CN103060409A - Method for extracting active collagen from sea cucumber - Google Patents

Abstract

The invention relates to a method for extracting active collagen from sea cucumber. The method comprises the following steps: removing internal organs from fresh and live sea cucumber; cutting into fragments; stirring with distilled water; after filtering, stirring with Tris-HCl; after filtering, adding collagenous fiber precipitate obtained by centrifugation into NaOH solution; stirring and centrifuging; adding the precipitate into pepsase-containing acetic acid solution; stirring and passing the night; centrifuging; and dialyzing the precipitate to obtain the active collagen, wherein all the operation is performed at the temperature of between 0 and 4 DEG C. The process is simple and high in extraction ratio; the three-dimensional spiral structure of the collagen is preserved well; and high scientific research and application value is achieved.

Description

A kind of extracting method of sea cucumber active collagen

Technical field

The present invention relates to collagen protein extractive technique field, be specifically related to a kind of extracting method of sea cucumber active collagen.

Background technology

Contain a large amount of collagen proteins in the sea cucumber, have anticoagulation, immunomodulatory, reducing blood-fat, the multiple pharmacological effect such as antiviral, antitumor, and the using value of the holothurian collagen of maintenance activity is higher, but the extraction process of collagen protein all can be destroyed its three-dimensional spiral structure at present, greatly reduces scientific research and the using value of its collagen protein.

Summary of the invention

For solving the problems of the technologies described above, the purpose of this invention is to provide a kind of extracting method of sea cucumber active collagen.

Technical scheme of the present invention is: comprise following extraction step:

(1) fresh and alive sea cucumbers is removed internal organ, reject the inwall muscle layer with scissors, the residue wall of sea cucumber Stichopus japonicus is cut into fragment by knife;

(2) distilled water of 10 times of weight of adding stirs 30 min, leaches with gauze, and the distilled water that adds again 10 times of weight is heavily washed once;

(3) then the sea cucumber fragment is put in the 0.1 mol/L Tris-HCl solution of 10 times of weight, pH value of solution is 8.0, also contains the EDTA of 4 m mol/L, and stirring is spent the night;

(4) with after the filtered through gauze, again with distillation washing 2 times, place the distilled water stirring of 10 times of weight to spend the night, namely be full of cotton-shaped collegen filament in the solution, leach the sea cucumber piece that does not dissociate with gauze, repeat aforesaid operations;

(5) with solution with centrifugal 30 min of 12000rpm;

(6) gained collegen filament precipitation adds in the 0.1 mol/L NaOH solution of 25 times of volumes, stirs 2d;

(7) the centrifugal 30min of 12000rpm, precipitation is washed to neutrality, adds 0.5 mol/L acetic acid of 25 times of volumes, adds 0.5% stomach en-, stirs, digests 2 days;

(8) the centrifugal 30min of 12000rpm, supernatant liquor slowly add the meticulous NaCl of grinding, and the solution final concentration is 0.9 mol/L, and stirring is spent the night;

(9) the centrifugal 30min of 12000rpm removes supernatant, and precipitation adds a little 0.5 mol/L acetic acid dissolving, change in the dialysis tubing 0.1 mol/L acetic acid dialysis 2d, use distill water dialysis 2d again, namely get active collagen after the lyophilize, above all operations all carries out under 4 ℃.

Technique of the present invention is simple, and extraction yield is high, and the three-dimensional spiral structure of collagen protein is intact, has higher scientific research and using value.

Embodiment

Embodiment 1

A kind of instant sea cucumber new process for processing comprises following extraction step:

(1) fresh and alive sea cucumbers is removed internal organ, reject the inwall muscle layer with scissors, the residue wall of sea cucumber Stichopus japonicus is cut into fragment by knife;

(2) distilled water of 10 times of weight of adding stirs 30 min, leaches with gauze, and the distilled water that adds again 10 times of weight is heavily washed once;

(3) then the sea cucumber fragment is put in the 0.1 mol/L Tris-HCl solution of 10 times of weight, pH value of solution is 8.0, also contains the EDTA of 4 m mol/L, and stirring is spent the night;

(4) with after the filtered through gauze, again with distillation washing 2 times, place the distilled water stirring of 10 times of weight to spend the night, namely be full of cotton-shaped collegen filament in the solution, leach the stichopus japonicus piece that does not dissociate with gauze, repeat aforesaid operations;

(5) with solution with centrifugal 30 min of 12000rpm;

(6) gained collegen filament precipitation adds in the 0.1 mol/L NaOH solution of 25 times of volumes, stirs 2d;

(7) the centrifugal 30min of 12000rpm, precipitation is washed to neutrality, adds 0.5 mol/L acetic acid of 25 times of volumes, adds 0.5% stomach en-, stirs, digests 2 days;

(8) the centrifugal 30min of 12000rpm, supernatant liquor slowly add the meticulous NaCl of grinding, and the solution final concentration is 0.9 mol/L, and stirring is spent the night;

(9) the centrifugal 30min of 12000rpm removes supernatant, and precipitation adds a little 0.5 mol/L acetic acid dissolving, change in the dialysis tubing 0.1 mol/L acetic acid dialysis 2d, use distill water dialysis 2d again, namely get active collagen after the lyophilize, above all operations all carries out under 4 ℃.

Above-described embodiment is not as a limitation of the invention, all any modifications of doing within the scope of the invention, is equal to replacement, improvement etc., all belongs to protection scope of the present invention.

Claims (1)

1. the extracting method of a sea cucumber active collagen is characterized in that: comprise following extraction step:

(1) fresh and alive sea cucumbers is removed internal organ, reject the inwall muscle layer with scissors, the residue wall of sea cucumber Stichopus japonicus is cut into fragment by knife;

(2) distilled water of 10 times of weight of adding stirs 30 min, leaches with gauze, and the distilled water that adds again 10 times of weight is heavily washed once;

(3) then the sea cucumber fragment is put in the 0.1 mol/L Tris-HCl solution of 10 times of weight, pH value of solution is 8.0, also contains the EDTA of 4 m mol/L, and stirring is spent the night;

(4) with after the filtered through gauze, again with distillation washing 2 times, place the distilled water stirring of 10 times of weight to spend the night, namely be full of cotton-shaped collegen filament in the solution, leach the sea cucumber piece that does not dissociate with gauze, repeat aforesaid operations;

(5) with solution with centrifugal 30 min of 12000rpm;

(6) gained collegen filament precipitation adds in the 0.1 mol/L NaOH solution of 25 times of volumes, stirs 2d;

(7) the centrifugal 30min of 12000rpm, precipitation is washed to neutrality, adds 0.5 mol/L acetic acid of 25 times of volumes, adds 0.5% stomach en-, stirs, digests 2 days;

(8) the centrifugal 30min of 12000rpm, supernatant liquor slowly add the meticulous NaCl of grinding, and the solution final concentration is 0.9 mol/L, and stirring is spent the night;

(9) the centrifugal 30min of 12000rpm removes supernatant, and precipitation adds a little 0.5 mol/L acetic acid dissolving, change in the dialysis tubing 0.1 mol/L acetic acid dialysis 2d, use distill water dialysis 2d again, namely get active collagen after the lyophilize, above all operations all carries out under 4 ℃.