Finely powdered fibroin and process for producing same

Finely powdered high-purity fibroin and a process for producing the same are disclosed. The fibroin is a fine powder of fibroin in nonfibrous and particulate form which has an average molecular weight of not less than 50,000, particle diameters of from 1 to 100.mu., and a bulk density of from 0.1 to 0.7 g/cm.sup.3 as measured in the dry state and which contains at least 50% by weight of hot-water-insoluble fibroin having the .beta.-configuration. The process comprises dissolving a degummed silk material in an aqueous salt solution containing from 5 to 80% by weight of an alkali metal salt or alkaline earth metal salt; dialyzing the resulting aqueous fibroin solution; adding from 1 to 150 parts by weight of an alcohol to 100 parts by weight of the dialyzed aqueous fibroin solution having a fibroin concentration of from 3 to 20% by weight to form a gel of fibroin; dehydrating and drying the gel so formed; and then pulverizing the resulting powder. The fine powder of fibroin thus obtained is useful as an additive for cosmetic and pharmaceutical preparations.

BACKGROUND OF THE INVENTION 
This invention relates to finely powdered high-purity fibroin and a process 
for producing the same. 
Powdered silk fibroin is considered to be useful as an additive for 
cosmetic and pharmaceutical preparations, because of its moderate moisture 
absorption and retention properties and its high affinity for the human 
skin. Currently available silk fibroin powders are generally produced by 
finely dividing silk thread with pulverizer. Such a silk fibroin powder 
consists of filamentous fibers cut in very short lengths rather than 
nearly globular particles and, when used as an additive for cosmetic and 
pharmaceutical preparations, gives rise to various difficulties. For 
example, in mixing the powder with other ingredients in globular form, it 
is so liable to aggregation that a homogeneous final product is hardly 
obtained. Even if such a product is obtained, it shows poor slip 
properties upon application to the human skin and may occasionally produce 
round agglomerates of silk fibroin. Thus, it can be said that these 
difficulties prevent us from making good use of the excellent properties 
of silk fibroin. 
With this background, the present inventors have made repeated studies on 
the production of a homogeneous fine powder of fibroin in globular 
particulate form rather than in fibrous form. In this field of art, for 
example, a process for producing silk fibroin suitable for use in 
chromatography is disclosed in Japanese Patent Publication No. 1941/'64. 
This process comprises dissolving silk fibroin in a cuprammonium solution 
or a solution of a copper complex (for example, a cupri-ethylenediamine 
solution), neutralizing the resulting solution with an acid, and adding an 
alcohol to the neutralized solution to form a white precipitate of silk 
fibroin. As a result of confirmatory tests made by the present inventors, 
it has been found that this process requires a very large amount of 
alcohol and, moreover, the resulting precipitate is too sticky to be 
separated by filtration. Another process for producing a powder of silk 
fibroin is disclosed in Japanese Patent Publication No. 4947/'51. This 
process comprises dissolving degummed silk fiber in a concentrated aqueous 
solution of a neutral salt such as calcium nitrate, dialyzing the 
resulting solution, and spray-drying the colloidal solution so formed. 
However, the powder of silk fibroin thus obtained is abnormally 
hydrophilic and, therefore, unsuitable for use as an additive for cosmetic 
preparations. 
In order to overcome the above-described difficulties, the present 
inventors have made great efforts to produce an improved fine powder of 
fibroin in nonfibrous and particulate form, and have thereby completed 
this invention. 
BRIEF SUMMARY OF THE INVENTION 
It is an object of this invention to provide a fine powder of fibroin in 
nonfibrous and particulate form. Another object of this invention is to 
provide a process for producing a fine powder of fibroin in nonfibrous and 
particulate form which permits its industrial production with great ease 
and at low cost. 
In accordance with one aspect of this invention, there is provided a fine 
powder of regenerated fibroin in nonfibrous and particulate form which has 
an average molecular weight of not less than 50,000, particulate diameters 
of from 1 to 100.mu., and a bulk density of from 0.1 to 0.7 g/cm.sup.3 as 
measured in the dry state and which contains at least 50% by weight of 
hot-water-insoluble fibroin having the .beta.-configuration. 
In accordance with another aspect of this invention, there is provided a 
process for producing such a fine powder of regenerated fibroin which 
comprises the steps of dissolving a degummed silk material in an aqueous 
salt solution containing from 5 to 80% by weight of an alkali metal salt 
or alkaline earth metal salt; dialyzing the resulting aqueous fibroin 
solution; adding from 1 to 150 parts by weight of an alcohol to 100 parts 
by weight of the dialyzed aqueous fibroin solution having a fibroin 
concentration of from 3 to 20% by weight to form a gel of fibroin; 
dehydrating and drying the gel so formed; and then pulverizing the 
resulting powder. 
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS 
The silk material which is used in the process of this invention can be 
cocoons, raw silk, waste cocoons, raw silk waste, bisu (unreelable 
cocoons), silk fabric waste, bourette, and the like. Prior to use, the 
silk material is degummed or freed from sericin by any conventional 
procedure. For example, it is washed in warm water containing a 
surface-active agent or any enzyme according to the need, and then dried. 
Using a suitable apparatus such as kneader, the degummed silk material is 
dissolved in a solvent preheated to a temperature of from 60.degree. to 
95.degree. C. and preferably from 70.degree. to 85.degree. C. The solvent 
is composed of an aqueous salt solution containing from 5 to 80% by weight 
of an alkali metal salt or alkaine earth metal salt, and used in an amount 
of from 2 to 50 parts by weight and preferably from 3 to 30 parts by 
weight per part by weight of the degummed silk material. The alkali metal 
salts and alkaline earth metal salts which can be used in the process of 
this invention include LiCl, LiBr, NaI, LiNO.sub.3, MgCl.sub.2, 
MgBr.sub.2, Mg(NO.sub.3).sub.2, ZnCl.sub.2, Zn(NO.sub.3).sub.2, and the 
like. However, CaCl.sub.2 and Ca(NO.sub.3).sub.2 are preferred because 
they permit the solubility and molecular weight of fibroin to be kept as 
high as possible. In the aforesaid aqueous salt solution, the 
concentration of the metal salt is generally from 5 to 80% by weight, 
preferably from 20 to 70% by weight, and most preferably from 40 to 60% by 
weight. If the concentration of the metal salt is less than 5% by weight, 
the rate of dissolution is low, while if it is greater than 80% by weight, 
the resulting fine powder of fibroin shows a reduction in molecular 
weight. It is preferable to add an alcohol to the aqueous salt solution 
for the purpose of further improving its dissolving powder. This alcohol 
may be added either before or during the dissolution of the degummed silk 
material, and the amount of alcohol added is generally from 20 to 60% by 
weight and preferably from 25 to 50% by weight. 
From the resulting aqueous fibroin solution, the salt contained therein is 
almost completely removed by means of a dialyzer using semipermeable 
membranes or hollow fibers, typically made of cellophane. In order that a 
gel of fibroin may be formed stably and rapidly, there must be a proper 
correlation between the volume of the solution to be dialyzed and the 
surface area of the dialysis membrane. More specifically, desalting should 
be carried out by the use of a multilayer membrane structure or bundled 
hollow-fiber structure satisfying the condition expressed by 
##EQU1## 
where the priming volume means the internal volume within the tubing or 
between the layers. If the value of the above-defined ratio is less than 
10, the removal of the salt through the membrane is not effected rapidly 
and, moreover, not a stable gel of fibroin but only a sticky precipitate 
is formed in the succeeding gelation step. In order to carry out the 
process of this invention smoothly and economically, the above-defined 
ratio should preferably have a value of not less than 30 and most 
preferably a value of not less than 50. In the case of a multilayer 
membrane structure, for example, it is necessary to keep the spacing 
between layers at 2 mm or less for the purpose of satisfying the aforesaid 
condition. In the case of a bundled hollow-fiber structure which is more 
suited to the satisfaction of the aforesaid condition, it is necessary to 
use hollow fibers having a diameter of 4 mm or less. 
In the process of this invention, the dialyzed aqueous fibroin solution has 
a very low residual salt concentration of from 0.003 to 0.06% by weight, 
so that an extremely high purity of fibroin can be achieved. 
Then, the dialyzed aqueous fibroin solution is transferred to the gelation 
step. First of all, the aqueous fibroin solution is adjusted to a fibroin 
concentration of from 3 to 20% by weight, preferably from 4 to 15% by 
weight, and most preferably from 5 to 10% by weight. If the fibroin 
concentration is less than 3% by weight, a homogeneous mass of gel is not 
formed and, upon addition of a large amount of alcohol, only a sticky 
precipitate is formed. On the other hand, if it is greater than 20% by 
weight, a stable mass of gel is formed but its dehydration becomes very 
difficult. Subsequently, the gelation step is carried out. In accordance 
with this invention, a homogeneous mass of gel can be formed by the 
addition of an alcohol to the aqueous fibroin solution having an 
appropriate fibroin concentration within the above-defined range. The 
amount of alcohol added should generally be from 1 to 150% by weight, 
preferably from 5 to 80% by weight, and most preferably from 10 to 60% by 
weight based on the weight of the aqueous fibroin solution. If the amount 
of alcohol added is less than 1% by weight, no gel is formed and, even if 
it is formed, a very long time is required. On the other hand, if it is 
greater than 150% by weight, a sticky precipitate rather than a stable 
mass of gel is formed and its dehydration becomes very difficult. There is 
a correlation between the amount of alcohol added and the fibroin 
concentration of the aqueous fibroin solution. More specifically, the 
product of the fibroin concentration (% by weight) and the amount of 
alcohol added (% by weight) should generally have a value of from 10 to 
1,000 and preferably from 15 to 500. The alcohol may be added to the 
aqueous fibroin solution, and vice versa. In either case, a homogeneous 
and stable mass of gel is usually formed in an instant or within several 
hours. 
The alcohols which can be used in the process of this invention include 
monohydric aliphatic alcohols such as methyl alcohol, ethyl alcohol, 
propyl alcohol, isopropyl alcohol, butyl alcohol, pentyl alcohol, octyl 
alcohol, etc.; dihydric aliphatic alcohols such as ethylene glycol, 
diethylene glycol, triethylene glycol, etc.; and trihydric alcohols such 
as glycerol. However, methyl alcohol, ethyl alcohol, and isopropyl alcohol 
are preferred because they permit a stable mass of gel to be formed 
easily. 
The mass of gel so formed is subjected to the dehydration step. This step 
is preferably carried out by the use of a centrifuge, and the stable mass 
of gel formed in accordance with this invention is generally dehydrated to 
a water content of the order of from 100 to 500% by weight based on the 
weight of the solid contained therein. During the dehydration step carried 
out by centrifugation, the mass of gel is broken to fragments of small 
sizes which can then be easily dried to the absolute dry state. This 
drying step is carried out at a temperature of from 60.degree. to 
120.degree. C. under normal or reduced pressure. 
The powder of fibroin thus obtained is subsequently pulverized by the use 
of a pulverizer such as pin mill or jet mill. The practice diameters 
should be adjusted to a range of from 1 to 100.mu., preferably from 4 to 
80.mu., and most preferably from 5 to 30.mu.. If the particle diameters 
are less than 1.mu., the resulting fine powder shows poor dispersibility 
and compatibility when used as an additive for cosmetic preparations, 
while if they are greater than 100.mu., the resulting fine powder has low 
affinity for the skin and poor slip properties on the skin. Since the 
present process for producing a fine powder of fibroin involves gelation 
followed by dehydration and drying, the resulting fibroin particles are 
considered to have very minute pores to which their good moisture 
absorption and retention properties are attributable. However, this may 
lead to the disadvantage that the fine powder of fibroin becomes 
excessively swollen in certain applications. If is desirable, therefore, 
to subject the resulting fine powder of fibroin to a wet heat treatment 
comprising exposure to saturated steam at a temperature of 50.degree. C. 
or above and preferably from 80.degree. to 120.degree. C. This treatment 
may be applied either to the powder ensuing from the dehydration and 
drying step or to the fine powder ensuing from the pulverization step. As 
a result, the resulting fine powder of fibroin is increased in the degree 
of crystallinity and the content of hot-water-insoluble fibroin. 
The fine powder of fibroin produced in accordance with this invention has a 
degree of crystallinity of not less than 20%, preferably not less than 
30%, and most preferably not less than 40%. The degree of crystallinity 
was determined in the following manner: An aqueous fibroin solution 
containing 5% by weight of a fibroin produced in accordance with this 
invention was poured on a Teflon plate and then dried at a temperature of 
50.degree. C. to form a fibroin film having a thickness of about 60.mu.. 
This fibroin film was regarded as amorphous (0%) while raw silk is 
completely crystalline (100%). The degree of crystallinity was expressed 
as a relative value on the scale defined between these standard points. 
The fine powder of fibroin produced in accordance with this invention has a 
bulk density of from 0.1 to 0.7 g/cm.sup.3 and preferably from 0.2 to 0.6 
g/cm.sup.3 as measured in the dry state. If the bulk density is less than 
0.1 g/cm.sup.3, the fine powder of fibroin has poor compatibility and 
dispersibility and, when used as an additive for cosmetic preparations, 
may produce a phase separation. If it is greater than 0.7 g/cm.sup.3, the 
fine powder of fibroin is decreased in moisture absorption and retention 
properties. The bulk density was measured in the most closely packed state 
by means of a commercially available powder tester (manufactured and sold 
by Hosokawa Tekkosho, Ltd.). 
The fine powder of fibroin produced in accordance with this invention 
contains at least 50% by weight, preferably at least 70% by weight, and 
most preferably at least 90% by weight of hot-water-insoluble fibroin 
having the .beta.-configuration. If the content of hot-water-insoluble 
fibroin is less than 50% by weight, the fine powder is extremely 
hydrophilic and liable to deterioration. Moreover, when used as a base 
material for cosmetic preparations, it shows a high degree of stickiness 
and gives a disagreeable feeling to the skin. The content of 
hot-water-insoluble fibroin (or rate of .beta.-configuration) was 
determined in the following manner: Ten g (absolute dry weight) of a fine 
powder of fibroin to be tested was boiled in 1 l of hot water at a 
temperature of 100.degree. C. for a period of 15 minutes, and the 
undissolved fraction of fibroin was absolutely dried and weighed. Then, 
the content of hot-water-insoluble fibroin was calculated from the 
equation: 
EQU Content of Hot-water-insoluble Fibroin =W/10.times.100 (% by weight) 
where W stands for the absolute dry weight (in g) of the undissolved 
fraction of fibroin. 
The fine powder of fibroin produced in accordance with this invention has a 
high purity as well as good moisture absorption and retention properties. 
Accordingly, it is very useful as an additive for cosmetic and 
pharmaceutical preparations. It is also suitable for use as an adsorbent 
in pharmaceutical and hygienic applications, because the fibroin particles 
contained therein have very minute pores owing to the special manner of 
production.

The present invention is further illustrated by the following examples. 
EXAMPLE 1 
In this example, raw silk waste was used as the starting material for the 
production of fine powders of fibroin. Three kg of raw silk waste was 
immersed in 100 l of water containing 0.3% by weight of marseille soap, 
stirred at 80.degree. C. for 1 hour, treated with a sericinolytic enzyme 
to remove the sericin almost completely, washed with water, and then 
dried. 
By stirring in a kneader, 1-kg portions of raw silk waste degummed as above 
were dissolved in 10 kg each of aqueous calcium chloride solutions having 
the respective calcium chloride concentrations indicated in Table 1. As 
can be seen from the data of this table, the process of this invention 
allowed the raw silk waste to be dissolved easily. However, when the 
calcium chloride concentration was less than 5% by weight as in Control 
Runs 1-(1) and 1-(2), it was hardly dissolved even after a long period of 
time (24 hours or more). 
The resulting aqueous fibroin solutions were then desalted by passing each 
of them through a dialyzer of the hollow-fiber type at a rate of 1 l/hr. 
This dialyzer was composed of 2,000 hollow fibers of regenerated cellulose 
having an internal diameter of 200.mu., a membrane thickness of 20.mu., 
and a length of 500 mm, both ends of these hollow fibers being bundled and 
sealed without blocking up their hollow bores. In this case, the ratio of 
membrane surface area (cm.sup.2) to priming volume (cm.sup.3) has a value 
of 100. After completion of the dialysis, the aqueous fibroin solutions 
had fibroin concentrations of 5.3-6.7% by weight and residual calcium 
chloride concentrations of 0.007-0.033% by weight. 
The molecular weight of the fibroin contained in each of the dialyzed 
aqueous fibroin solutions was measured by gel permeation chromatography. 
When the calcium chloride concentration was greater than 80% by weight as 
in Control Run 1-(15), the molecular weight was reduced to the order of 
40,000. In the aqueous fibroin solutions prepared in accordance with this 
invention, however, the fibroin contained therein had a molecular weight 
of not less than 50,000 and showed no appreciable degree of hydrolysis. 
TABLE 1 
__________________________________________________________________________ 
Calcium Amount of 
Chloride 
Ethyl Alcohol Molecular 
Concentration* 
Added** Temperature 
Time Weight 
(wt. %) (wt. %) (.degree.C.) 
(hr) 
Solubility*** 
(.times. 10.sup.4) 
__________________________________________________________________________ 
1-(1) 
Control Run 
3 0 95 24 X -- 
1-(2) 
" 3 30 85 24 X -- 
1-(3) 
Test Run 
5 30 85 5 .DELTA. 
8 
1-(4) 
" 10 30 80 5 .DELTA. 
9 
1-(5) 
" 30 30 80 2 O 9 
1-(6) 
" 40 0 95 2 O 7.5 
1-(7) 
" 40 30 80 2 .circleincircle. 
9.5 
1-(8) 
" 50 0 95 1 O 8 
1-(9) 
" 50 30 80 1 .circleincircle. 
10 
1-(10) 
" 50 50 95 1 .circleincircle. 
10 
1-(11) 
" 60 0 80 1 O 8.5 
1-(12) 
" 60 30 80 1 .circleincircle. 
11 
1-(13) 
" 60 30 70 1 .circleincircle. 
11 
1-(14) 
" 80 50 80 1 .circleincircle. 
5.5 
1-(15) 
Control Run 
90 60 80 1 .circleincircle. 
4 
__________________________________________________________________________ 
*Expressed as percentages based on the weight of the aqueous calcium 
chloride solution, exclusive of ethyl alcohol. 
**Expressed as percentages based on the weight of the aqueous calcium 
chloride solution. 
***Rated as insoluble (X), sparingly soluble (.DELTA.), soluble (O), or 
very soluble (.circleincircle.). 
EXAMPLE 2 
The procedure of Test Run 1-(12) in Example 1 was repeated except that the 
calcium chloride was replaced by calcium nitrate. After dialysis, the 
resulting aqueous fibroin solution had a fibroin concentration of 6.2% by 
weight and a residual calcium nitrate concentration of 0.015% by weight. 
This aqueous fibroin solution was either concentrated or diluted with 
water to prepare a series of aqueous fibroin solutions having the 
respective fibroin concentrations indicated in Table 2. 
An alcohol was added to each of the above aqueous fibroin solutions and the 
form of the resulting gel was observed. When the fibroin concentration was 
less than 3% by weight as in Control Runs 2-(1) and 2-(2) or when the 
amount of alcohol added was greater than 150% by weight of the aqueous 
fibroin solution as in Control Runs 2-(6), 2-(14) and 2-(18), not a 
homogeneous mass of gel but a white precipitate was formed. This 
precipitate was too sticky to be separated by conventional filtration 
under reduced pressure. When it was placed in a cloth bag and then 
centrifuged, it aggregated into a bulky and sticky mass which was very 
difficult of dehydration and drying. On the other hand, when the amount of 
alcohol added was less than 1 by weight as in Control Runs 2-(8) and 
2-(15), no gelation was recognized even after the mixture was allowed to 
stand for a whole day and night. Furthermore, when the fibroin 
concentration was greater than 20% by weight as in Control Run 2-(22), the 
resulting mass of gel was so tough that it could hardly be dehydrated by 
centrifugation. 
However, the process of this invention allowed a homogeneous mass of gel to 
be formed in an instant or within several hours after the addition of an 
alcohol. When dehydrated by centrifugation, the mass of gel was broken to 
fragments having sizes of the order of several millimeters and water 
contents of 110-480% by weight. Methyl alcohol, ethyl alcohol, and 
isopropyl alcohol made no substantial differences in the form of the 
resulting gel and the requirements for further treatment. The dehydrated 
gel was dried in a hot-air oven set at 90.degree.-100.degree. C. or a 
vacuum dryer set at 70.degree. C. to obtain a fine granular product of 
fibroin suitable for direct pulverization with a jet mill. Subsequently, 
the granular product was subjected to a wet heat treatment comprising 
exposure to saturated steam at 120.degree. C. for 20 minutes, and then 
pulverized with a jet mill to obtain a fine powder of fibroin consisting 
of nearly globular particles, 98% or more of which had diameters of 
5-40.mu.. 
When measurements of the bulk density were made, all the fine powders of 
fibroin, except that produced in Control Run 2-(22), were found to have 
values within the range of 0.1-0.7 g/cm.sup.3. The reason for this seems 
to be that the fibroin particles contained therein had minute pores owing 
to the special manner of production in which a powder is derived from a 
homogeneous mass of gel. 
In all the fine powders of fibroin produced in accordance with this 
invention, the content of hot-water-insoluble fibroin or rate of 
.beta.-configuration was found to be not less than 70% by weight and the 
degree of molecular orientation was found to be not greater than one-half 
that of natural silk. 
Furthermore, measurements of the rate of moisture absorption were made. 
More specifically, a sample of each fine powder of fibroin was allowed to 
stand in an atmosphere having a temperature of 20.degree. C. and a 
relative humidity of 65%, and the amount of water absorbed was measured 
after 2 hours. For purposes of comparison, a fine powder of fibroin 
produced by finely dividing silk yarn with a pulverizer had a rate of 
moisture absorption of 2-3% by weight as measured by the same method. 
The results of these measurements are given in Table 2. 
3 TABLE 2 
Dehydration by Fibroin Addition of Centrifugation Rate of 
Content of Degree of Concent- Alcohol Gelation Water Degree of Bulk 
Moisture Hot-Water- Crystallin- ration Amount Time Content** Dehydratio 
n*** Density Absorption insoluble ity (wt. %) Type* (wt. %) (min.) Form 
of Gel (wt. %) (wt. %) (g/cm.sup.3) (wt. %) Fibroin (%) (%) 
2-(1) Control Run 2 E 10 -- Precipitate -- -- -- -- 2 0 2-(2) " 2 E 
50 -- " -- -- -- -- 7 0 2-(3) Test Run 3 E 10 180 Soft mass 480 87 0.59 
4.2 76 43 2-(4) " 5 E 10 180"450 79 0.44 5.5 79 46 2-(5) " 5 E 50 5 " 
320 96 0.27 6.1 98 68 2-(6) Control Run 5 E 180 -- Precipitate -- -- -- 
-- 10 1 2-(7) Test Run 5 M 20 20 Soft mass 400 83 0.41 5.7 93 62 2-(8) 
Control Run 10 E 0.5 1,440 No gelation -- -- -- -- -- -- 2-(9) Test Run 
10 E 2 300 Somewhat hard mass 410 55 0.54 4.5 74 43 2-(10) " 10 E 10 180 
" 270 73 0.39 5.3 88 48 2-(11) " 10 E 20 20 " 150 86 0.44 5.1 90 60 
2-(12) " 10 E 50 5 " 110 92 0.21 6.2 98 65 2-(13) " 10 E 150 1 Soft mass 
180 92 0.50 4.8 83 42 2-(14) Control Run 10 E 200 -- Precipitate -- -- 
-- -- 17 3 2-(15) " 10 M 0.3 -- No gelation -- -- -- -- -- -- 2-(16) 
Test Run 10 M 20 20 Somewhat hard mass 200 82 0.45 5.2 85 48 2-(17) " 10 
P 20 20 " 210 81 0.45 5.1 77 43 2-(18) Control Run 10 P 200 -- Precipitat 
e -- -- -- -- 30 7 2-(19) Test Run 15 E 20 15 Somewhat hard mass 200 71 
0.41 5.0 81 48 2-(20) " 15 M 20 15 " 190 73 0.42 5.0 81 46 2-(21) " 20 E 
20 15 " 260 48 0.62 3.7 70 41 2-(22) Control Run 25 E 20 15 Hard mass -- 
-- 0.76 2.9 38 12 
*E stands for ethyl alcohol, M for methyl alcohol, and P for isopropyl 
alcohol. 
**Defined as the solvent content of the gel. 
***Defined as (the amount of solvent removed by centrifugation)/(the 
amount of solvent used during geletion) .times. 100. 
EXAMPLE 3 
One kg of spun silk waste (bourette) was immersed in 30 l of water 
containing 0.5% by weight of marseilles soap, stirred at 100.degree. C. 
for 1 hour to remove the sericin and oily matter almost completely, washed 
thoroughly with water, and then dried at 70.degree. C. In a kneader having 
a capacity of 50 l, a solution of 7.5 kg of calcium chloride in 5 kg of 
water and 4 kg of ethyl alcohol was prepared, and 3 kg of bourette 
degummed as above was dissolved therein by stirring at 80.degree. C. for 1 
hour. After completion of the dissolution, 9 kg of water preheated to 
80.degree. C. was added. The resulting fibroin solution was filtered to 
remove any insoluble foreign matter such as chrysalis refuse, and then 
desalted by passing it through the same dialyzer of the hollow-fiber type 
as used in Example 1. The aqueous fibroin solution thus obtained had a 
fibroin concentration of 6.2% by weight and a residual calcium chloride 
concentration of 0.021% by weight. 
When 10 kg of ethyl alcohol was added, with stirring, to 40 kg of an 
aqueous fibroin solution prepared as above and the resulting mixture was 
allowed to stand, a homogeneous mass of gel was formed in 20 minutes. This 
mass of gel was placed in a bag made of polyester fabric and then 
centrifuged to obtain fragments of gel having water contents of 250-350% 
by weight. After the dehydrated gel was dried in a vacuum dryer set at 
80.degree. C., a part of the resulting granular product was directly 
pulverized with a jet mill and the rest was subjected to a wet heat 
treatment comprising exposure to saturated steam at 100.degree. C. for 10 
minutes and then pulverized with a jet mill. Thus, a variety of fine 
powders of fibroin having the respective particle diameter ranges 
indicated in Table 3 were obtained. 
Using a mixer, each of the above fine powders of fibroin was blended with 
an equal weight of a commercially available fine powder of talc, and the 
compatibility and dispersibility of the resulting blend were examined 
under the microscope. It was found that its dispersibility became poor 
when the diameters of fibroin particles were excessively large or small. 
Especially when the fine powder of fibroin had particle diameters of less 
than 1.mu. as in Control Run 3-(1) or when it had particle diameters of 
greater than 100.mu. as in Control Runs 3-(12) and 3-(13), the fibroin and 
the talc tended to aggregate separately in the form of spots. When the 
diameters of fibroin particles were within the range of 1-100.mu. and 
particularly 4-80.mu. as taught by this invention, the fibroin and the 
talc showed very good compatibility and dispersibility. In addition, a 
sample of each fine powder of fibroin was placed on a hand and rubbed with 
fingers. As a result, the fine powders of fibroin produced in accordance 
with this invention spread uniformly over the skin without any appreciable 
degree of agglomeration and showed good slip properties. Especially the 
fine powders of fibroin subjected to a wet heat treatment were superior in 
slip and non-agglomeration properties to those subjected to no wet heat 
treatment. This is presumed to be due to the enhancement of crystallinity 
of fibroin caused by the wet heat treatment. In fact, when the fine 
powders of fibroin produced in Test Runs 3-(4) and 3-(5) were examined by 
X-ray diffraction analysis, the degree of crystallinity was 28% in the 
fine powder of fibroin subjected to no wet heat treatment and 45% in that 
subjected to a wet heat treatment. Moreover, the content of 
hot-water-insoluble fibroin or rate of .beta.-configuration was 52% by 
weight in the fine powder of fibroin subjected to no wet heat treatment 
and 98% by weight in that subjected to a wet heat treatment. However, the 
fine powder of fibroin produced in Control Run 3-(1), which had particle 
diameters of less than 1.mu., formed round agglomerates when placed on the 
skin and rubbed with fingers. On the other hand, the fine powders of 
fibroin produced in Control Runs 3-(12) and 3-(13), which had particle 
diameters of greater than 100.mu., felt rough and showed very poor slip 
properties. 
TABLE 3 
__________________________________________________________________________ 
Particle 
Wet Heat Dispersi- 
Range 
Treatment 
Bulk bility 
Diameter 
at 110.degree. C. 
Density 
with Slip on 
(.mu.) 
for 10 min. 
(g/cm.sup.3) 
Talc* 
the Skin* 
__________________________________________________________________________ 
3-(1) 
Control Run 
&lt;1 Done 0.16 X X (Agglome- 
ration) 
3-(2) 
Test Run 
1-10 " 0.22 .DELTA. 
.DELTA. 
3-(3) 
" 4-25 " 0.29 .circleincircle. 
.circleincircle. 
3-(4) 
" 5-30 " 0.44 .circleincircle. 
.circleincircle. 
3-(5) 
" 5-30 Not done 
0.37 .circleincircle. 
.circleincircle. 
3-(6) 
" 10-50 
Done 0.51 .circleincircle. 
.circleincircle. 
3-(7) 
" 10-50 
Not done 
0.48 O O 
3-(8) 
" 25-80 
Done 0.55 .circleincircle. 
O 
3-(9) 
" 5-100 
" 0.53 O .DELTA. 
3-(10) 
" 5-100 
Not done 
0.50 .DELTA. 
.DELTA. 
3-(11) 
" 50-100 
Done 0.64 .DELTA. 
.DELTA. 
3-(12) 
Control Run 
70-150 
" 0.78 X X 
3-(13) 
" &gt;100 " 0.83 X X 
__________________________________________________________________________ 
*Rated as very good (.circleincircle. ), good (O), inadequate (.DELTA.), 
or poor (X).