Method for making coffee husk reducing agent

Abstract

An efficient green method for the synthesis of noble metal/transition metal oxide nanocomposite comprising reducing noble metal salt and a templating metal oxide is disclosed. The method is a one-step method comprises mixing coffee seed husk extract, a noble metal precursor, and a transition metal precursor; and filtering and drying the nanocomposite. The nanocomposite prepared by the method of the invention displays all the characteristics and biocidal activity of a composite prepared by traditional methods.

Claims

1. A method of preparing and using a reducing agent, comprising: drying and grinding a seed husk to form a powder, adding the powder to a solvent to form a mixture, heating the mixture at a temperature in the range of 70-100° C. for a time in the range of 10-60 min to form a solid, and filtering away the solid to obtain the reducing agent, adding the reducing agent to an aqueous solution of a noble metal precursor to form a noble metal nanoparticle suspension, adding a solution of a transition metal oxide precursor to the noble metal nanoparticle suspension to form a noble metal/transition metal oxide suspension, filtering the noble metal/transition metal oxide suspension to isolate a noble metal/transition metal oxide nanocomposite precursor, then calcining the noble metal/transition metal oxide nanocomposite precursor to form a noble metal/transition metal oxide nanocomposite, wherein the noble metal/transition metal oxide nanocomposite has a particle size range of from 8 to 10 nm and meso porous/macro porous channels with an interior width of 1.37 to 1.5 μm.

2. The method of claim 1, wherein the seed husk is a Coffea species seed husk.

3. The method of claim 2, wherein the Coffea species is Coffea arabica or Coffea canephora.

4. The method of claim 1, wherein the solvent is water, alcohol, or combination thereof.

5. The method of claim 1, wherein the solvent is an alcohol selected from the group consisting of methanol, ethanol, propanol, isopropanol, butanol, and isobutanol.

6. The method of claim 1, wherein the reducing agent is added to the aqueous solution of the noble metal precursor while sonicating the aqueous solution of the noble metal precursor with ultrasound, and the solution of the transition metal oxide precursor is added to the noble metal nanoparticle suspension while sonicating the noble metal nanoparticle suspension with ultrasound.

7. The method of claim 6, wherein the noble metal/transition metal oxide nanocomposite comprises a noble metal in an amount in the range of 1 wt. % to 40 wt. % based on the total weight of the noble metal/transition metal oxide nanocomposite.

8. The method of claim 6, wherein the noble metal is silver, palladium, platinum, gold, ruthenium, rhodium, osmium, or iridium.

9. The method of claim 6, wherein the noble metal precursor is a noble metal salt.

10. The method of claim 9, wherein the noble metal salt is a noble metal chloride, bromide, iodide, acetate, or nitrate.

11. The method of claim 6, wherein the noble metal precursor is silver nitrate.

12. The method of claim 6, wherein the transition metal oxide is zinc oxide, titanium oxide, iron oxide, cobalt oxide, nickel oxide, zirconium oxide, niobium oxide or molybdenum oxide.

13. The method of claim 6, wherein the transition metal oxide precursor is a transition metal alkoxide, chloride, bromide, iodide, or acetate.

14. The method of claim 6, wherein the noble metal/transition metal oxide nanocomposite comprises silver and titanium oxide or zinc oxide.

15. The method of claim 6, wherein the noble metal precursor is silver nitrate and the transition metal oxide precursor is titanium tetraisopropoxide.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) A more complete appreciation of the invention and many of the attendant advantages thereof will be readily obtained as the same becomes better understood by reference to the following detailed description when considered in connection with the accompanying drawings, wherein:

(2) FIG. 1A shows a method of making coffee seed husk extract.

(3) FIG. 1B shows a schematic representation of preparing M/TiO.sub.2 using coffee seed husk extract.

(4) FIG. 1C shows colloidal suspensions of Au and Ag nanoparticles prepared using coffee seed husk extract.

(5) FIG. 2A shows XRD patterns of green Ag nanoparticles, TiO.sub.2 nanoparticles and Ag—TiO.sub.2 nanocomposites of various Ag ratios.

(6) FIG. 2B shows XRD patterns of green Au nanoparticles, TiO.sub.2 nanoparticles and 5% Au—TiO.sub.2 nanocomposite.

(7) FIG. 3 shows UV-vis diffuse reflectance spectra of the as prepared green TiO.sub.2 nanoparticles and Ag—TiO.sub.2 nanocomposites having 5 wt. %, 10 wt. %. and 20 wt. % of Ag of the total weight of the composite.

(8) FIG. 4 shows FTIR spectra of the as prepared green TiO.sub.2 nanoparticles and Ag—TiO.sub.2 nanocomposites having 5 wt. %, 10 wt. %. and 20 wt. % of Ag of the total weight of the composite.

(9) FIG. 5A shows SEM image of Ag nanoparticles.

(10) FIG. 5B shows SEM image of Ag/TiO.sub.2 nanocomposite.

(11) FIG. 6A shows TEM image of Ag nanoparticles synthesized by the green method.

(12) FIG. 6B shows TEM image of 5% Ag/TiO.sub.2 nanocomposite synthesized by the green method at 1× magnification.

(13) FIG. 6C shows TEM image of 5% Ag/TiO.sub.2 nanocomposite synthesized by a chemical method.

(14) FIG. 7A shows TEM image of 5% Ag/TiO.sub.2 nanocomposite synthesized by the green method at 4× magnification.

(15) FIG. 7B shows TEM image of 5% Ag/TiO.sub.2 nanocomposite synthesized by the green method at 8× magnification.

(16) FIG. 8A shows the growth of Aspergillus niger on an agar well containing Ag/TiO.sub.2 nanocomposite at the center of the plate.

(17) FIG. 8B shows the growth of Aspergillus niger on an agar well containing Ag nanoparticles at the center of the plate.

(18) FIG. 9A shows an optical micrograph of control mycelia of Aspergillus niger.

(19) FIG. 9B shows an optical micrograph of mycelia of Aspergillus niger treat with Ag/TiO.sub.2.

(20) FIG. 10 shows the effects of the green synthesized 5 mmol % Ag—TiO.sub.2 nanocomposites on radial growth of fungi. (a) Control sample, (b-e) at concentrations of 1, 2, 3, 4 mg/ml of Ag—TiO.sub.2 nanocomposite from left to right, respectively.

(21) FIG. 11A shows a UV-vis spectrum of the Coffee Husk Extract (CHE);

(22) FIG. 11B shows an FTIR of the Coffee Husk Extract (CHE).

(23) FIG. 12A shows the antifungal activity of Ag/TiO.sub.2 nanocomposites with 1% Ag concentration;

(24) FIG. 12B shows the antifungal activity of Ag/TiO.sub.2 nanocomposites with 5% Ag concentration;

(25) FIG. 12C shows the antifungal activity of Ag/TiO.sub.2 nanocomposites with 10% Ag concentration;

DETAILED DESCRIPTION

(26) Embodiments of the present disclosure will now be described more fully hereinafter with reference to the accompanying drawings, in which some, but not all embodiments of the disclosure are shown. The present disclosure will be better understood with reference to the following definitions.

(27) All publications mentioned herein are incorporated herein by reference in full for the purpose of describing and disclosing the methodologies, which are described in the publications, which might be used in connection with the description herein. Nothing herein is to be construed as an admission that the inventors are not entitled to antedate such disclosure by virtue of prior disclosure. Also, the use of “or” means “and/or” unless stated otherwise. Similarly, “comprise,” “comprises,” “comprising” “include,” “includes,” and “including” are interchangeable and not intended to be limiting.

(28) Terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention.

(29) The headings (such as “Background” and “Summary”) and sub-headings used herein are intended only for general organization of topics within the present invention, and are not intended to limit the disclosure of the present invention or any aspect thereof. In particular, subject matter disclosed in the “Background” may include novel technology and may not constitute a recitation of prior art. Subject matter disclosed in the “Summary” is not an exhaustive or complete disclosure of the entire scope of the technology or any embodiments thereof. Classification or discussion of a material within a section of this specification as having a particular utility is made for convenience, and no inference should be drawn that the material must necessarily or solely function in accordance with its classification herein when it is used in any given composition.

(30) It will be further understood that the terms “comprises” and/or “comprising,” when used in this specification, specify the presence of stated features, steps, operations, elements, and/or components, but do not preclude the presence or addition of one or more other features, steps, operations, elements, components, and/or groups thereof.

(31) As used herein, the term “and/or” includes any and all combinations of one or more of the associated listed items and may be abbreviated as “/”.

(32) Links are disabled by deletion of http: or by insertion of a space or underlined space before www. In some instances, the text available via the link on the “last accessed” date may be incorporated by reference.

(33) As used herein, the term “compound” is intended to refer to a chemical entity, whether in a solid, liquid or gaseous phase, and whether in a crude mixture or purified and isolated.

(34) As used herein, the term “salt” refers to derivatives of the disclosed compounds, monomers or polymers wherein the parent compound is modified by making acid or base salts thereof. Exemplary salts include, but are not limited to, mineral or organic acid salts of basic groups such as amines, and alkali or organic salts of acidic groups such as carboxylic acids. The salts of the present disclosure can be synthesized from the parent compound that contains a basic or acidic moiety by conventional chemical methods. Generally such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally non-aqueous media like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are preferred.

(35) As used herein, the term “about” refers to an approximate number within 20% of a stated value, preferably within 15% of a stated value, more preferably within 10% of a stated value, and most preferably within 5% of a stated value. For example, if a stated value is about 8.0, the value may vary in the range of 8±1.6, ±1.0, ±0.8, ±0.5, ±0.4, ±0.3, ±0.2, or ±0.1.

(36) Disclosure of values and ranges of values for specific parameters (such as temperatures, molecular weights, weight percentages, etc.) are not exclusive of other values and ranges of values useful herein. It is envisioned that two or more specific exemplified values for a given parameter may define endpoints for a range of values that may be claimed for the parameter. For example, if Parameter X is exemplified herein to have value A and also exemplified to have value Z, it is envisioned that parameter X may have a range of values from about A to about Z. Similarly, it is envisioned that disclosure of two or more ranges of values for a parameter (whether such ranges are nested, overlapping or distinct) subsume all possible combination of ranges for the value that might be claimed using endpoints of the disclosed ranges. For example, if parameter X is exemplified herein to have values in the range of 1-10 it also describes subranges for Parameter X including 1-9, 1-8, 1-7, 2-9, 2-8, 2-7, 3-9, 3-8, 3-7, 2-8, 3-7, 4-6, or 7-10, 8-10 or 9-10 as mere examples. A range encompasses its endpoints as well as values inside of an endpoint, for example, the range 0-5 includes 0, >0, 1, 2, 3, 4, <5 and 5.

(37) As used herein, the words “preferred” and “preferably” refer to embodiments of the technology that afford certain benefits, under certain circumstances. However, other embodiments may also be preferred, under the same or other circumstances. Furthermore, the recitation of one or more preferred embodiments does not imply that other embodiments are not useful, and is not intended to exclude other embodiments from the scope of the technology.

(38) The description and specific examples, while indicating embodiments of the technology, are intended for purposes of illustration only and are not intended to limit the scope of the technology. Moreover, recitation of multiple embodiments having stated features is not intended to exclude other embodiments having additional features, or other embodiments incorporating different combinations of the stated features. Specific examples are provided for illustrative purposes of how to make and use the compositions and methods of this technology and, unless explicitly stated otherwise, are not intended to be a representation that given embodiments of this technology have, or have not, been made or tested.

(39) The method of the invention utilizes an extract of waste plant materials as a reducing material. Many waste plant material comprises several organic compounds such as, but not limited to polyphenol, quinic acid, vanilic acid, ferulic acid, chlorogenic acid, caffeic acid, tannin, lignine, and caffeine, which may act alone or in combinations as reducing agent for noble metal salts. As used herein, the term “waste plant material” referrer to a byproduct of processing plants product such as seed husk, dry leaves, branches, wood cuttings, and the like. Such products ordinarily have no economic value and are usually discarded or burned.

(40) The seed husk used in the method of the invention may be any husk of any plant seed such as, but not limited to rice, wheat, corn, coffee, soy, and barley. In some embodiments, the husk is of a seed of a plant belong to the genus Coffea, which is a genus of flowering plants in the family Rubiaceae. Coffea species are shrubs or small trees native to tropical and southern Africa and tropical Asia. The seeds of some species, called coffee beans, are used to flavor various beverages and products. The fruits, like the seeds, contain a large amount of caffeine, and have a distinct sweet taste and are often juiced. The plant ranks as one of the world's most valuable and widely traded commodity crops and is an important export product of several countries, including those in Central and South America, the Caribbean and Africa. There are over 120 species of Coffea, which is grown from seed. The two most popular are Coffea arabica, commonly known as “Arabica” and Coffea canephora, known as “Robusta”. Arabica accounts for 60-80% of the world's coffee production and Robusta accounts for about 20-40%. As such, a considerable amount of coffee seed husks are produced as a byproduct of processing coffee beans. In some embodiments of the invention, the husk is obtained from Coffea arabica or Coffea canephora.

(41) In some embodiments of the present disclosure, the waste plant material is coffee seed husk, in particular, Coffea arabica or Coffea canephora seed husk. The husk is the outer layer of the coffee bean that breaks away and is discarded during the roasting process. Large quantities of coffee seed husk are produced every day after roasting coffee beans. An extract may be prepared by adding ground dry husk to a solvent and optionally heating it, e.g., heating to the boiling point of the solvent, for a time in the range of 0.1-3 h, preferably 0.2-2.5 h, preferably 0.3-2 h, preferably 0.4-1 h, preferably about 0.5 h. The solvent may be water, alcohol such methanol, ethanol, propanol, isopropanol, butanol, isobutanol, and the like, ether such as dimethyl ether, methyl ethyl ether, diethyl ether, halogenated hydrocarbons such as methylene chloride, chloroform, carbon tetrachloride, and the like, acetone, acetonitrile, and the like. In a preferred embodiment of the method, the solvent is water and the extract is prepared by boiling an amount of dried and comminuted coffee been husk in the range of 1-10 wt. %, preferably 2-9 wt. %, preferably 3-8 wt. %, preferably 4-7 wt. %, preferably 4.5-5.5 wt. % for about 0.5 h.

(42) The method of the invention comprises mixing a solution of a noble metal precursor with the coffee husk extract described herein for a time in the range of 1-48 h, preferably 2-24 h, preferably 3-15 h, preferably 4-12 h, preferably 5-10 h. As used herein, the term “noble metal” is a metal selected from the group consisting of ruthenium (Ru), rhodium (Rh), palladium (Pd), silver (Ag), osmium (Os), iridium (Ir), platinum (Pt), and gold (Au). As used herein the word “diameter” refers to the average diameter of a nanoparticle measured by transmission electron microscopy (TEM). The nanoparticles may contain nanoparticles of one or more of the noble metals such that the nanoparticles have a diameter in the range of 1-999 nm, preferably, 20-500 nm, preferably 30-100 nm, preferably 40-80 nm, preferably 50-70 nm, preferably 55-60 nm. In some embodiments of the invention, the noble metal nanoparticles are silver or gold having a diameter in the range of 5 nm to 100 nm, preferably, 10 nm to 80, preferably 20 nm to 60 nm, preferably 30 nm to 50 nm, preferably 35 nm to 45 nm. The noble metal precursor may be any salt or compound of a noble metal such as but not limited to AuF.sub.3, AuF.sub.5, (AuCl.sub.3).sub.2, Au.sub.4Cl.sub.8, AuBr, AuBr.sub.3, AuI, AuI.sub.3, Au.sub.2O.sub.3, Au.sub.2S, Au.sub.2S.sub.3, HAuCl.sub.4, PtCl.sub.2, PtCl.sub.4 PtF.sub.4, PtO.sub.2, H.sub.2PtCl.sub.6, palladium halide including but not limited to PdF.sub.2, PdF.sub.3, PdF.sub.4, PdF.sub.6, Pd(PdF.sub.6), palladium acetate, palladium acetylacetonate, PdCl.sub.2, PdBr.sub.2, palladium cyanide, Pd(NO.sub.3).sub.2, Ag.sub.2CO.sub.3, AgF.sub.2, AgNO.sub.3, silver acetate, OsO.sub.4, OsO.sub.2, RuO.sub.2, RuO.sub.4, K.sub.2RuO.sub.4, RuCl.sub.3, RuF.sub.3, RhCl.sub.3, RhF.sub.6, RhO.sub.2, Na.sub.2RhO.sub.3, IrO.sub.2, Ir.sub.2O.sub.3, IrO.sub.4, IrCl.sub.2, IrCl.sub.3, and the like.

(43) The noble metal nanoparticle may be separated from the reaction mixture by filtration or centrifugation. In some embodiments, the product may be calcined to remove any remnant of organic material at a temperature of at least 150° C., preferably 200° C., preferably 250° C., preferably 300° C., preferably 350° C., preferably 400° C. for a time in the range of 2-12 h, preferably 4-10 h, preferably 5-6 h.

(44) Another aspect of the invention is directed to a method of making a nanocomposite of a noble metal nanoparticle supported on a transition metal oxide that is similar to the method described herein above for making noble metal nanoparticles. The method comprises mixing a solution of noble metal precursor with the coffee husk extract as described herein above to form a suspension of noble metal nanoparticle in coffee husk extract. To the suspension, a solution of a transition metal oxide precursor is mixed with agitation and the mixture is allowed to stand for of 1-48 h, preferably 2-24 h, preferably 3-15 h, preferably 4-12 h, preferably 5-10 h.

(45) The amount of noble metal in the nanocomposite is in the range of 1 wt. % to 30 wt. %, preferably 2 wt. % to 25 wt. %, preferably of 5 wt. % to 20 wt. %, preferably 8 wt. % to 15 wt. % of the total weight of the nanocomposite.

(46) The transition metal oxide nanoparticles have a diameter in the range of 20 nm to 500 nm, preferably 30 nm to 250 nm, preferably 50 nm to 100 nm, preferably 60 nm to 80 nm. The transition metal oxide is an oxide of an element such as, but not limited to, Sc, Ti, V, Cr, Mn, Fe, Co, Ni, Zn, Y, Zr, Nb, and Mo. In some preferred embodiments of the invention, the metal oxide is selected from the group consisting of Zn, V, Cr, Mn, Fe, Co, and Ni. Preferably the metal oxide is that of Ti. Titanium dioxide, also known as titanium(IV) oxide or titania, is a naturally occurring oxide of titanium.

(47) Any suitable metal salt such as that of, but not limited to, Sc, Ti, V, Cr, Mn, Fe, Co, Ni, Zn, Y, Zr, Nb, and Mo may be utilized. Examples of the salts include, but are not limited to halides, carboxylate, carbonate, alkoxide and the like such as, but not limited to ZnCl.sub.2, ZnBr.sub.2, zinc acetate, zinc isopropoxide, Zn(OCH.sub.3).sub.2, titanium isopropoxide, ZrCl.sub.4, Zr.sub.4(OC.sub.2H.sub.5).sub.16, FeCl.sub.2, FeCl.sub.3, TiCl.sub.4, Ti.sub.4(OC.sub.2H.sub.5).sub.16, titanium tetraisopropoxide, VCl.sub.3, VBr.sub.3, vanadium acetate, vanadium oxytriisopropoxide, MnCl.sub.2, MnBr.sub.2, manganese acetate, and the like.

(48) In some embodiments, the mixture may be heated at a temperature in the range of 100-200° C., preferably in the range of 110-180° C., preferably in the range of 120-160° C., preferably at about 150° C. for a time in the range of 10 to 20 hours for a time in the range of 10 to 20 hours for a time in the range of 5 to 30 hours, preferably in the range of 10 to 25 hours, preferably, about 15 hours. The noble metal nanoparticles or nanocomposite are filtered, washed, and dried at a temperature in the range of 50-120° C., preferably 60-100° C., and preferably 70-80° C. In some embodiments, the nanocomposite product may be calcined to remove any remnant of organic material at a temperature of at least 150° C., preferably 200° C., preferably 250° C., preferably 300° C., preferably 350° C., preferably 400° C. for a time in the range of 2-12 h, preferably 4-10 h, preferably 5-6 h, preferably about 3 h.

(49) One of the advantages of the method of the invention is that it produces highly porous metal oxide nanoparticles. The highly porous nanoparticles are templated by some of the components of coffee seed husk extracts. The high porosity of the metal oxide of the composite makes it suitable for use in many applications including, but not limited to catalysis, electrochemical applications, and adsorption.

(50) Another aspect of the invention is directed to a method of preparing a reducing agent comprising washing a seed husk, drying and grinding the seed husk to form a powder, adding the powder to a solvent to form a mixture, heating the mixture at a temperature in the range of 70-100° C. for a time in the range of 10-60 min, and filtering away the solid to obtain the reducing agent. Any solvent may be use in the method such as but not limited to water, ethanol, methanol, propanol, isopropanol, butanol, isobutanol, pentane, cyclopentane, hexane, cyclohexane, acetone, diemthyl ether, diethyl ether, acetonitrile, dimethylformamide, chloroform, methylene chloride, carbontetrachloride and combination thereof.

(51) The husk used in the method of the invention may be any husk of any plant seed such as, but not limited to rice, wheat, corn, coffee, soy, and barley. In some embodiments, the husk is of a seed of a plant belong to the genus Coffea, in particular, Coffea arabica or Coffea canephora as large amounts of husk produced every day from processing coffee beans. In a particular embodiment, the method mixing water and a dry ground husk powder of seed husk from Coffea arabica or Coffea canephora and heating the mixture to a temperature in the range of 70-100° C., preferably 80-100° C., preferably 90-100° C., preferably 95-100° C. for a time in the range of 10-120 min, preferably 10-80 min, preferably 20-60 min, preferably 30-40 min. The amount of powder is in the range of 1-15 wt. %, preferably 2-13 wt. %, preferably 3-12 wt. %, preferably 4-10 wt. %, preferably 5-8 wt. % of the total weight

Example 1

(52) Preparation of Coffee Seed Husk Extract (CSHE):

(53) The coffee husks were washed several times with distilled water to remove the dust particles on their surface then air dried. The dried husk was grinded to obtain fine powder. Coffee husk powder (20 g) was boiled with 400 ml of distilled water for 30 min, then allowed to cool down slightly and finally filtered to obtain clear brown solution of CSHE (see FIG. 1A).

(54) Table 1 provides a qualitative comparison of various components in coffee husk and coffee beans.

(55) TABLE-US-00001 TABLE 1 Chemical compounds Chemical compounds in Coffee Husk in Coffee Been Caffeine (low level) Caffeine (high level) Tannin Tannin (high level) Lignin — Cellulose NA Hemicellulose NA Caffeic acid (low level) Caffeic acid (high level) Chlorogenic acid (low level) Chlorogenic acid (high level) Ferulic acid Ferulic acid Vanillic acid NA quinic acid NA Polyphenols Polyphenols Andrade et al. “Supercritical fluid extraction from spent coffee grounds and coffee husks: Antioxidant activity and effect of operational variables on extract composition” Talanta, vol. 88, pp. 544-552 (2012); .sup.2United Kingdom “P & A-084 In Vitro Antioxidant Activity of Coffee Compounds and Their” no. Ldl, pp. 6962-6969 (2007); .sup.3Dhand et al. “Green synthesis of silver nanoparticles using Coffea arabica seed extract and its antibacterial activity” Mater. Sci. Eng. C, vol. 58, pp. 36-43 (2016); .sup.4Louarn et al. “Caffeine, trigonelline, chlorogenic acids and sucrose diversity in wild Coffea arabica L. and C. canephora P. accessions” Food Chem., vol. 75, no. 2, pp. 223-230 (2001), .sup.5Mohan et al. “Biotechnological potential of coffee pulp and coffee husk for bioprocesses” Biochem. Eng. J., vol. 6, pp. 153-162 (2002); .sup.6Brand et al. “Biological detoxification of coffee husk by filamentous fungi using a solid state fermentation system” Enzyme Microb. Technol., vol. 27, no. 1-2, pp. 127-133 (2000); .sup.7Mussatto et al. “Production, Composition, and Application of Coffee and Its Industrial Residues” Food Bioprocess Technol., vol. 4, no. 5, pp. 661-672 (2011); .sup.8Castaldo et al. “Study of the Chemical Components Bioactivity and Antifungal Properties of the Coffee Husk” J. Food Res., vol. 7, no. 4, p. 43 (2018); .sup.9Al-Yousef et al. “Essential oil of Coffee arabica L. Husks: A brilliant source of antimicrobial and antioxidant agents” Biomed. Res., vol. 29, no. 1, pp. 174-180 (2018), .sup.10 Campos-Vega et al. “Spent coffee grounds: A review on current research and future prospects” Trends Food Sci. Technol., vol. 45, no. 1, pp. 24-36 (2015); .sup.11 Mazzafera, P. “Degradation of caffeine by microorganisms and potential use of decaffeinated coffee husk and pulp in animal feeding” Sci. Agric., vol. 59, no. 4, pp. 815-821, (2005); and .sup.12 Oliveira and A. S. Franca, An Overview of the Potential Uses for Coffee Husks. Elsevier Inc. (2014)—each incorporated by reference.

Example 2

(56) Preparation of Silver Nanoparticles and Nanocomposite:

(57) Silver nanoparticles were prepared by dropwise addition of CSHE on an aqueous solution of silver nitrate agitated with ultrasound to form silver nanoparticles. Ag—TiO.sub.2 nanocomposite was prepared by dropwise addition of an aqueous solution of titanium tetraisopropoxide to a solution containing the desired amount of silver nanoparticles and CSHE with agitation at ambient temperature (see FIGS. 1B and 1C). The resulting precipitate was filtered, washed with deionized water, dried, and calcined at 300° C. for one hour to produce porous nanocomposite of Ag—TiO.sub.2. The same procedure was used to obtain TiO.sub.2 from titanium tetraisopropoxide and gold nanoparticles from gold salt (see FIG. 1C).

(58) Characterization of the Nanocomposite:

(59) Crystalline structure, morphology and optical properties of the produced green nanomaterials and nanocomposites have been examined by XRD, UV-vis diffuse reflectance spectroscopy, and FTIR spectroscopy. The results show that the green prepared nanomaterials are pure and have high crystallinity and modified size and shape with narrow particle distribution as compared to the corresponding nanomaterials prepared by chemical methods. The nanomaterials have meso porous/macro porous channels with 1.37-1.5 μm interior width which are well arranged, particle size ranged from 8 to 10 nm. FTIR and UV-Vis spectra of FIGS. 11B and 11A, respectively, reveal that phenolic compounds are present in CSHE and act efficiently as reducing and templating/stabilizing agents during synthesis process. The FTIR spectra of the obtained CSHE shows broad absorption peak from 3600 to 3200 cm.sup.−1 which can be assigned to an —OH group of polyphenolic compounds. The absorption at 1700 cm.sup.−1 corresponds to the carbonyl stretching vibration of free caffeine. A peak at 1655 cm.sup.−1 of vs (CO)+v (CC) in free caffeine. The Quadratal CN stretching in imidazole ring appearing at 1403 cm.sup.−1 of the caffeine. The vibrations of free caffeine appearing at 2900, 1700, 1683, 1456 and the broad bands at 1284-973, 925 cm.sup.−1.

(60) CHE was further analyzed by UV-vis absorption spectroscopy measurement (FIG. 11A) with absorption in the range 200-400 nm. Two peaks were observed, one peak at 275 nm is attributed to caffeine and the peak at 320 nm is attributed to Caffeic acid. The results indicate the presence of polyphenolic, Caffeic acid and Caffeine which act as bio-templating agents for the synthesis of the porous TiO.sub.2 nanoparticles. [See U. T. C. P. Souto, M. F. Barbosa, H. V. Dantas, A. S. Pontes, W. S. Lyra, P. H. G. D. Diniz, M. C. U. Araújo, E. CSilva, Food Science and Technology, 2015, 63, 1037-1041, and Diding Suhandy & Meinilwita Yulia, International Journal of Food Properties, 2017, 20, S331-S339—each incorporated by reference].

(61) The effect of the ratio of Ag nanoparticles in the nanocomposite was studied. It was found that no noticeable change in the antifungal activity was observed among various Ag/TiO.sub.2 samples with various Ag ratios (see FIGS. 12A, 12B and 12C).

(62) FIG. 2A shows XRD patterns of the as prepared green Ag, TiO.sub.2 nanoparticles, and Ag—TiO.sub.2 nanocomposites having 5 wt. %, 10 wt. %. and 20 wt. % of Ag of the total weight of the composite. FIG. 2B shows XRD of Au, TiO.sub.2, and composite of 5% Au/TiO.sub.2.

(63) FIG. 3 shows UV-vis diffuse reflectance spectra of the as prepared green TiO.sub.2 nanoparticles and Ag—TiO.sub.2 nanocomposites having 5 wt. %, 10 wt. %. and 20 wt. % of Ag of the total weight of the composite.

(64) FIG. 4 shows FTIR spectra of the as prepared green TiO.sub.2 nanoparticles and Ag—TiO.sub.2 nanocomposites having 5 wt. %, 10 wt. %. and 20 wt. % of Ag of the total weight of the composite.

(65) FIGS. 5A and 5B show the SEM images of Ag nanoparticles and 5% Ag/TiO.sub.2 nanocomposite, respectively, prepared by the method of invention.

(66) FIG. 6A shows a TEM image of Ag nanoparticles prepared by the method of invention. FIGS. 6B and 6C show the TEM image of 5% Ag/TiO.sub.2 nanocomposite, respectively, prepared by the method of invention and by conventional method, respectively, at 1× magnification. FIGS. 7A and 7B show TEM images at 4× and 8× magnification, respectively.

Example 3

(67) Biocidal Activity of the Nanoparticles:

(68) The biocidal activity of the nanoparticles and nanocomposites prepared by the method of invention were examined by the agar well diffusion method in cultures of Aspergillus flavus and Aspergillus niger. Different concentrations of Ag and Ag/TiO.sub.2 nanocomposite were added to 90 ml of molten potato dextrose agar, pH 6. PDA petri plates without any nanoparticles and nanocomposite were incubated as a control. All plates were inoculated in the center with 6 mm diameter fungal disc. All the plates were incubated at room temperature 28±2° C. for 4 days. The growth of the fungal pathogen in treated plates was measured when the growth of the organism in control plate was full (FIGS. 8A and 8B). Each experiment was repeated thrice and the average values were recorded. The presence of clearance zone indicates the high fungal sensitivity to Ag/TiO.sub.2 nanocomposite more than Ag nanoparticles. The negative control (water) had no effect on the tested fungi (plate (a) in FIG. 10 in which the concentration of the nanocomposite used in experiments b-e was in the following order: 1 mg/ml, 2 mg/ml, 3 mg/ml, 4 mg/ml].

(69) Ag/TiO.sub.2 and Ag inhibited the growth of tested fungi and the results are summarized in Tables 1-3.

(70) TABLE-US-00002 TABLE 1 Antifungal activity of Ag and Ag/TiO.sub.2 against Aspergillus niger and Aspergillus flavus. Fungal Treatment with Ag Treatment with Ag/TiO2 strains 0.5 mg/ml 1 mg/ml 2 mg/ml 3 mg/ml 4 mg/ml 0.5 mg/ml 1 mg/ml 2 mg/ml 3 mg/ml 4 mg/ml Inhibition zones (mm) A. 13 ± 0.43 14.5 ± 0.23 15 ± 0.32 16.5 ± 0.35 16.5 ± 0.3 13.5 ± 0.30 15 ± 0.33   16 ± 0.25 17 ± 0.33 20 ± 0.45 niger A. 14 ± 0.65 15.5 ± 0.25 16 ± 0.25 17.5 ± 0.35 17.5 ± 0.2   17 ± 0.30 19 ± 0.5  18.75 ± 0.25 19 ± 0.32 19 ± 0.35 flavus

(71) TABLE-US-00003 TABLE 2 Percentage of radial growth inhibition* Fungal Treatment with Ag Treatment with Ag/TiO2 strains 0.5 mg/ml 1 mg/ml 2 mg/ml 3 mg/ml 4 mg/ml 0.5 mg/ml 1 mg/ml 2 mg/ml 3 mg/ml 4 mg/ml Mycelium growth inhibition (%) A. niger 68 ± 3.5 87 ± 2.5 100 100 100 65 ± 2.5 82 ± 2.8 100 100 100 A. flavus 76 ± 4   94 ± 3.3 100 100 100 100 100 100 100 100 *Values are given as mean ± S.D. of three experiments

(72) TABLE-US-00004 TABLE 3 Minimum Inhibitory concentration (MIC) and MFC values recorded against Aspergillus flavus and Aspergillus niger Treatment (mg/ml) Ag/TiO.sub.2 Flucanazole* Fungi MIC MFC MIC MFC A. flavus 0.2 0.35 0.1 0.1 A. niger 0.5 1 0.1 0.1 *Antigoni Elefanti, Johan W. Mouton, Katerina Krompa, Rafal Al-Saigh, Paul E. Verweij, Loukia Zerva, andJoseph Meletiadis, Inhibitory and Fungicidal Effects of Antifungal Drugs against Aspergillus species in the presence of serum, Antimicrob Agents Chemother. 2013 Apr; 57(4): 1625-1631. doi: 10.1128/AAC.01573-12. MFC: Minimum Fungicidal Concentration with concentration in mg/ml.

(73) The maximum growth inhibition of A. flavus was observed at concentration of 1 mg/mL of Ag/TiO.sub.2 nanocomposite and 3 mg/ml of Ag nanoparticles. In case of A. niger, Ag/TiO.sub.2 inhibited completely the growth at concentration of 3 mg/ml, while Ag completely inhibited the growth at concentration of 4 mg/ml. For the two strains, the inhibition is irreversible. Thus, the growth sensitivity of Aspergillus strains to Ag/TiO.sub.2 nanocomposite and Ag nanoparticles was species dependent and concentrations dependent in the case of Aspergillus niger. The morphology of mycelia of Aspergillus niger treated with Ag/TiO.sub.2 nanocomposite was significantly altered from that intact mycelia of the control (compare FIGS. 9A and 9B).

(74) In order to explore the impact of Ag/TiO.sub.2 nanocomposite and Ag nanoparticles on intracellular pH level, concentrations of 1 mg/mL and 4 mg/mL of Ag/TiO.sub.2 were tested respectively against A. flavus and A. niger. The initial pH was adjusted to pH 6. The prepared suspension was kept in a shaking incubator at speed of 200 rpm at 28° C. The pH changes in suspension were measured using pH meter. The result demonstrates that only control fungi maintain constant internal pH while treated strains shows increase in internal acidification to provide a culture acidity of pH 6. The increase was more pronounced in cultures of A. flavus. The result suggests that, the produced green nanomaterials exhibit antifungal activity by targeting H.sup.+-ATPase located in the fungi membranes causing intracellular acidification which leads to cell death.