HYDROCOLLOIDS OBTAINED FROM PORTULACA OLERACEA AND THE USE THEREOF
The present invention relates to hydrocolloids obtained from Portulaca Oleracea, (hereinafter "the plant") and to the use thereof .
The plant is known as is the composition thereof. (See Ali I Mohamed et . al . ; Chemical Composition of Portulaca Oleraca; Plant Foods for Human Nutrition, 45, 1994, pp. 1-9).
It is known that the plant comprises 3.5-4.5% of a hydrocol- loid which is partially soluble in water. However, no use of said hydrocolloids is known so far.
The hydrocoUoid may be obtained by extracting same from the plant by a method which is substantially described in G. Wenzel et . al.; The Viscous Mucilage from the Weed Portulaca Oleracea, L; Applied Biochemistry and Biotechnology, Vol. 24/25, 1990, pp. 341-353. Said authors analyzed the hydrocoUoid to a certain extent but did not analyze all parts of same. The results of the analysis performed by the present inventors are shown in Table I.
Table I
The hydrocoUoid was extracted in accordance with the method described by G . Wenzel et . al . The white powder obtained was analyzed, in particular the sugar contents of the fibers; and the ash portion of the hydrocoUoid.
Said analysis gave the following results:
The sugar contents of the fibers are shown in Table II
Table II
D-galactose L-arabinose L-rhamnose D-xylose D-galacturonic aci 40 : 20 : 5 : 1 : 31
Said values are substantially within the limits of the results given by Wenzel e . al .
The analysis of the ash portion which had so far not been performed gave the results shown in Table III.
Table III
The presence of transition and heavy metals in the ash portion was also determined and it was found that they were present, if at all, in very small portions. Therefore, the presence of said transitional metals and/or heavy metals may be disregarded.
The present invention thus consists in a hydrocoUoid obtained by extraction from Portulaca Oleracea having the composition shown in Table I in which the sugar contents in the fibres are as shown in Table II and the composition of the ash portion is as given in Table III.
The specific characteristic features regarding the molecular
weight, the solubility in water, the viscosity, the surfac activity and the interface activity of the hydrocoUoid were als determined.
It has been found that the molecular weight of the fractio of the hydrocoUoid has a distribution as shown in Table IV.
Table IV
34% up to 104 daltons
12% between 104 ■ - 10b daltons
33% between 105 - - 107 daltons
11% between 107 - - 108 daltons
10% over 108 daltons
It has been found that the solubility of said hydrocoUoid i water is as shown in Table V.
Table V
It has been found that the viscosity of said hydrocoUoid i as shown in Table VI .
Table VI
Said data show that said hydrocolloids are not very viscous, which means that one may obtain a relatively concentrated solution which cannot be obtained with other hydrocolloids .
Moreover, said hydrocoUoid has a good surface tension. Thus, e.g it may reduce the surface tension of water in a concentration of 0.6% of the weight of the material, from 72 dyn/cm to 47 dyn/cm.
The interface tension between tetra-decane and water is in general 44 dyn/cm and in the presence of the hydrocoUoid according to the present invention it is reduced to 18 dyn/cm.
The ratio between the concentration of the hydrocoUoid and the surface tension and of the interface tension is shown in Table VII.
Table VII
All the above data regarding the molecular weight, the solubility in water, the viscosity, the surface tension and the interface tension of the hydrocoUoid according to the present invention have so far not been known.
It has surprisingly been found that the hydrocoUoid according to the present invention may be used, inter alia, in the following ways :
1. It may be part of an emulsion;
2. It can be part of a pharmaceutical preparation which may be used, e.g. for a. reducing the sugar level in the blood; and b. reducing the blood pressure.
The present invention thus consists also in emulsions comprising as active ingredient a hydrocoUoid according to the present invention.
A preferred emulsion is that of , e.g. a paraffin oil, such as tetra-decane, which is dispersed in an aqueous solution of the hydrocoUoid.
However, the present invention is not restricted to the above emulsions. Thus similar emulsions may be prepared with other oils, e.g. soya oil, olive oil, coconut oil, castor oil, tricaprylin oil, sunflower oil, etc.
A stable emulsion can also be obtained when toluene is the oil phase .
When the hydrocoUoid solution is further purified by centrifugation the fraction having a higher molecular weight is precipitated. The fraction remaining in the solution enables the preparation of emulsions which are stable for a long period having comprising drops having a much lower average size.
The lower the pH of the hydrocoUoid dispersion a more stable emulsion is being obtained having a lower average drop size (1-2 micrometers) .
The reduction of the ion strength also causes an increase of
the stability of the emulsion.
The hydrocoUoid dispersion which is prepared at room temperature also yields a more stable emulsion than those prepared at other temperatures .
The stability of the emulsion according to the present invention may be increased by the addition of another suitable emulsifier, e.g Tween 20; sorbitan monolaurate-20-ethoxylate ; sucrose esters; etc.
The present invention also consists in pharmaceutical preparations comprising as active ingredient a hydrocoUoid according to the present invention. Said pharmaceutical preparation is also used in particular for reducing the sugar content in the blood and/or for reducing the blood pressure.
The pharmaceutical composition may be an emulsion according to the present invention. However, it may be any suitable tablet, capsule, solution, etc. comprising any suitable conventional carrier, solvent, etc.
The amount of hydrocoUoid to be given to the patient is preferably 5 - 10 g in half glass of water or of yoghurt XA hour before the meal once or twice a day.
The present invention will now be illustrated with reference to the following Examples and accompanying Figs., without being restricted by them. Example 1
The plant Portulaca Oleracea, was crushed for 2-3 minutes in an electric blender in the presence of ethanol in a ratio of 1:1 (wet plant : ethanol) .
After crushing, the solid fraction was dried in an oven and extracted with acetone. The fraction rich with chlorophyll and
other dirty particles was deleted. Said fraction constitutes 0.5% of the weight of the dry plant. Thereafter, the remainder was extracted in a soxlet with a mixture of toluene : ethanol in a ratio of 1:2. The lipid fraction was thus removed. Said fraction comprises most of the lipids, the fats, the carotenoids, the phospholids and the wax. Said fraction constitutes 3% of the weight of the dry plant. Thereafter, the remainder was extracted with water. Said extraction may be performed at room temperature or at temperatures up to 100°C. The higher said temperature, the shorter the extraction time. In general, said extraction was performed in the course of 4 hours at 50°C.
The aqueous fraction was then centrifuged in order to remove the rest of the plant . Ethanol was then added to the aqueous fraction in a ratio of ethanol : aqueous solution 3 : 1. The water soluble hydrocoUoid was precipitated.
After drying in an oven and in vacuum at room temperature said hydrocoUoid constitutes about 3-5% of the weight of the plant. The product obtained is a white powder which is partly soluble in water. Some further part may be dispersed in water (see Table V) . Example 2
The hydrocoUoid was dissolved in water at room temperature with magnetic stirring overnight. The oil was dripped into the solution and the stirring was performed in a homogenizer (9500 rpm) . After 5 minutes all the oil had been dripped into the emulsion. Stirring was continued for another 10 minutes. Thereafter, if required, stirring is performed in a micro-fluidizer (3-4 turnovers) .
By the above method the following emulsions were prepared: a. 5% of tetra-decane was dropped into 95% aqueous solution
of 0.5% hydrocoUoid with homogenization by ultraturrax (9500 rpm) and followed by micro-fluidization (4 - 5 turnovers) . The emulsion obtained was stable more than a year. The size of the droplets was less than 5 μm. b. In a similar manner as described in a. above an emulsion of 20% tetra-decane and 80% of a 0.5% aqueous hydrocoUoid solution was prepared. The emulsion obtained was stable more than a fortnight . c. In a similar manner as described in a. above an emulsion of 30 tetra-decane and 70% of a 0.9 aqueous solution was prepared. The emulsion obtained was stable more than a fortnight. d. In a similar manner as described in a. above an emulsion of toluene (5%), of hydrocoUoid (0.5 %0 and water (94.5%). was prepared. The emulsion obtained was so stable and transparent that nearly no drops could be seen.
Example 3
A serie of emulsions having different pH of the aqueous phase was prepared in a similar manner as described in Example 2. The concentration of the hydrocoUoid was 0.5% of the emulsion. The oil was tetra-decane.
The hydrocoUoid was dissolved, in the course of the night, in water with magnetic stirring. The pH was adjusted to 1.7, 3. 6.6, 9 and 11 by the addition of HC1 or NaOH, respectively, without the addition of a buffer. The solution maintains the desired pH for an extended period of time. The oil was dripped into the solution with magnetic stirring.
The influence of the concentration of an electrolyte on the stability of the emulsion was also tested.
When the electrolyte was 0.015 M NaCl it was added to the
hydrocoUoid solution before the oil was added.
The results are shown in Table VIII and in annexed Fig. 1. ESI in accordance with the following table is: ESI = the ratio between the percentage of the oil drops (in accordance with the volume) in the range of 2- 10 μm a week after the day of preparation.
Table VIII
As can be seen the stability of the emulsion is improved when the pH is decreased. The most stable emulsion is obtained around pH 3. Example 4
The activity of the hydrocoUoid prepared as described in Example 2 after being subject to ultracentrifugation (32000 rotations/30 minutes was tested. The results are given in Table IX.
Table IX
It can be seen that the hydrocoUoid which was treated by ultracentrifugation was much more stable than the untreated
hydrocoUoid Example 5
In order to compare the activity of the hydrocoUoid obtained from Portulaca Oleracea, i.e. that according to the present invention with emulsions comprising hydrocolloids of other origins there were prepared, in the same manner as described in Example 2 (the time of homogenisation was reduced from 15 to 5 minutes) emulsions comprising 0.5% hydrocoUoid and 5% tetra-decane. The compared hydrocolloids were:
Arabic, Guar and Xanthan.
The results are shown in Table X.
ESI in accordance with the following table is:
% (of Volume) of drops 2 -10 μm
ESI = in a week
% (of Volume) of drops in the range of 2-10 μm at the day of preparation
Table X
As can be seen the hydrocoUoid according to the present invention gives the most stable emulsion in comparison with the other tested hydrocolloids. Example 6
A serie of emulsions was prepared changing the dissolving
temperature of the hydrocoUoid. The hydrocoUoid was at first dissolved over the night at room temperature. Thereafter it was heated for 10 minutes to 50°, 70° and 90°C, respectively. Thereafter the emulsion was cooled slowly to room temperature. Then the emulsion was prepared as described in Example 2.
The results are shown in Table XI and in Fig. 2.
Table XI
As can be seen the most stable emulsion is obtained when the hydrocoUoid is not heated before the emulsion is prepared. Example 7
A test was made to combine the hydrocoUoid according to the present invention with a monomeric emulsifier.
A serie of emulsions was prepared in which the hydrocoUoid and Tween 20 were prepared in 2 steps:
At the beginning a 0.3% emulsion was prepared in the following manner :
After it has been dissolved in the conventional manner, oil (tetra-decane) was dripped in, with homogenisation, into the solution. The homogenisation was performed for 15 minutes in a homogenisator. To the emulsion obtained was added Tween 20 in concentrations of 0.5 - 4%. The emulsion was stirred for 1 hour with a magnetic stirrer. Thereafter the Z-potential of the oil drops was measured. As the result of the measurement the composi-
tion of the drops was determined. With the increase of the concentration of the Tween 20 there occurs a gradual substitution of hydrocoUoid by the Tween 20. An absolute substitution occurs in the ratio of the weight concentrations
Hydrocolloid/Tween 20 = 0.3wt%/4 wt%. The concentration of Tween 20 required for the absolute substitution is relatively high to the proteins?, i.e. when the concentration of the Tween 20 is lower than 4% both emulsifiers are on the oil drops. Example 8.
For checking the change of the blood pressure 5 - 10 g of the hydrocoUoid in % a glass of water or yoghurt were given to 10 patients ^ hour before the meal. The tests were continued for a fortnight . The blood pressure was measured at the beginning of the test and at the end thereof. The blood pressure was reduced from 160 + 20 /110 + 20 to 125 + 20 / 90 + 20, respectively. Example 9
6.4 mg/ml of sugar were dissolved in a in vitro/in vivo test on one side of a rat intestine. On the other side of the intestine only water was present. When a hydrocoUoid emulsion was added to the sugar solution the amount thereof in the other side was only 1.7 mg/ml. Example 10
The contents of the sugar in the blood was tested. 5 - 10 g of the hydrocoUoid in % a glass of water or yoghurt in were given to 10 patients hour before the meal. The tests were continued for a fortnight . The sugar content was measured at the beginning of the test and at the end thereof . At the end of the test the amount of glucose in the blood for those patients being fed with the
hydrocoUoid was 135 mg glucose/dl blood, whereas the amount in the blood of untreated patients was 180 mg glucose/dl blood.