METHOD FOR TREATMENT OF MULBERRY LEAVES AND FOR ANTI-BACTERIAL SILK PRODUCTION

Abstract

Method for treatment of mulberry leaves for feeding silkworms in order to obtain a feedstuff for silkworms, doped with at least a metal selected among silver and/or zinc, comprising the steps of: a. preparing a solution comprising water, a photo-reducer agent and a precursor of said at least metal; b. treating said mulberry leaves with said solution; c. exposing said treated mulberry leaves to UV radiation, so that the photo-reduction of said precursor of said at least metal occurs, with the consequent deposition on said leaves of nanoparticles of said at least metal.

Claims

1. A method for treatment of mulberry leaves for feeding silkworms in order to obtain a feedstuff for silkworms, doped with at least one metal selected among silver and/or zinc, comprising the steps of: a) preparing a solution comprising: water; a photo-reducer agent; a precursor of said at least one metal; b) treating said mulberry leaves with said solution; c) exposing said treated mulberry leaves to UV radiation, so that the photo-reduction of said precursor of said at least one metal occurs, with the consequent deposition on said leaves of nanoparticles of said at least one metal.

2. The method according to claim 1, further comprising, prior to step b), the step of chopping said mulberry leaves and additivating them with nutrient substances in order to obtain a feedstuff for silkworms, and characterized in that said feedstuff, after carrying out steps v) and c), can be stored and used for breeding desasonalization of silkworms.

3. The method according to claim 1, characterized in that said at least one metal is silver, said photo-reducer agent is an alcohol, and said precursor is a silver salt.

4. The method according to claim 3, characterized in that said alcohol is methyl alcohol, and its concentration in said solution is between 1 and 20% and in that said silver salt is silver nitrate and its concentration in said solution is between 0.1 and 1.1%.

5. The method according to claim 1, characterized in that said treatment at step b) is carried out by means of spray coating or by means of dip coating and in that the quantity of solution used for said treatment is between 25 and 100 g per square meter of mulberry leaves.

6. The method according to claim 1, characterized in that said exposition to UV radiation at step c) occurs by means of exposition to solar light for a time interval between 4 and 8 hours.

7. The method according to claim 1, characterized in that said exposure to UV radiation of point c) occurs by means of exposure to radiation produced by a lamp with emission peak between 350 and 400 nm, arranged at such a distance that the power is between 5 W/m.sup.2 and 100 W/m.sup.2 for a time interval between 1 and 30 minutes.

8. The method according to claim 3, further comprising, after step c), the step of chopping said mulberry leaves treated with said at least one metal to obtain feedstuff for silkworms.

9. The method according to claim 1, characterized in that said treatment occurs by spraying said solution on mulberry leaves on the tree before they are picked.

10. The method according to claim 1, characterized in that said at least one metal comprises both zinc and silver.

11. A method for production of intrinsically antibacterial silk, doped with silver and/or zinc, comprising the steps of: feeding the silkworms up to when they climb to the wood on said mulberry leaves treated with the method according to claim 1; and picking cocoons produced by the silkworms and processing them for obtaining zinc and/or silver doped silk.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0017] As preliminary remarks to the detailed description of the invention it is convenient to precise that the biologic cycle of the silkworm begins in the egg phase and it is articulated in about four weeks up to the production of the cocoon. The growth phases of the silkworm are called larval instars with molts occurring at intervals during which the exoskeleton of the worm grows; the cocoon is completed at the end of the 5 th larval instar. After about 12-15 days there occur the moth emerging, the coupling and the deposition of new eggs. During the development, the food requirements of the silkworm increase and the silkworm needs always more quantity of food.

[0018] It is convenient to note that in addition to the mulberry leaves, the silkworms can be fed also on artificial feedstuff in order to obviate the seasoning of the mulberry and to allow the sericulture also during the dormant season of the plant.

[0019] The feedstuff obtained from mulberry leaves dried and additivated with other nutrient substances such as flours of various kind, vitamins etc. in powder form can be treated with silver and/or zinc by using the same technologies proposed in the present invention for the treatment of new mulberry leaves. Each strategy can be evaluated according to the results expected in the silkworms by suitably managing the process variables.

[0020] For example it is possible to define the concentration of silver to be used for the treatment of leaves according to the antibacterial level desired in the silk yarn produced by the silkworm by increasing the concentration of the silver precursor to obtain silk yarns with more efficient antibacterial properties.

[0021] Embodiments of the method with silver nanoparticles The first step of the method provides the treatment of mulberry leaves (or feedstuff obtained therewith) with silver nanoparticles.

[0022] For realizing this step there can be used solutions containing various silver precursors. The method of deposition provides the step of impregnation by means of solutions of silver precursor by using the dip coating or spray coating techniques and the next exposition to ultraviolet light.

[0023] The next step of the process provides the silkworm feeding, and also in this case various methods can be used. Since the growth of the silkworm develops in five larval instars to each one of which a different quantity of food to be administered corresponds, it is possible to differentiate the silkworm feeding according to their growth phases. For example, it is possible to feed the silkworms on silver- and/or zinc-treated leaves or as an alternative on silver- and/or zinc-treated feedstuff since the first instar so that the silkworm gets used to the food administered since the first phases of life; on the other hand, it is possible to choose to feed the silkworms on silver and/or zinc treated food only during the 5 th instar after which the silkworm will begin the production of the cocoon; as an alternative it is possible to feed the silkworm on silver and/or zinc treated food between the 1 st and the 5 th instars. The choice to use treated food only during the 5 th instar is the most economically advantageous since it allows to limit the volume of food to be treated with silver and/or zinc to the needs of the silkworm in the last instar, thus leading anyway the risk of a lower intake of food by the worm due to the modification of its diet.

[0024] The silk yarn obtained from the silkworms subjected to the modified diet has the antibacterial properties and can be further worked thanks to the use of techniques commonly used in sericulture.

[0025] In the specific case of biomedical application, from such silk it is possible to extract fibroin and sericin for the production of devices with both antibacterial and regeneration properties.

[0026] In particular, such process of extraction of fibroin and sericin can be carried out by means of degumming, during which the cocoons are immersed in hot water with possible addition of chemical substances with the aim of separating the sericin, protein of the silk with adhesive function between the fibres. In the following, fibroin and sericin are processed by means of suitable working techniques in order to produce biomaterials with intrinsic regeneration and antibacterial properties.

[0027] The properties of silk fibres produced by the silkworm reflect both the process parameters adopted for silver and/or zinc treatment of leaves and/or feedstuff (first step of the method) and the feeding method of the silkworms (second step).

[0028] In particular, the feeding method (in terms of duration and choice of instars at which the silkworm have to be fed) and the percentage of silver and/or zinc to be used for the treatment of leaves and/or feedstuff influence the concentration of silver and/or zinc present in the cocoon.

[0029] Embodiments of the method with nanoparticles of zinc or of zinc and silver

[0030] As yet said, yarns and fabrics treated with zinc or zinc and silver have efficient antibacterial properties as well.

[0031] Therefore it is possible to use methods similar to the ones which will be described for silver, to obtain feedstuff doped with zinc or zinc and silver. Therefore the method according to the present invention which uses zinc particles (alone or in association with silver particles) follows the same steps as yet described for the method which uses silver nanoparticles.

PREFERRED EMBODIMENTS

Example No 1Treatment of Mulberry Leaves by Means of Photo-Chemical Deposition of Silver Particles

[0032] According to a preferred embodiment, the treatment with silver of mulberry leaves and/or feedstuff is carried out by using a hydro-alcoholic solution of a silver precursor, preferably silver nitrate.

[0033] Leaves and/or feedstuff are impregnated with such solution, preferably using the spray coating technique, and in the following exposed to a UV lamp or as an alternative to solar light for inducing the photo-reduction reaction in order to convert the silver precursor in metal silver particles. Finally the treated leaves are washed and stored in refrigerator, preferably at a temperature of about 4 C. In case of new leaves only, these last ones can be stored in refrigerator for five days maximum up to their consumption. In case of leaves treated with silver directly on the tree, these maintain the antibacterial properties in the time and can be picked also after various weeks after treatment.

[0034] In the following the leaves and/or feedstuff treated with silver are used for feeding the silkworms and at the same time, as control, the silkworms are fed on not treated leaves or feedstuff. Both the families of silkworms are bred in the same conditions up to the time of climbing to the wood, after which the silkworms produce the cocoon. The cocoon are picked and processed according to the traditional procedures dictated by the tradition of sericulture.

[0035] In the following there are described in detail some preferred embodiments to obtain intrinsically antibacterial silk thanks to the use of suitable techniques of treatment with silver and/or zinc of leaves and/or feedstuff:

1. Preparation of the Solution Containing the Silver Precursor

[0036] A hydro-alcoholic solution containing the silver precursor, preferably silver nitrate, is prepared with a concentration between 0.1 and 1%, preferably 0.5% AgNO.sub.3. The solution contains also the photo-reducer agent, preferably an alcohol and preferably methyl alcohol CH.sub.3OH, in quantity between 1 and 20%, preferably 10% CH.sub.3OH. The use of water in the solution is provided as solvent.

[0037] The reagents are mixed at ambient temperature using a suitable mixing technique for obtaining the complete dissolving of salt, preferably by magnetic stirring for a time interval between 5 and 15 minutes, preferably 10 minutes.

[0038] The mixing step occurs in a non-transparent and suitably sealed vessel to avoid the interaction of the solution with solar light and the alcohol evaporation, respectively.

2. Deposition of the Solution Containing the Silver Precursor on Mulberry Leaves

[0039] Mulberry leaves are arranged on a working table avoiding that they overlap to each other and in the following they are treated with the silver based solution by using the a spray coating system, i.e. the spraying of the solution. The quantity of sprayed solution is between 25 and 100 g/m.sup.2, and preferably 50 g/m.sup.2.

3. Exposition to Ultraviolet Light of Mulberry Leaves

[0040] Such step can be carried out according to the two following methods:

a) immediately after the spray coating, the mulberry leaves are exposed to UV radiation, by using a lamp with an emission peak at wavelength between 350 and 400 nm, arranged at such a distance that the power emitted is between 5 and 100 W/m.sup.2. Preferably the distance is equal to 20 cm, the time is between 1 and 30 minutes and preferably equal to 15 minutes, and the intensity of UV radiation is equal to 55 W/m.sup.2.
b) As an alternative the photo-reduction reaction can occur thanks the direct or indirect exposition of the treated mulberry leaves to solar light in a maximum time interval equal to at least 4 hours in case of direct light (preferably equal to 60 minutes) and 8 hours maximum in case of indirect light (preferably equal to 120 minutes).

4. Washing and Storing

[0041] In the following the leaves treated with silver are washed in water to remove the not reacted silver precursor (if present), and in the following stored in refrigerator up to their use.

Example No 2Treatment of Feedstuff Obtained From Mulberry Leaves Chopped and Additivated by Means of Photo-Chemical Deposition of Silver Particles

[0042] The procedure of treatment provides to prepare a solution in the same way as in example no 1, and to apply such solution on mulberry leaves in powder dried, chopped and additivated with nutrient substances (feedstuff).

[0043] In the following a layer of uniform thickness of feedstuff is arranged on a surface. Preferably the layer thickness is lower than 3 mm.

[0044] The next step consists in impregnating with the solution described in example no 1 the powder of mulberry leaves dried by means of spray coating. Preferably a quantity of solution equal to about 50 g/m.sup.2 is sprayed.

[0045] In the following there occurs the exposition to ultraviolet light (preferably with emission spectra peak between 350 and 400 nm) to obtain the photo-reduction of silver nitrate in metal nitrate after an exposition time between 1 and 30 minutes and which is preferably equal to 15 minutes. Preferably it can be used a UV lamp configured so that it produces a radiation intensity about 55 W/m.sup.2.

[0046] As an alternative, the photo-reduction reaction can occur thanks to the direct or indirect exposition of the impregnated feedstuff to solar light in a maximum time interval equal to at least 4 hours in case of direct light (preferably equal to 60 minutes) and maximum 8 hours in case of indirect light (preferably equal to 120 minutes).

Example No 3Treatment by Means of Photo-Chemical Deposition of Silver Particles on Mulberry Leaves Directly on the Tree Before Picking

[0047] The procedure of treatment provides to prepare a solution in the same way as in example no 1, and to spray by means of the spray coating technique a quantity between 25 and 100 g/m.sup.2, and preferably equal to 50 g/m.sup.2 of solution on leaves before picking them from the tree. The photo-reduction reaction will occur thanks to the direct or indirect exposition to the solar light in a maximum time interval equal to at least 4 hours in case of direct light (preferably equal to 60 minutes) and maximum 8 hours in case of indirect light (preferably equal to 120 minutes). The leaves will be then picked and subjected to the normal working cycles (washing, chopping etc.).

[0048] The advantages associated to these treatment modes are the following: [0049] the problem concerning the storing time of the treated leaves is eliminated; in this case the leaf treated with silver can be picked also after various weeks after the treatment; [0050] such mode of treatment directly on the plant or tree can be carried out only using the photo-reduction technique which allows to obtain a strong adhesion and stability in the time of silver to the leaf; a treatment with colloidal silver o other techniques widely reported in literature would not resist in the time especially if exposed to atmospheric agents (wind, rain etc.); [0051] lower overheating and/or partial deterioration of the leaf with respect to the technique providing the use of UV lamps; [0052] simplification of the process which does not provide the use of industrial systems for UV radiation of the leaves treated with the solution containing the silver precursor; [0053] lower environmental impact of the present technique.

Example No 4

[0054] Treatment with mulberry leaves or feedstuff obtained from mulberry leaves by photo-chemical deposition of silver and colloidal zinc particles The procedure of treatment aiming at promoting a synergistic action induced by the contemporaneous presence of silver and zinc provides to prepare a solution in the same way as in example no 1 and to add a solution of colloidal zinc thereto.

[0055] The solution of colloidal zinc comprises nanoparticles between 10 and 100 nm, preferably 60 nm, and can be added in concentration between 25 and 100 ppm, preferably 70 ppm, in quantity between and 30% with respect to the silver based solution, preferably 10%.

[0056] The following operations are the same as in example and for brevity of exposition will be not reported.

Example No 5

[0057] Treatment of mulberry leaves or feedstuff obtained from mulberry leaves by means of photo-chemical deposition of silver and micronized zinc particles The procedure of treatment aiming at promoting a synergistic action induced by the contemporaneous presence of silver and zinc provides to prepare a solution in the same way as in example no 1 and to add a solution of micronized zinc thereto.

[0058] The solution of micronized zinc comprises nanoparticles between 0.1 and 100 micron, and can be added in concentration between 25 and 100 ppm.

[0059] The following operations are the same as in example and for brevity of exposition will be not reported.