Method for separating estrogen from placenta

Abstract

The present invention provides a method of separating estrogen from placenta, which uses supercritical fluid technique to load the placenta powder of human body or sheep, pig, deer and other animals into an extraction tank. Under the operating conditions of preset pressure and temperature, supercritical solvent is added into the extraction tank to extract estrogen from placenta, so as to acquire de-estrogen placenta powder and placenta extract liquor. Under the same condition, the de-estrogen placenta extract liquor and supercritical solvent are added by a preset volume flow ratio into an adsorption tank. The estrogen in the placenta extract liquor is adsorbed by the adsorption tank to obtain de-estrogen placenta extract. It is then eluted with ethanol solution by gradient proportion to obtain purified natural estrogen.

Claims

1. A method of separating estrogen from placenta, comprising steps of: performing an extraction under 24-26MPa and 40 Degree Celsius in an extraction tank made of stainless steel monomer materials, comprising: loading 1 Kilogram of a placenta powder into the extraction tank, wherein the placenta powder is a dry placenta powder of an animal; adding a supercritical solvent into the extraction tank by a flow ratio of 15:1, wherein the supercritical solvent is a supercritical carbon dioxide or a supercritical ethanol; and producing a placenta liquor by mixing the placenta powder with the supercritical solvent inside the extraction tank for 2-3 Hours, wherein the placenta liquor is constituted of an estrogen extract and a remaining substance; performing an adsorption under 24-26MPa and 40 Degree Celsius in an adsorption tank, comprising: filling an adsorbent into the adsorption tank, wherein the adsorbent is a silica gel, a sephadex or a resin; adding the placenta liquor into the adsorption tank; and obtaining the estrogen extract by rendering the adsorbent to adsorb the estrogen extract and not to adsorb the remaining substance; and performing a gradient elution, comprising: providing an ethanol solution with a gradient proportion ranged from 50% to 80%; and desorbing the estrogen extract from the adsorbent by executing the ethanol solution to the adsorbent.

2. The method of separating estrogen from placenta of claim 1, wherein a rejection rate reaches 90% by performing the extraction under 24-26 MPa and 40 Degree Celsius in the extraction tank and adding the supercritical solvent into the extraction tank by the flow ratio of 15:1.

3. The method of separating estrogen from placenta of claim 1, wherein the extraction tank is a stainless steel tank with an internal diameter of 60 mm, an external diameter of 130 mm and a height of 130 mm.

4. The method of separating estrogen from placenta of claim 1, wherein the adsorption tank is a stainless steel tank with an internal diameter of 36 mm, an external diameter of 48 mm and a height of 597 mm.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) FIG. 1 is a flow chart of a preferred embodiment of the present invention.

(2) FIG. 2 is a diagram, showing the estrogen rejection data.

(3) FIG. 3 is a diagram, showing the analysis of de-fishy aroma, color and bitterness and the other data.

DETAILED DESCRIPTION OF THE INVENTION

(4) The following is the detailed description of a preferred embodiment of the present invention with reference to the accompanying drawings:

(5) FIG. 1 shows a preferred embodiment of the method for separating estrogen from placenta 100, whereby estrogen can be removed from placenta to acquire de-estrogen placenta powder, placenta extract and purified estrogen.

(6) Step 1 extraction 110: under the operating pressure of 20-30 MPa (e.g., 20, 22, 24, 26, 28, 30 MPa) and at the temperature of 40 C., load 1 Kg placenta powder into an extraction tank and add supercritical solvent into the extraction tank by a flow ratio of 1020:1 (supercritical carbon dioxide/ethanol), so as to extract estrogen from placenta powder and produce placenta extract liquor after 2-3 h reaction time. The placenta powder may be dry placenta powder of human body or sheep, pig, deer or other animals. The supercritical solvent is a supercritical carbon dioxide (SCCO.sub.2)/ethanol solvent. The extraction tank is a stainless steel tank of ID 60 mm, OD 130 mm and height 130 mm filled with stainless steel monomer pieces.

(7) Step 2 adsorption 120: under the same operating condition (pressure 20-30 MPa and temperature 40 C.), add the de-estrogen placenta extract liquor and supercritical solvent into an adsorption tank by a flow ratio of 1020:1, allow the estrogen in the placenta extract liquor to be adsorbed by the adsorption tank to acquire de-estrogen placenta extract. The adsorption tank is a stainless steel tank of ID 36 mm, OD 48 mm and height 597 mm filled with silica gel, sephadex or resin and other adsorbents that can adsorb estrogen.

(8) Step 3 gradient elution 130: after the estrogen is adsorbed in the adsorption tank, elute 50% ethanol solution to 80% ethanol solution by gradient proportion to acquire purified natural estrogen.

(9) The following is an implementation of said method under different operating conditions (changing the pressure and volume flow ratio of supercritical solvent). The residual extracting and extracted examples are quantitatively analyzed for their estrogen concentration and color change. The estrogen concentration is quantified by ELISA on progesterone and estriol. The color is measured with Japan-made Denshoku 90 colorimeter, indicated by Hunter L, a, b value. The color L value closer to 100 indicates a higher sample transparency. Sensory evaluations for acceptance including odor (fishy aroma), bitterness, color and overall are rated. The rating is from 0 to 10, where 10 is the most acceptable, and 0 is the most unacceptable. The highest rejection of estrogen (%), deodorization, bitterness removal, and overall acceptance are selected to evaluate the optimal reaction condition:

(10) FIG. 2 depicts the estrogen rejection data. When the pressure is 2426 MPa and temperature is 40 C., and the flow ratio of supercritical solvent is 15:1, which is the optimal operating condition, the rejection rate may reach 90%. This is mainly because the supercritical solvent under high pressure exerts high solubility and mass transportation rate, and under low temperature has low viscosity and high density of supercritical fluid, which may increase the reactant contract opportunity to accelerate the removal of estrogen from placenta powder. Therefore, the supercritical solvent under high pressure and low temperature may resolve the shortcomings when using larger number of organic solvent in hydrolyzing estrogen with acid, alkali and enzyme.

(11) In addition, the color change data of placenta before and after extraction measured with a colorimeter are shown in Table 1. The de-estrogen sample color L value is 62.4, which is higher than placenta powder color L value of 39.2 before processing. This suggests that the de-estrogen sample presents higher transparency. According to Table 1, the de-estrogen sample in present invention has deodorization, bitterness removal, high color and overall acceptance.

(12) TABLE-US-00001 TABLE 1 Analytical Items Placenta Powder SCCO2/EtOH Color L 39.2 62.4 a 17.2 14.6 b 44.7 44.2 Acceptance evaluation Odor 1.5 8.4 Color 6.8 8.2 Bitterness 1.7 7.4 Overall 3.7 8.1

(13) As shown in FIG. 3, the de-estrogen sample and extract prepare in the present invention has bright appearance, deodorized taste without bitterness, de-estrogen and good acceptance, unlike the placenta powder with odor and bitter taste, and estrogen ingredient.

(14) The method of the present invention for separating estrogen from placenta uses safe and nontoxic supercritical solvent. Physical extraction, adsorption and gradient elution are used to prepare de-estrogen placenta and its extract under supercritical CO.sub.2, which exerts the properties of high solubility, low viscosity and high mass transportation efficiency and no safety concern of residual solvent. The environmentally friendly and safe supercritical solvent can be recycled. Compared with the conventional method of using acid, alkali and enzyme for decomposition, and using organic solvent for separation, which may lead to safety concern of residual solvent, the present invention is an improved method and has practical value.