LT5802B - Cationic starch-triiodide bactericide - Google Patents

Abstract

The present invention relates to chemistry and can be used in manufacture of polymeric biocide for disinfecting water and biological liquids. The efficiency of polymeric bactericides consisting of molecular iodine bound to polymer macromolecules vianon-covalent bonds is determined by the iodine releasing from polymer. The bactericide claimed is prepared by cross-linking of microgranulated native potato starch with epichlorhydrine (y = 0.05) followed by etherification with glycidylmethylammonium chloride and iodination in aqueous solution with monovalent anions of iodine derivatives. The bactericidal product with improved efficiency is cross-linked N-(2-hydroxy)propyl-3-trimethylammonium starch chloride having substitution degree 0.14 « x« 0.3.@

Description

The invention relates to the field of chemistry and can be used in the production of a polymeric bactericide for the disinfection of water and biological fluids.

The starting material for polymeric bactericides in which the molecular iodine is attached to the polymer macromolecules by non-covalent linkages can be synthetic and natural polymers. They incorporate iodine into the polymer matrix by treatment with iodine vapor, solutions in organic solvents or aqueous solutions of anion (triiodide, pentajodide) of monovalent iodine derivatives. Their antibacterial activity is determined by the molecular iodine released from the polymer into the environment.

Among the most effective and widely used iodophores are complexes of styrendivinylbenzene anion exchangers with ions of monovalent iodine derivatives [J. Mills. U.S. Patent 3462363, 210/62, 1969; J. Lamber, L.Fina U.S. Patent 3923665, 424/79, 1975; P-J. Messier. U.S. Patent 5,639,452, 424 / 78.1, 1997], polyvinylpyrrolidone-iodine charge transfer complexes [E. Shanbrom, U.S. Patent 5,360,605, 424 / 78.08, 1994; E. Shanbrom et al., U.S. Patent 5,370,869, 424 / 78,22, 1994], polydialyldimethylammonium iodide [J. Barkauskaitė and others. LT patent 4497B, C08F126 / 02, 1999]. The raw materials for the aforementioned iodophores are carbohydrate biodegradable polymers. This property limits the use of the mentioned polymeric iodophores in modern water and biological fluid decontamination technologies.

Organic and biodegradable raw materials for iodophores are natural polysaccharides and their ionogenic derivatives (chitosan, starch). Amylose iodine preparations containing 30 mg / g of starch iodine for medical purposes, obtained from amylose enriched starch isolated from genetically modified wheat grains [W.R. Ghent, B.A.Eskin, U.S. Patent 5,955,101, 424/451, 1999].

Soluble ionic chitosan-iodine complexes were obtained by reacting a polymer with anions of monovalent iodine derivatives in an acidified solution containing polyhydric compounds. The proposed antiseptic hydrogel, which contains the active ingredient iodine derivative of chitosanLT 5802 B, accelerates wound healing [E.M. Hassan. U.S. Patent 6521243, 424/404, 2003]. Iodine may be immobilized on water-dispersible, water-biodegradable cross-linked dextrins, dextran, sorbitol, polyvinyl alcohol or derivatives thereof containing diethylaminoethyl, carboxy groups [J.A.O. Johansson. U.S. Patent 4010259, 424/150, 1977]. These bactericides, belonging to the cadoxemero family, are used to kill bacteria and pathogens from protein solutions [E. Shanbrom, U.S. Patent 5,814,225, 210/656, 1998; E. Shanbrom et al., U.S. Pat. No. 6096216, 210/638, 2000], or blood [S. J. Miekka et al., U.S. Pat. No. 6,106,773, 422/28, 2000] in the treatment of infectious diseases because they release molecular skin differently from the polyvinylpyrrolidone-iodine complex.

The prototype of the present invention is an ionic complex of triiodide and a cross-linked N- (2-hydroxyl) propyl-3-trimethylammonium starch having a cationic exchange ratio of 0.32 [J. Bendoraitienė, R. Kavaliauskaitė, N. Plačenytė, V. Valikonytė, A. Žemaitaitis, Antimicrobial Effect of Cationic Starch and Its Iodine Compounds // Chemical Technology ISSN 1392 - 1231, 2007. No. 2 (44), p. 61-69],

It is an object of the present invention to provide crosslinked N- (2-hydroxyl) propyl-3-trimethylammonium starch triiodide complexes having improved bactericidal activity.

The present invention relates to an insoluble crosslinked N- (2-hydroxyl) propyl-3-trimethylammonium starch triiodide starting material - a crosslinked N- (2-hydroxyl) propyl-3-trimethylammonium starch chloride having a degree of substitution of 0.14 <x <0.3 by N- (2-hydroxyl) propyl-3-trimethylammonium groups per anhydroglucose unit (AGL).

Only in such a raw material is the molecular iodine incorporated into several molecular iodine complexes: ionic cationic starch-triiodide (TKK-triiodide) and insertion amylose iodine (TK-iodine) complexes. This is evidenced by the varying degrees of substitution of the cationic starch suspension particles in iodine solution. The cross-linked starch acquires a blue color of the amylose-iodine complex, the cationic starch having a PL> 0.32 is a yellow-brown color. The color of the crosslinked N- (2-hydroxyl) propyl-3-trimethylammonium starch chloride having a degree of substitution of 0.14 <x <0.3 according to the N- (2-hydroxyl) propyl-3-trimethylammonium groups is determined by both complexes. Table 1 shows the abbreviated designation of substances.

table. Cationic starch and its complexes with iodine

Starch derivative

Notation

Tinned ^ .0 starch

OH

N ⁺ cr l \

TKKpl = x chloride

OH

Crosslinked ^ .0 starch

where: x - degree of cationic starch substitution (PL) according to N (2-hydroxy) propyl-3 IPCC = L complex of trimethylammonium

The bactericide is obtained by cross-linking natural potato starch in the form of microgranules with epichlorohydrin (EPH) followed by cationization of glycidyltrimethylammonium chloride (GTAC) and iodination of monovalent iodine derivatives (Il _n ) 'in anionic solution as follows:

A well-suspended aqueous solution of epichlorohydrin (0.05 mol / anhydroglucosidic residue (AGL)) is added to a 50% starch-water suspension alkaline with NaOH solution to pH = 11. The reaction mixture was maintained at 45 ° C for 24 h. At the end of the reaction, the cross-linked starch (TK) was washed with copious amounts of water and dried at room temperature;

N- (2-hydroxyl) propyl-3-trimethylammonium starch chloride (IPC) was obtained by etherification of the cross-linked starch with glycidyltrimethylammonium chloride (GTAC) in alkaline medium. The cationization mixture is prepared by mixing NaOH and GTAC solutions, adding TK to such mixture and mixing thoroughly. The molar ratio of reactants in the reaction mixture was AGL: GTAC: NaOH: H2O = 1: z: 0.04: w. Here, z and w, respectively, are the number of moles of GTAC and water added to the reaction mixture. The reaction was carried out under heterogeneous conditions at 45 ° C for 24 h. After the reaction, the crosslinked N- (2-hydroxyl) propyl-3-trimethylammonium starch (IPC) chloride is washed with water and filtered.

Iodine is used in iodine solution with a molar L / KI ratio of 3. The cationic starch-triiodide bactericide is prepared by adding 550 ml of 0.05 M iodine solution to 10 g of TCB. The suspension is kept at 20-25 ° C for 30 min periodically with the degree of substitution (PL) from the formula:

1400-1 S. V 'stirring. The TKK-triiodide complex and the remaining solution are separated by filtration, the bactericide washed well with water and dried at 40 ° C.

Determination of cationic groups in the IPCC.

Prior to analysis, cationic starch samples were extracted with methanol in a Soxhlet apparatus for 24 h. After determination of the nitrogen content (N,%) in the sample by the Kjeldahl method, the content of 2-hydroxypropyltrimethylammonium in the modified starch is calculated as: N,% is the nitrogen content of the sample, 162 is the mass of AGL.

Determination of iodine content in TKK-iodine complex.

The amount of iodine incorporated in iodophor is determined by the standard procedure using sodium thiosulphate [K. Dunkas. Quantitative analysis. Vilnius, 1961.]

Determination of antibacterial activity of IPC-iodine complex

Cultures of bacteria, including pathogenic ones, were used to assess antibacterial activity. Selected test cultures: Listeria monocytogenes, ATCC 19117, Pseudomonas aeruginosa, ATCC 27853, Salmonella typhimurium (ATCC 14028), Staphylococcus aureus (ATCC 25923), Escherichia coli (ATCC 25922), Bacillus subtilis (spores and vegetative) (ATCC 29212). Surface water containing coliform and heterotrophic bacteria was used.

Bacterial cultures were grown at 37 ° C on sloping agar for 18 h to assess antibacterial activity by agar diffusion method according to LST EN ISO 20645: 2005. The washed bacterial suspension was diluted according to Mc Farland standard no. 0.5, well mixed in a mini-shaker and added to the reconstituted and cooled to 47 ° C agar medium for total bacterial count. Once again, well mixed until uniform distribution of cells. The suspension of bacterial cells and medium prepared in this way was dispensed into 10 ml 90 mm glass Petri dishes. When the medium was stationary, wells were made (8 mm diameter) containing samples of test substance and 50 μl of sterile distilled water. The antimicrobial activity of bacterial cultures was evaluated in millimeters after incubation at 37 ° C for 24 or 48 hours according to the diameter of the clear areas around the wells. The width (H) of the damping zone is measured in millimeters. Where the attenuation zone is not a circle but an oval, the maximum and minimum widths of the zone are measured and their arithmetic mean calculated [LST EN ISO 20645: 2005 en]. After measuring the entire diameter and knowing the diameter of the well, the width of the damping zone is calculated using the formula:

D - d where: H - Width of damping zone, mm; D - total diameter of sterile zone and well, mm; d is the diameter of the well, mm

The antimicrobial efficacy data of cationic starch-triiodide bactericides are shown in Tables 2 and 3. It follows that the crude N- (2-hydroxyl) propyl-3-trimethylammonium chloride feedstock having a degree of substitution of 0.14 <x <0.3 according to the N- (2-hydroxy) propyl-3-trimethylammonium groups gives bactericides have a higher bactericidal activity than the prototype material.

table. Effect of cationic starch-triiodide bactericides on growth of coliform and heterotrophic bacteria by agar diffusion method according to LST EN ISO 20645: 2005

Cationic starch and triiodide complex Inhibition of growth of coliform bacteria area size, mm Growth of heterotrophic bacteria size of damping zone, mm after 24 h after 48 h after 24 h after 48 h TKKpL = c, 3, Example 2 43 45 36 40 TKKpl = o, 32, prototype 40 42 30th 30th

Note: 5 mg each was used to evaluate bactericidal effects. samples of the test substance

The invention is illustrated by the following examples Example 1 (prototype).

99% epichlorohydrin (Sigma-Aldrich, Germany) was added to 50% starch (SP AB "Stumbras" Antanavas, Lithuania) - aqueous suspension, which was basified to pH = 11 with NaOH solution. The amount of epichlorohydrin was 0.05 mol / anhydroglucosidic residue (AGL). The reaction mixture was maintained at 45 ° C for 24 h. After completion of the reaction, the cross-linked starch (TK) was washed with copious amounts of water and dried at room temperature. N- (2-hydroxyl) propyl-3-trimethylammonium starch (TKK) chloride was obtained by etherification of the cross-linked starch in a 70% solution of glycidyltrimethylammonium chloride (GTAC) (Fluka, Germany) under alkaline conditions. The cationization mixture is prepared by mixing NaOH and glycidyltrimethylammonium chloride (GTAC) solutions and adding TK to such mixture and mixing the components thoroughly. The molar ratio of reactants in the cationization mixture was AGL: GTAC: NaOH: H ₂ O = 1: 0.33: 0.04: 8. Here, 0.33 is the number of GTAC moles added to the reaction mixture. The reaction was carried out under heterogeneous conditions at 45 ° C for 24 h. After the reaction, the crosslinked N (2-hydroxyl) propyl-3-trimethylammonium starch (TKK) chloride is washed with water and filtered. The degree of substitution of the resulting IPC chloride is 0.32 (IPCsl = o, 32) ·

The cationic starch-iodine complex is prepared by adding 550 g of 0.05 M iodine solution to 10 g of TCA. The suspension is kept at 20-25 ° C for 30 minutes with periodic stirring. The TKK-triiodide polycomplex and the remaining solution are separated by filtration. The microgranules of the TKK-triiodide polycomplex are well washed with water and dried at 40 ° C. The iodine content of the cationic starch bactericide is 28%. The bactericidal activity data are presented in Table 2.

example.

The iodinated cationic starch is prepared as in the first example, except that the mole ratio of the reactants in the cationization mixture was AGL: GTAC: NaOH: H ₂ O = 1: 0.34: 0.04: 3. The resulting TKK chloride has a degree of substitution of 0.30 (TKKsl = o, 3o) · The iodine content of the cationic starch is 27%. The bactericidal activity data are presented in Tables 2 and 3.

example.

The iodinated cationic starch was prepared as in the first example, except that the molar ratio of reactants in the starch cationization mixture was AGL: GTAC: NaOH: H? O = 1: 0.71: 0.04: 3. The resulting TKK chloride has a degree of substitution of 0.62 (TKKsl = o). , 62) · Iodine content of cationic starch bactericide is 38%. The bactericidal activity data are presented in Table 3.

example.

Iodinated cationic starch is prepared as in the first example, except that the molar ratio of reactants in the starch cationization mixture was AGL: GTAC: NaOH:

H2O = 1: 0.24: 0.04: 3. The resulting TKK chloride has a degree of substitution of 0.21 (TKKsl = o, 2i) · The iodine content of the cationic starch bactericide is 21%. The bactericidal activity data are presented in Table 3.

example.

The iodinated cationic starch is prepared as in the first example, except that the molar ratio of reactants in the starch cationization mixture was AGL: GTAC: NaOH: H? O = 1: 0.16: 0.04: 5. The resulting TCC chloride has a degree of substitution of 0.14 (TCCsl = o). , i4) · The iodine content of the cationic starch bactericide is 20%. The bactericidal activity data are presented in Table 3.

example.

The iodinated cationic starch was prepared as in the first example, except that the molar ratio of reactants in the starch cationization mixture was AGL: GTAC: NaOH: H2O = 1: 0.05: 0.04: 8. The resulting TCC chloride has a degree of substitution of 0.04 (TCC1 = o, O4). ). The cationic starch bactericide content is 10%. The bactericidal activity data are presented in Table 3.

table. Antimicrobial efficacy of cationic starch and iodine complexes in bacterial growth by diffusion to agar method according to LST EN ISO 20645: 2005

sauaSojfoouoiu DUaJSig 12.00 ± 0.00 29.00 ± 0.81 28.00 ± 0.81 14.00 ± 0.00 i 14.00 ± 0.00 O O o O \ o O o m o CN o -H o -H -H o -H so -H cn o o o cn CN CN in cc c *? CN CN O O O O o o o O o o o θ ' o * o o s s i s s o o o o o snaMV snoaūooifydvis is? cn θ ' O Tf c? Etc. θ ' in o 00 O O o o o umumutųdy CN -H θ ' • + I o • 'rt' -H O C? £ o m o o o Ol [3UOUfIOS r- CN CN cn • Π CN r- χζ o. oo vsouiSntaO θ ' -H ΠΊ CN -H Ό C? -H O 0.00 o Ό o -H SOUouiopnasj 00 ΓΏ CO σ? <o O \ m O o o so O 00 00 o (soMds) S θ ' s CN -H m c? -H o o i o o m o o stjJiqns snitch V © CN o m <? cn CO CN CD o O \ Os o o m (enrolled +1 o 0 ± 0. if r- cn • Ή CD iJ Ό §3x) Skirt snÄpsag o o • * 3 · cc k © SO CN is? co oo * ' V? cn C <? CN o t> o o tJ- C? C? O -H -H O s o O O O o SJV33O / Sn33030A3} Ug θ ' C **) K? ΟΊ CD Linen and iodine ksas Vi S S Ό Vi Ό V5 «3 Ό Ό O CN Ό O · —J cn O • ”“> CN O O • “i S §0-2 o Ό O * II L = 0 O Λ o Λ <o Λ go to * ft. 0h eu 0- -4 μ — i · —I H H P P P o 2 -2 .§ £ $ · m CN o u 5-> 25 {X «i - a. 'Rt-

Note: The iodine content of the sample is 1,1 mg. Growth inhibition zone (mm) after 24 h incubation.

Claims (1)

Definition of the Invention Insoluble in cross-linked epichlorohydrin (y = 0.05), a cationic starch-triiodide bactericide in which the triiodide anions are complexed with the modified polysaccharide macromolecules [CHOrOEl fO-CH, -CH-CH, -O) (O-CH _f CH-CH.N (CHA) 1 xnH ^ϋ fi · 3-2y-x J ** v ^ώ I ~ OH OH characterized in that the highly effective bactericidal feedstock is cross-linked N- (2-hydroxyl) propyl-3-trimethylammonium starch chloride with a degree of substitution of 0.14 <x <0.3 ·