<% vol = 20 number = 1 prevlink = 35 nextlink = 44 titolo = "IN VITRO ANTI-STAPHYLOCOCCAL ACTIVITY OF HONEY AND TWO STANDARD ANTIBIOTICS (CLOXACILLIN AND AMPICILLIN)" volromano = "XX" data_pubblicazione = "March 2007" header titolo %>

Adeleke O.E.1, Anyacho C.E.2, Onakoya T.M.2

1 Department of Pharmaceutical Microbiology, Faculty of Pharmacy, University of Ibadan, Ibadan, Nigeria
2 Department of Microbiology, College of Natural Sciences, University of Agriculture, Abeokuta, Nigeria

SUMMARY. Fifty bacterial isolates comprising Staphylococcus aureus and Staphylococcus albus from six pathological sources were exposed to honey and its fractional dilutions as also as to two standard antibiotics - cloxacillin (5 mg/ml) and ampicillin (10 mg/ml) - in an agar-cup diffusion sensitivity test. The undiluted honey showed antibacterial activity against almost 100% of the bacterial isolates while its 1:2 to 1:10 fractional dilutions also had antibacterial property against the isolates in decreasing order of activity. Cloxacillin recorded higher antibacterial activity than ampicillin but both exerted less antibacterial activity than honey. The therapeutic application of honey, following clinical trials, should be of relevance in treating bacterial infections and, in particular, those due to the two Staphylococcus species tested.


Staphylococcus aureus and Staphylococcus albus have both been recognized as part of the normal microbial flora and persistent pathogens in man. More specifically, varieties of superficial and suppurative infections have been associated with S. aureus, while bacteraemia has been linked to S. albus following infections of cannulae, indwelling catheters, shunts, and other appliances positioned in the body.3 Pathogenic strains of S. albus have been reported4 to owe their virulence to the production of slime, which forms a biofilm on the surface of prosthetic devices, while S. aureus produces a number of enzymes and toxins, notable among which are coagulase, deoxyribonuclease, and enterotoxin B, which accounts for food-poisoning.

R plasmid genes mediating production of antibiotic-inactivating enzymes have been largely implicated in the resistance of S. aureus to antibiotics, particularly the b-lactams, including cloxacillin, cefuroxime, cefotaxime, methicillin, and vancomycin.4-6 S. albus has a record of 75% of its strains having resistance to naficillin and it can also sequester itself from the circulation and antimicrobial drugs when it occurs in prosthetic devices.

Honey is widely regarded as a food supplement owing to its greater rate of absorption than that of table sugar, its nutritive properties, and its easy digestibility.8-11 It is also a recognized income-earner.12 To this regard, harvesters, collectors, and hawkers of honey have significantly increased in our country, Nigeria.

Interestingly, honey has been associated with curative and antimicrobial effects. Heart and liver diseases, coughs, constipation, gastrointestinal disturbances, measles, ulcers, and wounds have all been treated with honey.13 Higher antibacterial activity against infections in burn wounds has been reported for honey than for the antibacterial ointment silver sulphadiazine.14 Laboratory isolates of S. aureus, Pseudomonas aeruginosa, and Escherichia coli, as also diarrhoeal bacterial isolates, had their respective growth inhibited by honey.

In environments where sanitation remains at a low ebb, S. aureus and S. albus are important environmental pollutants, especially in tropical developing countries. Hence the need for a readily available and affordable treatment for infections due to these organisms, in view of their resistance to the cheaper b-lactam antibiotics. This study reports on the comparative activities of honey and two standard antibiotics (ampicillin and cloxacillin) against two staphylococcal species from six pathological sources.

Materials and methods

Bacterial isolates

Pure cultures of bacterial isolates from different pathological sources (Table I) were obtained from the Routine Laboratory, Department of Medical Microbiology and Parasitology, University College Hospital, Ibadan, Nigeria. They were confirmed by conventional tests17 and preserved on tryptone soya agar (OXOID) slants in a refrigerator at 4 °C.

<% createTable "Table I ","The bacterial isolates and their pathological sources",";Pathological source; S. aureus; S. albus@;Eye swab; 8; 10 @;Wound swab; 13; 3@;Urine; 3; -@;Wound biopsy; 1; -@;Aspirate; -; 10@;Ear swab; -; 10@;Unidentified source; 5; 5 @;;30; 20","",4,300,true %>


Honey was obtained from a pure natural honey collection centre in Ibadan, South West Nigeria. It was used undiluted and also as fresh fractional dilutions in sterile distilled water (1:2, 1:4, 1:6, 1:8, and 1:10).

Ampicillin and cloxacillin (as sodium salt BP)

The two antibiotics were obtained in a local pharmacy store in Ibadan in ampoule vials as powder for injection. Cloxacillin was used at 5 mg/ml and ampicillin at 10 mg/ml in aqueous dilution, reflecting hospital practice in the use of antibiotic discs.

Sensitivity test

An overnight nutrient broth (OXOID) culture of each bacterial isolate, equivalent to 107 cells/ml, was diluted 1:100 in sterile distilled water. A seeded pour-plate was prepared from a mixture of 0.1 ml from the 1:100 dilution and 20 ml molten nutrient agar. Wells were dug with a flame-sterilized size 3 cork-borer of 6.5 mm internal diameter. The undiluted honey and its respective fractional dilutions, as well as the 5 mg/ml and 10 mg/ml of cloxacillin and ampicillin, were filled in turn into the wells. The culture plates were allowed to remain on the bench for a pre-incubation diffusion period of 2 h, followed by incubation in an upright position for 24 h at 37 °C. The bacterial isolates were then assessed for sensitivity or resistance, using the zone of growth inhibition as an index.


The sensitivity test revealed for S. aureus and S. albus a better susceptibility pattern to undiluted honey and its lower dilutions (1:2 and 1:4) than to cloxacillin and ampicillin. This was evident in the zones of growth inhibition produced by undiluted honey, which varied from 6 to 33 mm against the two Staphylococcus spp. isolates. Cloxacillin produced zones of growth inhibition varying from 6 to 24 mm against S. aureus and 9.5 to 18 mm against S. albus. With ampicillin, the range was 8 to 28 mm against S. aureus and 10 to 21 mm against S. albus (Table II).

<% createTable "Table II ","Some results of sensitivity tests on honey and two standard antibiotics against the clinical isolates of S. aureus and S. albus",";Isolate; Clinical source §1,5§Honey - zone of growth inhibition (mm) §1,2§ Antibiotics@;   0*; 1:2; 1:4; 1:6; 1:8; Cloxacillin(5 mg/ml); Ampicillin (10 mg/ml)@;S. aureus; Eye swab; 13; 9; 9; –; –; 19; –@;S. aureus; Eye swab; 16; 10; 10; –; –; 8.5; –@;S. aureus; Eye swab; 10.5 ;10; 10; –; –; –; 10 @;S. aureus; Eye swab; 23; 19; 19; –; –; –; 20 @;S. aureus; Eye swab; 31; 19; 18; 10; 6; 21; 15 @;S. aureus; Wound swab; 15; 7; 5; –; –; 16; – @;S. aureus; Wound swab; –; –; –; –; –; 20; –@;S. aureus; Wound swab; 23; 21; 9; –; –; –; –@;S. aureus; Wound biopsy;15 ;15; 15 ;7 ;– ;12 ;21@;S. aureus; Urine; 32; 13; 13; –; –; 20; – @;S. albus; Eye swab; 23; 15; 15; –; –; –; 10.5@;S. albus; Eye swab; 31 ;16; 16; 10; 10; 13; 15 @;S. albus; Eye swab; 29 ;20; 17; 10; –; –; –@;S. albus; Eye swab; 31; 18; 16; 10; 10; –; –@;S. albus; Eye swab; –; –; –; –; –; 14.5; –@;S. albus; Wound swab; 32; 18; 18; –; –; 11; 16 @;S. albus; Wound swab; 22; 21; 18; 15; 13; 11.5; –@;S. albus; Wound swab; 9; 6; 6; –; –; –; –@;S. albus; Wound biopsy;11; 11; 10; –; –; 9.5; 21 @;S. albus; Wound swab; 33; 30 ;10; –; –; 18; 12","",4,300,true %>

Undiluted honey thus recorded 96.6% antibacterial activity, followed by 93.3% for 1:2 dilution, and 86.6% for 1:4 dilution against S. aureus. Against S. albus, undiluted honey gave respectively 95%, 95%, 60%, and 50% of antibacterial activity. Two strains of the two Staphylococcus spp. (SA6 and SB18) showed total resistance to honey (Table III).

<% createTable "Table III ","Relative percentage resistance of Staphylococcus spp. to honey and the two standard antibiotics",";Staph. spp; Honey §1,7§Antibiotic @;; 0*; 1:2; 1:4; 1:6; 1:8 ;1:10 ;Cloxacillin (5 mg/ml); Ampicillin (10 mg/ml)@;S. aureus ;3.3%; 6.6%; 13.3%; 73.3%; 86.6%; 100%; 36.6% ; 60%@;S. albus; 5.0%; 5.0%; 5.0%; 4.0%; 50%; 80%; 50%; 55%","",4,300,true %>

With respect to the two standard antibiotics, 63.3% of S. aureus isolates and 50% of S. albus isolates were sensitive to cloxacillin, in sharp contrast to the 40% and 45% of S. aureus and S. albus, respectively, against ampicillin. Of the 23 isolates of S. aureus that showed resistance to either of the two antibiotics, only six (i.e. 26%) were resistant to both antibiotics. With S. albus, eight strains (66.6%) of the 12 similarly resistant isolates were resistant to both antibiotics. Table III shows the relative percentage resistance of the two Staphylococcus spp. to honey and the two standard antibiotics.

Discussion and conclusion

The antistaphylococcal activity recorded in this study for honey was consistent with earlier reports on its curative and antibacterial properties.11,14,16,19,20 S. albus showed a resistance level of 5.0% to undiluted honey and of 3.3% to S. aureus. Zones of growth inhibition of 10 mm or more for 50% and 75% dilutions of honey in agar-cup diffusion as the basis for offering honey as a unique treatment in staphylococcal infections16 found support in this study.

Compared with honey and its 1:2 to 1:6 dilutions, cloxacillin and ampicillin used in concentrations of 5 mg/ml and 10 mg/ml were less active on both S. aureus and S. albus. The resistance levels among the S. aureus isolates were respectively 36.6% and 60% for cloxacillin and ampicillin compared with 3.3%, 6.6%, and 13.3% for undiluted honey, 1:2, and 1:4 dilutions respectively. With S. albus, the picture was similar. These results find an analogy in an earlier report14 of more effective treatment of bacterial infection with honey in burn wounds than with silver sulphadiazine, a recognized antibacterial ointment.

This study buttresses the potential of honey as a suitable therapeutic agent against bacterial infections, particularly those due to resistant staphylococci, in the search for a solution to the problem of antibiotic resistant strains.

RESUME. Cinquante isolats bactériens composés de Staphylococcus aureus et de Staphylococcus albus provenant de six sources pathologiques ont été exposés au miel et à ses dilutions fractionnelles comme aussi à deux antibiotiques standard - cloxacilline (5 mg/ml) et ampicilline (10 mg/ml) - dans un test de sensibilité de diffusion «agar-cup». Le miel non dilué présentait une activité antibactérienne à presque 100% des isolats bactériens tandis que ses dilutions fractionnelles de 1:2 jusqu’à 1:10 possédaient des propriétés antibactériennes contre les isolats en ordre décroissant d’activité. La cloxacilline présentait une activité antibactérienne plus élevée par rapport à l’ampicilline mais toutes les deux exerçaient une activité antibactérienne inférieure à celle du miel. L’application thérapeutique du miel, à la suite des épreuves cliniques, devrait être importante dans le traitement des infections bactériennes et en particulier les infections causées par les deux espèces de Staphylococcus testées.


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<% riquadro "This paper was received on 31 May 2005.
Address correspondence to: Dr O.E. Adeleke, Unibadan P.O. Box 22039, Orita U.I. Ibadan, Nigeria. E-mail: adelzek@yahoo.com" %>