TECHNICAL FIELD
[001] The subject matter described herein in general relates to antibacterial compositions, and in particular, relates to the antibacterial compositions that are effective against Gram negative bacteria.
BACKGROUND OF INVENTION
[002] Antibiotics are the major drugs used for the treatment of infectious diseases caused by bacteria. In recent years, antibiotic resistance has been increasing, and is becoming a serious problem in infection control. The failure of the existing antibiotics to control infections makes it critical to find alternative agents which overcome the drawbacks of the synthetic antibiotics.
[003] Herbal alternatives are being increasingly used as desirable alternatives to curb the harmful effects of bacteria. For example, US9655838 discloses an antibacterial combination of essential oils comprising bisabolol and one or more essential oils selected from citral, carvacrol, oregano extract, and rosemary, or comprising carvacrol and one or more essential oils selected from bisabolol, thymol, and rosemary. WO2010091415 relates to a preservative or antimicrobial composition with broad spectrum antimicrobial activity comprising low concentrations of essential oil (and/or one or more component thereof) and a botanical extract in synergistic combination with a fruit acid and alkanediol, and optionally a solvent. US5658584 discloses an antimicrobial composition containing a mixture of hinokitiol and citronellic acid in a ratio of about 1:1 to about 3:1 by weight. The antimicrobial composition according to the invention is safe for humans and has a high antimicrobial activity and a broad antimicrobial spectrum [004] However, these compositions have met with minimal acceptance in the marketplace owing to poor microbicidal effects and similar instances/symptoms of skin irritancy experienced as with synthetic chemical agents. Considering the above evidence, there exists a need to develop herbal compositions that is devoid of synthetic ingredients, that is capable of imparting an antimicrobial effect.

SUMMARY OF THE INVENTION
[005] In an aspect of the present invention, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is in a range of 1:1 to 17:1.
[006] In another aspect of the present invention, there is provided a process for preparing the composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is in a range of 1:1 to 17:1, said process comprising: a) obtaining Acacia concinna extract; b) obtaining lactic acid; and c) contacting the Acacia concinna extract with lactic acid to obtain the composition.
[007] These and other features, aspects, and advantages of the present subject matter will be better understood with reference to the following description and appended claims. This summary is provided to introduce a selection of concepts in a simplified form. This summary is not intended to identify key features or essential features of the claimed subject matter, nor is it intended to be used to limit the scope of the claimed subject matter.
DETAILED DESCRIPTION OF THE INVENTION
[008] Those skilled in the art will be aware that the present disclosure is subject to
variations and modifications other than those specifically described. It is to be
understood that the present disclosure includes all such variations and
modifications. The disclosure also includes all such steps, features, compositions,
and compounds referred to or indicated in this specification, individually or
collectively, and any and all combinations of any or more of such steps or features.
Definitions
[009] For convenience, before further description of the present disclosure,
certain terms employed in the specification, and examples are delineated here.
These definitions should be read in the light of the remainder of the disclosure and
understood as by a person of skill in the art. The terms used herein have the
meanings recognized and known to those of skill in the art, however, for
convenience and completeness, particular terms and their meanings are set forth
below.

[0010] The articles “a”, “an” and “the” are used to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article.
[0011] The terms “comprise” and “comprising” are used in the inclusive, open sense, meaning that additional elements may be included. It is not intended to be construed as “consists of only”.
[0012] Throughout this specification, unless the context requires otherwise the word “comprise”, and variations such as “comprises” and “comprising”, will be understood to imply the inclusion of a stated element or step or group of element or steps but not the exclusion of any other element or step or group of element or steps.
[0013] The term “including” is used to mean “including but not limited to”. “Including” and “including but not limited to” are used interchangeably. [0014] Ratios, concentrations, amounts, and other numerical data may be presented herein in a range format. It is to be understood that such range format is used merely for convenience and brevity and should be interpreted flexibly to include not only the numerical values explicitly recited as the limits of the range, but also to include all the individual numerical values or sub-ranges encompassed within that range as if each numerical value and sub-range is explicitly recited. For example, a ratio range of about 1:1 to 17:1 should be interpreted to include not only the explicitly recited limits of about 1:1 to about 17:1, but also to include sub¬ranges, such as 1:1, 4:1, 8:1, 16:1 and so forth, as well as individual amounts, including fractional ratios, within the specified ranges, such as 4.33:1, and 8.6:1, for example.
[0015] The term “at least one” is used to mean one or more and thus includes individual components as well as mixtures/combinations.
[0016] The term “extract” refers to a preparation containing the active ingredient of a substance in concentrated form. Exacts may be in the form of solids, powders, and liquids.
[0017] A composition comprising “synergistic activity” or a “synergistic composition” is a combination of compounds which exhibits increased biological or functional activity as a non-linear multiple of the biological or functional

activity of the individual compounds. In other words, the combined biological or functional activity of two or more compounds being tested is significantly greater than the expected result based on independent effects of the compounds when tested separately. Synergy may be apparent only at some ranges or weight percentages.
[0018] Excipients are substances that serve as mechanisms to improve the delivery and the effectiveness of drugs. The term “lactic acid, or salts and derivatives thereof” refers to either lactic acid, or its derivative, or its salt. Any well-known salt or derivative can be considered to be covered by the term salts and “derivative thereof”.
[0019] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the disclosure, the preferred methods, and materials are now described. All publications mentioned herein are incorporated herein by reference.
[0020] The present disclosure is not to be limited in scope by the specific embodiments described herein, which are intended for the purposes of exemplification only. Functionally-equivalent products, compositions, and methods are clearly within the scope of the disclosure, as described herein.
[0021] As mentioned previously, conventionally used synthetic antimicrobials have side-effects such as, irritation on skin and mucous membrane of the user along with a threat of spreading antibiotic resistance. Although herbal alternatives are being increasingly used as a desirable alternative, so far, these compositions have met with minimal acceptance in the marketplace owing to poor microbicidal effect and similar instances/symptoms of skin irritancy experienced as with synthetic chemical agents. The present disclosure aims to solve the said problem by using a combination of Acacia concinna extract and lactic acid, which when used in defined w/w ratios is effective in providing a synergistic effect against microorganisms, particularly Gram-negative bacteria.

[0022] In an embodiment of the present disclosure, there is provided a composition
comprising: a) Acacia concinna extract; and b) lactic acid, or salts and derivatives
thereof, wherein the Acacia concinna extract, and lactic acid, or salts and
derivatives thereof w/w ratio is in a range of 1:1 to 17:1. In another embodiment of
the present disclosure, the Acacia concinna extract, and lactic acid, or salts and
derivatives thereof w/w ratio is in a range of 4:1 to 17:1.
[0023] In an embodiment of the present disclosure, there is provided a composition
comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia
concinna extract, and lactic acid w/w ratio is in a range of 1:1 to 17:1. In another
embodiment of the present disclosure, the Acacia concinna extract, and lactic acid
w/w ratio is in a range of 4:1 to 17:1.
[0024] In an embodiment of the present disclosure, there is provided a composition
comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia
concinna extract, and lactic acid w/w ratio is 4.16:1.
[0025] In an embodiment of the present disclosure, there is provided a composition
comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia
concinna extract, and lactic acid w/w ratio is 8.33:1.
[0026] In an embodiment of the present disclosure, there is provided a composition
comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia
concinna extract, and lactic acid w/w ratio is 16.67:1.
[0027] In an embodiment of the present disclosure, there is provided a composition
comprising a) Acacia concinna extract; and b) lactic acid, wherein the Acacia
concinna extract, and lactic acid w/w ratio is 2.08:1.
[0028] In an embodiment of the present disclosure, there is provided a composition
as described herein, wherein the Acacia concinna extract has a weight percentage
in a range of 0.3 – 3% with respect to the extract; and wherein lactic acid has a
weight percentage in a range of 0.03 - 0.4% with respect to the composition. In
another embodiment of the present disclosure, Acacia concinna extract has a
weight percentage in a range of 0.31 – 2.5% with respect to the extract; and
wherein lactic acid has a weight percentage in a range of 0.03 - 0.3% with respect
to the composition.

[0029] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is in a range of 1:1 - 17:1, and wherein the Acacia concinna extract has a weight percentage in a range of 0.3 – 3% with respect to the extract; and wherein lactic acid has a weight percentage in a range of 0.03 -0.4% with respect to the composition.
[0030] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is 4.16:1, and wherein Acacia concinna extract has a weight percentage in a range of 0.3 – 3% with respect to the extract; and wherein lactic acid has a weight percentage in a range of 0.03 - 0.4% with respect to the composition.
[0031] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is 8.33:1, and wherein Acacia concinna extract has a weight percentage in a range of 0.3 – 3% with respect to the extract; and wherein lactic acid has a weight percentage in a range of 0.03 - 0.4% with respect to the composition.
[0032] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is 16.67:1, and wherein Acacia concinna extract has a weight percentage in a range of 0.3 – 3% with respect to the extract; and wherein lactic acid has a weight percentage in a range of 0.03 - 0.4% with respect to the composition.
[0033] In an embodiment of the present disclosure, there is provided a composition as described herein, wherein the Acacia concinna extract comprises saponins having a weight percentage in a range of 1 – 4% with respect to the extract. In another embodiment of the present disclosure, the Acacia concinna extract comprises saponins having a weight percentage in a range of 2 – 2.5% with respect to the extract.

[0034] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is in a range of 1:1 - 17:1, and wherein the Acacia concinna extract comprises saponins having a weight percentage in a range of 1 – 4% with respect to the extract.
[0035] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is 4.16:1, and wherein the Acacia concinna extract comprises saponins having a weight percentage in a range of 1 – 4% with respect to the extract.
[0036] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is 8.33:1, and wherein the Acacia concinna extract comprises saponins having a weight percentage in a range of 1 – 4% with respect to the extract.
[0037] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is 16.67:1, and wherein the Acacia concinna extract comprises saponins having a weight percentage in a range of 1 – 4% with respect to the extract.
[0038] In an embodiment of the present disclosure, there is provided a composition comprising a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is 2.08:1, and wherein the Acacia concinna extract comprises saponins having a weight percentage in a range of 1 – 4% with respect to the extract.
[0039] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is in a range of 1:1 - 17:1, and wherein the Acacia concinna extract has a weight percentage in a range of 0.3 – 3% with respect to the extract, and wherein lactic acid has a weight percentage in a range of 0.03 to 0.4% with respect to the composition, and wherein Acacia concinna extract

comprises saponins having a weight percentage in a range of 1 – 4% with respect to the extract.
[0040] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is in a range of 1:1 - 17:1, and wherein the Acacia concinna extract has a weight percentage in a range of 0.31 – 2.5% with respect to the extract; and wherein lactic acid has a weight percentage in a range of 0.03 - 0.3% with respect to the composition; and wherein the Acacia concinna extract comprises saponins having a weight percentage in a range of 2 – 2.5% with respect to the extract.
[0041] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is 4.16:1, wherein the Acacia concinna extract has a weight percentage in a range of 0.31 – 2.5% with respect to the extract; and wherein lactic acid has a weight percentage in a range of 0.03 - 0.3% with respect to the composition; and wherein the Acacia concinna extract comprises saponins having a weight percentage in a range of 2 – 2.5% with respect to the extract.
[0042] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is 8.33:1, wherein the Acacia concinna extract has a weight percentage in a range of 0.31 – 2.5% with respect to the extract; and wherein lactic acid has a weight percentage in a range of 0.03 - 0.3% with respect to the composition; and wherein the Acacia concinna extract comprises saponins having a weight percentage in a range of 2 – 2.5% with respect to the extract.
[0043] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is 16.67:1, and wherein the Acacia concinna extract has a weight percentage in a range of 0.31 – 2.5% with respect to the extract; and wherein lactic acid has a weight percentage in a range of 0.03 -

0.3% with respect to the composition; and wherein the Acacia concinna extract comprises saponins having a weight percentage in a range of 2 – 2.5% with respect to the extract.
[0044] In an embodiment of the present disclosure, there is provided a composition comprising: a) Acacia concinna extract; and b) lactic acid, wherein the Acacia concinna extract, and lactic acid w/w ratio is 2.08:1, and wherein the Acacia concinna extract has a weight percentage in a range of 0.31 – 2.5% with respect to the extract; and wherein lactic acid has a weight percentage in a range of 0.03 -0.3% with respect to the composition; and wherein the Acacia concinna extract comprises saponins having a weight percentage in a range of 2 – 2.5% with respect to the extract.
[0045] In an embodiment of the present disclosure, there is provided a process for preparing a composition comprising: a) Acacia concinna extract; and b) lactic acid; wherein the Acacia concinna extract, and lactic acid w/w ratio is in a range of 1:1 to 17:1, said process comprising: a) obtaining the Acacia concinna extract; b) obtaining lactic acid; and c) contacting the Acacia concinna extract with lactic acid to obtain the composition.
[0046] In an embodiment of the present disclosure, there is provided a process for preparing a composition comprising: a) Acacia concinna extract; and b) lactic acid, or salts and derivatives thereof; wherein the Acacia concinna extract, and lactic acid, or salts and derivatives thereof w/w ratio is in a range of 1:1 to 17:1, said process comprising: a) obtaining the Acacia concinna extract; b) obtaining lactic acid, or salts and derivatives thereof; and c) contacting the Acacia concinna extract with lactic acid, or salts and derivatives thereof to obtain the composition. [0047] In an embodiment of the present disclosure, there is provided a process for preparing a composition comprising: a) Acacia concinna extract; and b) lactic acid; wherein the Acacia concinna extract, and lactic acid w/w ratio is in a range of 1:1 to 17:1, and wherein the Acacia concinna extract has a weight percentage in a range of 0.3 – 3% with respect to the extract; and wherein lactic acid has a weight percentage in a range of 0.03 to 0.4% with respect to the composition, said process comprising: a) obtaining the Acacia concinna extract; b) obtaining lactic acid; and

c) contacting the Acacia concinna extract with lactic acid to obtain the composition.
[0048] In an embodiment of the present disclosure, there is provided a process for preparing a composition comprising: a) Acacia concinna extract; and b) lactic acid; wherein the Acacia concinna extract, and lactic acid w/w ratio is in a range of 1:1 to 17:1, and wherein the Acacia concinna extract has a weight percentage in a range of 0.3 – 3% with respect to the extract; and wherein lactic acid has a weight percentage in a range of 0.03 to 0.4% with respect to the composition, and wherein the Acacia concinna extract comprises saponin having a weight percentage in a range of 1 – 4% with respect to the extract, said process comprising: a) obtaining the Acacia concinna extract; b) obtaining lactic acid; and c) contacting the Acacia concinna extract with lactic acid to obtain the composition.
[0049] In an embodiment of the present disclosure, there is provided a process for preparing a composition comprising: a) Acacia concinna extract; and b) lactic acid; wherein the Acacia concinna extract, and lactic acid w/w ratio is in a range of 1:1 to 17:1, and wherein the Acacia concinna extract has a weight percentage in a range of 0.31 – 2.5% with respect to the extract, and wherein lactic acid has a weight percentage in a range of 0.03 - 0.3% with respect to the composition, and wherein the Acacia concinna extract comprises saponin having a weight percentage in a range of 1 – 4% with respect to the extract, said process comprising: a) obtaining the Acacia concinna extract; b) obtaining lactic acid; and c) contacting the Acacia concinna extract with lactic acid to obtain the composition.
[0050] In an embodiment of the present disclosure, the composition inhibits the growth of E. coli.
[0051] In an embodiment of the present disclosure, the composition as described herein is an antibacterial composition. The applications include, personal care products such as shampoos, cosmetics, and soaps, and the like.
EXAMPLES
[0052] The disclosure will now be illustrated with working examples, which is intended to illustrate the working of disclosure and not intended to take restrictively to imply any limitations on the scope of the present disclosure. Unless

defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of ordinary skill in the art to which this disclosure belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice of the disclosed methods and compositions, the exemplary methods, devices and materials are described herein. It is to be understood that this disclosure is not limited to particular methods, and experimental conditions described, as such methods and conditions may apply. [0053] Conventionally used antibacterials involve the use of antibiotics which are associated with several side effects. Although herbal alternatives such as essential oils are being increasingly used as desirable alternatives, these oils are known to cause irritation on skin and mucous membrane of the user. Further, these compositions have met with minimal acceptance in the marketplace owing to the poor microbicidal effects and narrow spectrum of antimicrobial activity. The present disclosure aims to solve the said problem by using a combination of Acacia concinna extract and lactic acid which when used in defined w/w ratios is effective in providing a synergistic effect against bacteria, particularly Gram-negative bacteria.
Materials and methods
[0054] In order to test the antimicrobial activity of the actives, Acacia concinna
extract was procured from Harika Chemicals (Product number-8906070780946).
Although any plant part may such as leaf, flower, root, stem, fruit can be used to
prepare the extract, the composition present disclosure uses fruit extract of Acacia
concinna. Anhydrous lactic acid was procured from Fluka Analytics, catalog
number-69775.
Example 1: Preparation of the inoculum
[0055] 10% Acacia concinna solution was prepared by dissolving 10g of Acacia concinna extract in 100ml of water. A working stock solution of 5% Acacia concinna extract (A) was prepared in Luria Bertani (LB) broth in a 96 well plate. The lower concentrations were obtained by 2-fold serial dilutions. These concentrations were tested in combination with lactic acid (L) (0.15% and 0.075%). Combinations were prepared by simple mixing and ensuring complete

mixing. The combinations tested were A+L (0.625%+0.15%), A+L
(1.25%+0.075%), A+L (0.625%+0.075%). E. coli – DH5 alpha and S. aureus –
ATCC25923 inoculum of 106 CFU/mL density was added to the wells, such that
the total volume in each well is 200uL. LB broth with the inoculum alone served as
the growth control. Separate media blanks (without the inoculum) was maintained
for each active and the combinations. The wells were incubated at 37°C with
shaking for 18 hours and their optical density (OD) at every 10 minutes were read
using a spectrophotometer. From the data collected the average lag phase, doubling
time and the maximum OD relative to control was calculated; and the results are
presented below from Table 1 – 4.
[0056] Table 1 depicts the lag delay of E. coli in presence of Acacia concinna
extract (A), and lactic acid (L), alone and in combination. Lag delay is the period
between the introduction of a microorganism into a culture medium and the time it
begins to increase exponentially. A short lag phase is indicative of rapid
multiplication of microorganisms.
Table 1
A: L Expected
ratio lag delay
(w/w) Lag delay relative
Concentration Average relative to to
Active (%) lag (h) control control
Control 4.00
A 1.25 4.29 0.29
A 0.625 4.00 0.00
A 0.3125 3.50 -0.50
L 0.075 4.25 0.25
L 0.15 4.88 0.88
A+L 1.25+0.075 16.67:1 7.71 3.71 0.54
A+L 0.625+0.15 4.16:1 10.67 6.67 0.88
A+L 0.625+0.075 8.33:1 4.38 0.38 0.25
[0057] As can be observed in Table 1, the average lag with Acacia concinna extract, and lactic acid, at all concentrations tested, was found to be comparable with the control. However, unexpectedly, and surprisingly, for the various combination ratios tested, it can be observed that the lag phase for the combination ratios 4.16:1 and 16.67:1 was found to be 10.6 hours, and 7.7 hours respectively,

which is much higher to any of the individual concentrations tested. Although an
increase in lag phase is observed for all the combination ratios tested, a synergistic
increase in lag phase is observed particularly at w/w ratio of 4.16:1. This delay in
lag phase relative to the control suggests that the rate of bacterial growth has been
synergistically reduced with the composition of the present disclosure, particularly
at w/w ratio of 4.16:1.
[0058] Table 2 depicts the lag delay of S. aureus in presence of Acacia concinna
extract (A), and lactic acid (C), alone and in combination.
Table 2
A:L ratio Lag delay
Concentration (w/w) relative to
Active (%) Average lag (h) control
Control 4.00
A 1.25 4.67 0.67
A 0.625 3.92 -0.08
L 0.075 4.33 0.33
L 0.15 17.25 13.25
A+L 1.25+0.075 16.67:1 8.75 4.75
A+L 0.625+0.15 4.16:1 17.33 13.33
A+L 0.625+0.075 8.33:1 4.83 0.83
[0059] As can be observed in Table 2, the lag delay relative to the control was substantially higher with 0.15% of lactic acid, and for the combination ratio’s 16.67:1 and 4.16:1. However, no synergistic increase in lag was observed for either of these ratio’s. Although a significant increase in lag was observed for the combination ratio of 16.67:1, however, combination at the ratio of 4.16:1 was comparable to that of lactic acid. This suggests although the combination is effective against S. aureus, a synergistic effect in terms of bacterial (S. aureus) growth reduction was not observed with the composition of the present disclosure. [0060] Table 3 depicts a doubling time of E. coli in presence of Acacia concinna extract, and lactic acid, alone and in combination. Doubling time is the time taken by the bacteria to double in number during a specified period. It varies with different organisms. For example, E. coli divides around every 20 minutes, hence its doubling time is 20 minutes, and for Staphylococcus aureus it is 30 minutes. Table 3

A:L Average
Concentration ratio doubling
Active (%) (w/w) time
Control 34.48
A 1.25 37.22
A 0.625 32.31
A 0.3125 32.29
L 0.075 39.08
L 0.15 58.10
A+L 1.25+0.075 4.16:1 130.29
Growth
A+L 0.625+0.15 16.67:1 inhibition
A+L 0.625+0.075 8.33:1 39.15
[0061] From Table 3 it can be observed there is a significant lag in doubling ti in comparison to the control for all the combination ratios tested. It can be furth observed that the rate at which the bacteria multiplied was about 4 times slow than the control for the combination (A:L) ratio 4.16:1; and a complete inhibiti in bacterial growth was observed for the combination (A:L) ratio 16.67:1. A high average doubling time is indicative of slow multiplication, thereby suggesting th the composition of the present disclosure is effective in reducing the bacter growth (E.coli) at defined w/w ratios.
[0062] Table 4 depicts a doubling time of Acacia concinna extract, and lac acid, alone and in combination against S. aureus. Table 4
A:L ratio
Concentration Average
(w/w)
Actives (%) doubling time
Control 34.31
A 1.25 42.94
A 0.625 26.71
67.03
L 0.075
Growth inhibition
L 0.15
4.16:1 186.11
A+L 1.25+0.075
16.67:1 Growth inhibition
A+L 0.625+0.15
8.33:1 53.67
A+L 0.625+0.075

[0063] As can be observed in Table 4, a complete growth inhibition was observed with 0.15% of lactic acid, and at 16.6:1 w/w ratio of Acacia concinna extract, and lactic acid.
[0064] Further, the effect of the actives, Acacia concinna extract, and lactic acid, alone and in combination at various w/w ratios was tested against E. coli, and the percentage inhibition in bacterial growth was calculated. The results are presented below in Table 5.
Actives Concentration (%) Ratio % inhibition
A 2.5 88
A 1.25 24
A 0.625 -1
A 0.3125 -10
A 0.015 -31
L 0.3 99
L 0.15 31
L 0.075 9
A+L 1.25+0.15 8.33:1 99
A+L 1.25+0.075 16.67:1 69
A+L 0.625+0.15 4.16:1 87
A+L 0.3125+0.15 2.08:1 75
A+L 0.015+0.3 0.05:1 100
A+L 2.5+0.075 33.3:1 99
[0065] For various combination ratios tested, it can be observed that a synergistic effect is observed for all the ratios ranging from 2.08:1 to 16.67:1. However, no synergistic effect was observed for the w/w ratios 0.05:1 and 33:3:1. The percentage inhibition exhibited by the individual actives (2.5% of the Acacia concinna extract and 0.075% of lactic acid) was found to be 88% and 9% respectively; the additive effect being 97%. However. when a combination of actives (2.5% of A+0.075% of L) was used, the percentage inhibition of bacterial growth was found to be 99%, which is comparable to the additive effect observed with the individual concentration of actives. Similarly, no synergistic effect was observed for the w/w ratio 0.05:1 (0.015% of Acacia concinna extract, and 0.3% lactic acid). The percentage inhibition exhibited with 0.3% lactic acid alone is 99%. No increase in percentage inhibition was observed when a combination of actives

(0.015% of Acacia concinna extract, and 0.3% lactic acid). The data suggests that the composition of the present disclosure in defined w/w ratios, i.e., 2.08:1 to 16.6:1 is very effective inhibiting the growth of E. coli. The percentage inhibition exhibited by the combination of actives at these w/w ratios is far greater than the additive effect exhibited by the individual concentrations of actives.
[0066] Overall, the data suggests that composition of the present disclosure shows an anti-bacterial activity against both E. coli (Gram negative) and S. aureus (Gram positive) microbes. However, a surprising antimicrobial synergy was observed only against E. coli at specific w/w ratios. No synergy was observed against S. aureus.
Advantages of the present disclosure
[0067] The present disclosure discloses a composition comprising herbal
ingredients that is effective against Gram-negative bacteria, particularly against E.
coli. The composition comprising Acacia concinna extract; and lactic acid in a
range of 1:1 to 17:1, was found to synergistically inhibit or slow down the growth
of bacteria, particularly Gram-negative bacteria. Also, since the composition is
devoid of any synthetic components, there are no side-effects associated with the
use of the composition.

I/We Claim:
1. A composition comprising:
a) Acacia concinna extract; and
b) lactic acid, or salts and derivatives thereof,
wherein the Acacia concinna extract, and lactic acid or salts, and derivatives thereof w/w ratio is in a range of 1:1 to 17:1.
2. The composition as claimed in claim 1, wherein the Acacia concinna extract, and lactic acid, or salts and derivatives thereof has a w/w ratio of 4.16:1.
3. The composition as claimed in claim 1, wherein the Acacia concinna extract, and lactic acid, or salts and derivatives thereof has a w/w ratio of 8.33:1.
4. The composition as claimed in claim 1, wherein the Acacia concinna extract, and lactic acid, or salts and derivatives thereof has a w/w ratio of 16.67:1
5. The composition as claimed in any of the claims 1-4, wherein the Acacia concinna extract has a weight percentage in a range of 0.3 – 3% with respect to the extract; and wherein lactic acid, or salts and derivatives thereof has a weight percentage in a range of 0.03 to 0.4% with respect to the composition.
6. The composition as claimed in any one of the claims 1 to 5, wherein the Acacia concinna extract comprises saponins having a weight percentage in a range of 1-4% with respect to the extract.
7. A process for preparing the composition as claimed in claim 1, said process comprising:

a) obtaining Acacia concinna extract;
b) obtaining lactic acid, or salts and derivatives thereof; and
c) contacting the Acacia concinna extract with lactic acid, or salts and derivatives thereof to obtain the composition.
8. The composition as claimed in any one of the claims 1 to 6, wherein said
composition inhibits growth of Gram-negative bacteria.

9. The composition as claimed in claim 8, wherein said composition inhibits growth of E. coli.