GB2432528A

GB2432528A - Use of flavonoids and fruit acid as anti-viral compositions

Info

Publication number: GB2432528A
Application number: GB0524375A
Authority: GB
Inventors: Ian Ripley
Original assignee: Individual
Current assignee: Individual
Priority date: 2005-11-29
Filing date: 2005-11-29
Publication date: 2007-05-30
Anticipated expiration: 2025-11-29
Also published as: GB0524375D0; GB2432528B

Abstract

The composition comprises flavonoids, preferably flavanone glycosides (bioflavonoids) and fruit organic acids, preferably ascorbic, malic or citric acids, which may be partially neutralised with 2-hydroxyethyltrimethyl ammonium hydroxide (choline) base. Preferably the flavonoid and acid are dissolved in a non-toxic solvent mixture comprising water and a co-solvent e.g. glycerine, polyhydric alcohols. A preferred embodiment comprises 15% soluble bioflavonoid, 15% citric acid, 15% malic acid, 5% ascorbic acid, 15% aqueous solution choline (45%n solution), 15% glycerine, 20% water.

Description

Novel Liquid Corn position In J'itro BIolo2jcaJJy Active To Viral

Infections Viral me4hanjs,,,s Preliminary work by an independent laboratory (2) testing a number of non-enveloped viruses (e.g. avian influenza virus H5N1) against our novel liquid composjtio has shown significant log reductions in viral count (Table 1).

The mechanism, which underpins the effectiveness of our innovative approach, is that non-enveloped viruses gain entry to their host cells by endocytosis, a process which requires an acidic intercellular compartment for penetration into the cytoplasm. There follows a step called "uncoating" in which the genetic information (RNA) of the virus is released. The RNA is used to produce new viral components in the nucleus of the cell and the cytoplasm Many new viruses begin to form at the cell membranes, an enzyme neuramjnj&,se' -a protein on the surface of the virus -enables the virus to cut itself off the membrane and so be released from the cell.

The virus now infects other cells and the replication is repeated. The death of cells and the immune response lead to typical flu symptoms.

The novelty of our system for in vitr9 applications is that the bioflavonoi wrap around and encapsulate the virus, thus very effectively preventing the spread of viral infection.

Endocytosis is obligatory in order for non-enveloped viruses to penetrate the host cells (3) and a specific study on canine parvovirus (I) proved that the mechanjs of endocytosis is pH dependent.

2) DESRIPTION -Relates to a non-toxic composition manufactured by a novel method which can be used for antiviral in vitro applications. The invention relates to a liquid "supplement" that includes a mixture of flavanone glycosides (also generally classified as "bioflavonoids"), and fruit organic acids which are partially neutralised with a Choline (2hydroxylethY1-trimethYl ammonium hydroxide) base.

More specifically this invention relates to mixture of soluble flavanone glycosides and organic acids dissolved in a non-toxic solvent mixture comprising of water and a water diluteable co-solvent (glycerine, a polyhydric alcohol, has been selected on toxicity grounds) but in cases where toxicity is not an issue other water soluble functional materials, such as aldehydes, ketones, alcohols and other water soluble reagents including nitrogen containing compounds such as amines and hydroxylamines, may be used.

Prior Art

We have noted that a preparation of water insoluble flavones, i.e., aglycones which are flavones free of glycoside, in low concentrations combined with organic acids such as citric acid, malic acid and ascorbic acid, plus a very large concentration of glycerol (more than 75 wt%), has been promoted for the commercial treatment of topical bacterial infections on plant food and food animal surfaces (4). The amount of ascorbic acid in the formulation caused it to discolour on standing. Substantial dilution of the water-soluble high glycerol concentrate with more citric acid and glycerol, plus a large amount of water, is carried out before the mixture is used for topical treatment of solid surfaces. It is not known to have any in vitro antiviral properties but challenged according to the BE EN 1276 protocol it gives satisfactory microbacterial results of greater than a 5 log reduction.

Because of the discoloration problem, that formulation was reformulated to remove ascorbic acid. The new formulation added propiomc acid to this very dilute solution but when tested against human cells was found to be a toxin.

Patent DE 4340438 discloses the use of bio-flavonoids, especially rutin for retrovirus inactivation, again DE 4339486 -discloses the use of bioflavonoids such as rutin and quercetin for retrovirus inactivation.

The distinguishing feature between the present application and the closest prior art is an inventive step represented by a "synergistic solubility" effect, not previously reported in the literature, whereby the spread of viral infection is prevented, not by inhibiting the viral neuraminidase (5), but by encapsulating the virus with bioflavonoids (flavones). The glucoside derivatives have the ability to breakdown on demand, releasing the flavanones and glucoside fragments as independent components. Only the flavanone fragments together with choline ascorbate are involved in the viral deactivation process.

3) Composition 3.1 Specifically the range of preferred active compounds should not exceed the following maximum / minimum values but in any case should not in total constitute more than 80% of the blended mixture (i.e.) the solvent content should be a minimum of 20%; 1 -20 % of bioflavonoids 1 -20 % of citric acid Active 1 -20 % of malic acid Components 1 -20 % of ascorbic acid 1 -20 % of Choline hydroxide % of glycerine / water (solvent) 3.2 The preferred ratio of components are; % soluble bioflavonoids % citric acid % malic acid % ascorbic acid (vitamin C) % of a 45% aqueous solution of Choline hydroxide % glycerine % water * The preferred soluble flavonoids are mixtures of soluble flavanone glycosides (i.e.) typically composed of, but not restricted to, a ratio of; naringin 40% -60%, hesperidin 20% -40%, and neohesperidin 5% -15%, plus other minor soluble flavanone components (maximum content 5%) such as, but not limited to didymin, neoeriocitrin, poncirin as extracted from citrus fruits, preferably bitter oranges.

Typical Flavanone Glycoside Structure H<VCY * Choline salt may be any Choline base but preferably Choline hydroxide.

* The best optional antiviral properties are obtained by neutralisation of the ascorbic acid component with Choline hydroxide.

The preferred formulated product (as noted in 3.2) is manufactured to the following procedure: Part (a) Blend water, glycerine and ascorbic acid together at ambient temperature -Increase the temperature to 50 C -add Choline hydroxide to neutralise the ascorbic acid Initial pH 1.2 Final pH 5.5 -6.0 The neutralisation temperatures should not be less than 25 C or greater than 55 C Optima! results were obtained at 50 C without any significant thermal degradation of the ascorbic acid which occurs rapidly at temperature values greater than 55 C. Part (b) Add remaining acids, (citric and malic) and finally the soluble bioflavonoids. (typical pH value of bioflavonoidS = 4.7) InitialpH 5.5-6.0 FinalpH 2.3-3.5 4) Results -based on neutralised variations of the preferred ratio of components as outlined in 3.2 Comparison of viral properties (avian influenza (H5N 1) on part neutralised, fully neutralised (all acids) and un-neutralised acids showed log reduction as noted in Table 1.

Table 1

Optimal Log Contact time/mm Reduction pH % kill 1. Ascorbic acid neutralised 5 5.7 2.3 99.9998 (all acids present) 2. Citric acid neutralised 5 3.5 3.0 99.00 (all acids present) 3. Malic acid neutralised 5 3.5 3.0 99.00 (all acids present) 4. Fully neutralised 5 4.0 6.5 99.99 (all acids present) 5. Un-neutralised 5 3.5 2.0 99.00 (all acids present) 6. Ascorbic acid only 5 3.0 2.5 97.90 (no other acids present) 7. Choline ascorbate only 5 3.5 6.5 99.00 (no other acids present) Bioflavonoids (typical pH 4.7) present in formulations 1 -7 above.

Claims

CLAIMS

What we claim is an antiviral composition based on: (i) A mixture of soluble bioflavonoidS from the following main groups; -FlavanoneS -Flavones -Flavanols Other classes of "bioflavoflOids" we wish to cover in the application includes, but is not limited to, anthocyanidins, proanthocyaflidins, procyanidolic oligomers, catechins, biflavans, polyphenols, rutinosides.

But preferably water-soluble glyco side derivatives from the flavanone group.

(ii) The soluble flavanone glyco sides mixed with organic acids of natural or synthetic origin, preferably fruit acids which contain a low order of toxicity -specifically citric, malic and ascorbic acids.

(iii) Partial neutralisation of the ascorbic acid with Choline hydroxide to give optimum results on H5N1 kill rates for any given ratios of the flavonoids and acids.(Table 1.) Note: -Similar results have been achieved on urbani SARS, Human Immunodeficiency Virus (HIV), Human influenza type A, and human Rhinovirus.

References 1 Suikken et al 2002 2 Test Laboratory details 3 Brabec et al 2003 4 BS EN 1276/ EN 1276: 1997 "Chemical disinfectants and antiseptics-Quantitative suspension test for the evaluation of bactericidal activity of chemical disinfectants and antiseptics used in food, industrial, domestic, and institutional areas Pharmasquare -Tamiflu Mode of Action