it into a clean beaker, add deionized water, and place it in an ultrasonic cleaner for repeated cleaning until the water after cleaning becomes extremely clear.

[0051] (2) After dipping the moisture on the surface of the tuber with sterile filter paper, proceed as follows: first soak the tuber with 75% (V / V) ethanol for 2 minutes, rinse it with sterile water for 3 times; Soak in -6% sodium hypochlorite solution for 2 minutes, then rinse continuously with a large amount of sterile water for 4 times, and...

Embodiment 2

[0055] (1) Take the endophytic fungus EF036 bacterial classification of the present invention, under aseptic conditions, pick a small amount of hyphae with an inoculation needle, insert into a sterilized PDA medium test tube, and activate and cultivate for 72 hours at 28°C;

[0056] (2) Take the strains after the activated culture, transfer them into the sterilized liquid potato seed culture medium under aseptic conditions, and shake and cultivate them at 180 rpm for 72 hours at 28° C. to obtain seeds;

[0057] (3) The prepared described liquid fermentation medium is packed into 250ml triangular flasks respectively, about 100ml of each bottle, sterilized, and cooled for standby; under aseptic conditions, insert seeds according to 10% inoculum size, 28 Cultivate with shaking at 180rpm for 7 days at ℃;

[0058] (4) After the fermentation is completed, the fermented solid-liquid mixture is placed in an ultra-low temperature refrigerator to be quickly frozen into a solid state, an...

technical field

[0001] The invention relates to the technical field of microorganisms, in particular to an endophytic fungus of Ophiopogon japonicus. The endophytic fungus is an endophytic Penicillium oxalicum capable of producing steroidal saponins from Ophiopogon japonicus. Background technique

[0002] Xiangmaidong steroidal saponins are national geographical indication products, authentic Chinese medicinal plants in Hubei——Xiangmaidong Liriope spicatavar. prolifera The main active ingredient of Y. T. Ma tuber root, its saponin basic skeleton belongs to the derivative of Spirostane (Spirostane, which has 27 carbon atoms in total), according to the configuration of C25 and the ring closure state of the ring in the Spirostane structure, it can be It is divided into spirostanols (C25 is S configuration), isospirostanols (C25 is R configuration), furostanols (F ring is an open-chain derivative), deformed spirostanols Alcohols (F ring is a five-membered tetrahydrofuran rin...

Examples

Embodiment 1

[0048] The Xiangmaidong living tuber used for the isolation of the Xiangmaidong endophytic fungus EF036 strain of the present invention comes from Xiangmaidong GAP base, the place of origin of Xiangmaidong in Oumiao Town, Xiangyang City, Hubei Province.

[0049] The endophytic fungus of the present invention is separated and obtained according to the following steps:

[0050] (1) Use a large amount of tap water to fully clean the soil on the surface of the root tuber, then put it into a clean beaker, add deionized water, and place it in an ultrasonic cleaner for repeated cleaning until the water after cleaning becomes extremely clear.

[0051] (2) After dipping the moisture on the surface of the tuber with sterile filter paper, proceed as follows: first soak the tuber with 75% (V / V) ethanol for 2 minutes, rinse it with sterile water for 3 times; Soak in -6% sodium hypochlorite solution for 2 minutes, then rinse continuously with a large amount of sterile water for 4 times, and...

Embodiment 2

[0055] (1) Take the endophytic fungus EF036 bacterial classification of the present invention, under aseptic conditions, pick a small amount of hyphae with an inoculation needle, insert into a sterilized PDA medium test tube, and activate and cultivate for 72 hours at 28°C;

[0056] (2) Take the strains after the activated culture, transfer them into the sterilized liquid potato seed culture medium under aseptic conditions, and shake and cultivate them at 180 rpm for 72 hours at 28° C. to obtain seeds;

[0057] (3) The prepared described liquid fermentation medium is packed into 250ml triangular flasks respectively, about 100ml of each bottle, sterilized, and cooled for standby; under aseptic conditions, insert seeds according to 10% inoculum size, 28 Cultivate with shaking at 180rpm for 7 days at ℃;

[0058] (4) After the fermentation is completed, the fermented solid-liquid mixture is placed in an ultra-low temperature refrigerator to be quickly frozen into a solid state, an...

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