manuka honey
DESCRIPTION
antimicrobial : honeyTRANSCRIPT
Survey of the antimicrobial activity
of Australian honey
Julie Irish
Dee Carter
Shona BlairSchool of Molecular and Microbial Biosciences
University of Sydney
Antimicrobial activity of honey
Honey has been well studied for its antimicrobial and wound healing properties
Low water activity ~80% sugars
Low pH Gluconic acid pH 3.2 – 4.5
Hydrogen peroxide Glucose oxidase
Antimicrobial activity of honey
Leptospermum honey Leptospermum honey + catalase
Brush box honey Brush box honey + catalase
Staphylococcus aureus
Floral factors Non-peroxide activity (Leptospermum spp.) Also known as Unique Manuka Factor (UMF®) Advantage in clinical situations
Non-peroxide activity is caused by methylglyoxal (MGO) High concentrations of MGO in manuka
honey that correlate strongly with non-peroxide activity
MGO is naturally produced in all cells as a by-product of metabolism
Highly toxic Modifies DNA, RNA, and proteins Targets them for degradation by the cell’s own
machinery Some bacteria and fungi have well-defined
detoxification systems
How does this fit with the current theory about medicinal honey? MGO produced by stressed plants May explain why some organisms are more sensitive
to honey than others Combination of MGO and honey makes it non-toxic to
human cells?
Need further studies on source, mode of action, standardisation in medicinal honeys
Non-peroxide activity is caused by methylglyoxal (MGO)
Survey of Australian honeys for antibacterial activity
Important to find the most highly active honeys for therapeutic use
345 New Zealand honeys tested (Allen et al. (1991))
No published data for Australian honey
Overview of activity of Australian honey
Testing the antibacterial activity of honey Compares the antibacterial activity of a given honey to
that of phenol Staphylococcus aureus is mixed into agar plate Holes cut into agar Various solutions added to wells
25% honey in water
(tests total activity)
OR 25% honey in catalase solution
(tests non-peroxide activity)
OR Solutions of 2 to 7% phenol
(standard curve)
Solutions diffuse out of the wells during incubation
If they prevent the growth of S. aureus
zones of inhibition Zones are measured,
mean diameter squared Standard curve generated
from phenol solutions
Testing the antibacterial activity of honey
Testing the antibacterial activity of honey
y = 137.86x - 108.1
R2 = 0.9889
0
100
200
300
400
500
600
700
800
900
0 1 2 3 4 5 6 7 8
Phenol concenration (% w/v)
Mea
n d
iam
eter
sq
uar
ed (
mm
)
Testing the antibacterial activity of honey
Honey compared to phenol (x 4.69 for dilution and density)
400 mm2 zone of inhibition = 17.3% phenol equivalent
The higher the number, the more active the honey
Allows comparison between different honeys
y = 137.86x - 108.1
R2 = 0.9889
0
100
200
300
400
500
600
700
800
900
0 1 2 3 4 5 6 7 8
Phenol concenration (% w/v)
Mea
n d
iam
eter
sq
uar
ed (
mm
)
Results
503 honey samples tested 477 Apis mellifera 26 Trigona spp.
Activity (phenol equivalent) varies greatly <5 Insignificant therapeutic value 5-10 Low activity >10 Therapeutically beneficial >20 Highly active
Total activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 93 7 135 40 275 (57.7%) 175 (64%)
VIC 14 1 2 4 21 (4.4%) 6 (29%)
TAS 23 2 20 1 46 (9.6%) 21 (46%)
QLD 12 0 7 9 28 (5.9%) 16 (57%)
SA 10 0 5 0 15 (3.2%) 5 (33%)
WA 39 2 20 31 92 (19.2%) 51 (55%)
Sum 191
(40%)
12
(2.5%)
189
(39.6%)
85
(17.9%)
477
(100%)
274
(57.5%)
Total activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 93 7 135 40 275 (57.7%) 175 (64%)
VIC 14 1 2 4 21 (4.4%) 6 (29%)
TAS 23 2 20 1 46 (9.6%) 21 (46%)
QLD 12 0 7 9 28 (5.9%) 16 (57%)
SA 10 0 5 0 15 (3.2%) 5 (33%)
WA 39 2 20 31 92 (19.2%) 51 (55%)
Sum 191
(40%)
12
(2.5%)
189
(39.6%)
85
(17.9%)
477
(100%)
274
(57.5%)
Total activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 93 7 135 40 275 (57.7%) 175 (64%)
VIC 14 1 2 4 21 (4.4%) 6 (29%)
TAS 23 2 20 1 46 (9.6%) 21 (46%)
QLD 12 0 7 9 28 (5.9%) 16 (57%)
SA 10 0 5 0 15 (3.2%) 5 (33%)
WA 39 2 20 31 92 (19.2%) 51 (55%)
Sum 191
(40%)
12
(2.5%)
189
(39.6%)
85
(17.9%)
477
(100%)
274
(57.5%)
Total activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 93 7 135 40 275 (57.7%) 175 (64%)
VIC 14 1 2 4 21 (4.4%) 6 (29%)
TAS 23 2 20 1 46 (9.6%) 21 (46%)
QLD 12 0 7 9 28 (5.9%) 16 (57%)
SA 10 0 5 0 15 (3.2%) 5 (33%)
WA 39 2 20 31 92 (19.2%) 51 (55%)
Sum 191
(40%)
12
(2.5%)
189
(39.6%)
85
(17.9%)
477
(100%)
274
(57.5%)
Total activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 93 7 135 40 275 (57.7%) 175 (64%)
VIC 14 1 2 4 21 (4.4%) 6 (29%)
TAS 23 2 20 1 46 (9.6%) 21 (46%)
QLD 12 0 7 9 28 (5.9%) 16 (57%)
SA 10 0 5 0 15 (3.2%) 5 (33%)
WA 39 2 20 31 92 (19.2%) 51 (55%)
Sum 191
(40%)
12
(2.5%)
189
(39.6%)
85
(17.9%)
477
(100%)
274
(57.5%)
Total activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 93 7 135 40 275 (57.7%) 175 (64%)
VIC 14 1 2 4 21 (4.4%) 6 (29%)
TAS 23 2 20 1 46 (9.6%) 21 (46%)
QLD 12 0 7 9 28 (5.9%) 16 (57%)
SA 10 0 5 0 15 (3.2%) 5 (33%)
WA 39 2 20 31 92 (19.2%) 51 (55%)
Sum 191
(40%)
12
(2.5%)
189
(39.6%)
85
(17.9%)
477
(100%)
274
(57.5%)
Total activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 93 7 135 40 275 (57.7%) 175 (64%)
VIC 14 1 2 4 21 (4.4%) 6 (29%)
TAS 23 2 20 1 46 (9.6%) 21 (46%)
QLD 12 0 7 9 28 (5.9%) 16 (57%)
SA 10 0 5 0 15 (3.2%) 5 (33%)
WA 39 2 20 31 92 (19.2%) 51 (55%)
Sum 191
(40%)
12
(2.5%)
189
(39.6%)
85
(17.9%)
477
(100%)
274
(57.5%)
Total activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 93 7 135 40 275 (57.7%) 175 (64%)
VIC 14 1 2 4 21 (4.4%) 6 (29%)
TAS 23 2 20 1 46 (9.6%) 21 (46%)
QLD 12 0 7 9 28 (5.9%) 16 (57%)
SA 10 0 5 0 15 (3.2%) 5 (33%)
WA 39 2 20 31 92 (19.2%) 51 (55%)
Sum 191
(40%)
12
(2.5%)
189
(39.6%)
85
(17.9%)
477
(100%)
274
(57.5%)
Total activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 93 7 135 40 275 (57.7%) 175 (64%)
VIC 14 1 2 4 21 (4.4%) 6 (29%)
TAS 23 2 20 1 46 (9.6%) 21 (46%)
QLD 12 0 7 9 28 (5.9%) 16 (57%)
SA 10 0 5 0 15 (3.2%) 5 (33%)
WA 39 2 20 31 92 (19.2%) 51 (55%)
Sum 191
(40%)
12
(2.5%)
189
(39.6%)
85
(17.9%)
477
(100%)
274
(57.5%)
Total activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 93 7 135 40 275 (57.7%) 175 (64%)
VIC 14 1 2 4 21 (4.4%) 6 (29%)
TAS 23 2 20 1 46 (9.6%) 21 (46%)
QLD 12 0 7 9 28 (5.9%) 16 (57%)
SA 10 0 5 0 15 (3.2%) 5 (33%)
WA 39 2 20 31 92 (19.2%) 51 (55%)
Sum 191
(40%)
12
(2.5%)
189
(39.6%)
85
(17.9%)
477
(100%)
274
(57.5%)
Total activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 93 7 135 40 275 (57.7%) 175 (64%)
VIC 14 1 2 4 21 (4.4%) 6 (29%)
TAS 23 2 20 1 46 (9.6%) 21 (46%)
QLD 12 0 7 9 28 (5.9%) 16 (57%)
SA 10 0 5 0 15 (3.2%) 5 (33%)
WA 39 2 20 31 92 (19.2%) 51 (55%)
Sum 191
(40%)
12
(2.5%)
189
(39.6%)
85
(17.9%)
477
(100%)
274
(57.5%)
Total activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 93 7 135 40 275 (57.7%) 175 (64%)
VIC 14 1 2 4 21 (4.4%) 6 (29%)
TAS 23 2 20 1 46 (9.6%) 21 (46%)
QLD 12 0 7 9 28 (5.9%) 16 (57%)
SA 10 0 5 0 15 (3.2%) 5 (33%)
WA 39 2 20 31 92 (19.2%) 51 (55%)
Sum 191
(40%)
12
(2.5%)
189
(39.6%)
85
(17.9%)
477
(100%)
274
(57.5%)
60% of all samples had antibacterial activity
57.5% of all samples had therapeutically beneficial antibacterial activity
Distribution of total activity
020406080
100120140160180200
<5 5-10 10-15 15-20 20-25 25-30 30-35
Total antibacterial activity (% (w/v) phenol equivalent)
Nu
mb
er o
f sa
mp
les
Of the honeys that have antibacterial activity, most are in the therapeutically beneficial range
Average total activity (n=286): 17.8 5 (7.4 – 34.3)
Non-peroxide activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 209 6 50 10 275 (57.7%) 60 (22%)
VIC 21 0 0 0 21 (4.4%) 0 (0%)
TAS 43 0 3 0 46 (9.6%) 3 (6.5%)
QLD 22 1 1 4 28 (5.9%) 5 (18%)
SA 14 1 0 0 15 (3.2%) 0 (0%)
WA 88 4 0 0 92 (19.2%) 0 (0%)
Sum 397
(83%)
12
(2.6%)
54
(11.4%)
14
(3.0%)
477
(100%)
68
(14.3%)
Non-peroxide activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 209 6 50 10 275 (57.7%) 60 (22%)
VIC 21 0 0 0 21 (4.4%) 0 (0%)
TAS 43 0 3 0 46 (9.6%) 3 (6.5%)
QLD 22 1 1 4 28 (5.9%) 5 (18%)
SA 14 1 0 0 15 (3.2%) 0 (0%)
WA 88 4 0 0 92 (19.2%) 0 (0%)
Sum 397
(83%)
12
(2.6%)
54
(11.4%)
14
(3.0%)
477
(100%)
68
(14.3%)
Non-peroxide activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 209 6 50 10 275 (57.7%) 60 (22%)
VIC 21 0 0 0 21 (4.4%) 0 (0%)
TAS 43 0 3 0 46 (9.6%) 3 (6.5%)
QLD 22 1 1 4 28 (5.9%) 5 (18%)
SA 14 1 0 0 15 (3.2%) 0 (0%)
WA 88 4 0 0 92 (19.2%) 0 (0%)
Sum 397
(83%)
12
(2.6%)
54
(11.4%)
14
(3.0%)
477
(100%)
68
(14.3%)
Non-peroxide activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 209 6 50 10 275 (57.7%) 60 (22%)
VIC 21 0 0 0 21 (4.4%) 0 (0%)
TAS 43 0 3 0 46 (9.6%) 3 (6.5%)
QLD 22 1 1 4 28 (5.9%) 5 (18%)
SA 14 1 0 0 15 (3.2%) 0 (0%)
WA 88 4 0 0 92 (19.2%) 0 (0%)
Sum 397
(83%)
12
(2.6%)
54
(11.4%)
14
(3.0%)
477
(100%)
68
(14.3%)
Non-peroxide activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 209 6 50 10 275 (57.7%) 60 (22%)
VIC 21 0 0 0 21 (4.4%) 0 (0%)
TAS 43 0 3 0 46 (9.6%) 3 (6.5%)
QLD 22 1 1 4 28 (5.9%) 5 (18%)
SA 14 1 0 0 15 (3.2%) 0 (0%)
WA 88 4 0 0 92 (19.2%) 0 (0%)
Sum 397
(83%)
12
(2.6%)
54
(11.4%)
14
(3.0%)
477
(100%)
68
(14.3%)
Non-peroxide activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 209 6 50 10 275 (57.7%) 60 (22%)
VIC 21 0 0 0 21 (4.4%) 0 (0%)
TAS 43 0 3 0 46 (9.6%) 3 (6.5%)
QLD 22 1 1 4 28 (5.9%) 5 (18%)
SA 14 1 0 0 15 (3.2%) 0 (0%)
WA 88 4 0 0 92 (19.2%) 0 (0%)
Sum 397
(83%)
12
(2.6%)
54
(11.4%)
14
(3.0%)
477
(100%)
68
(14.3%)
Non-peroxide activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 209 6 50 10 275 (57.7%) 60 (22%)
VIC 21 0 0 0 21 (4.4%) 0 (0%)
TAS 43 0 3 0 46 (9.6%) 3 (6.5%)
QLD 22 1 1 4 28 (5.9%) 5 (18%)
SA 14 1 0 0 15 (3.2%) 0 (0%)
WA 88 4 0 0 92 (19.2%) 0 (0%)
Sum 397
(83%)
12
(2.6%)
54
(11.4%)
14
(3.0%)
477
(100%)
68
(14.3%)
Non-peroxide activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 209 6 50 10 275 (57.7%) 60 (22%)
VIC 21 0 0 0 21 (4.4%) 0 (0%)
TAS 43 0 3 0 46 (9.6%) 3 (6.5%)
QLD 22 1 1 4 28 (5.9%) 5 (18%)
SA 14 1 0 0 15 (3.2%) 0 (0%)
WA 88 4 0 0 92 (19.2%) 0 (0%)
Sum 397
(83%)
12
(2.6%)
54
(11.4%)
14
(3.0%)
477
(100%)
68
(14.3%)
Non-peroxide activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 209 6 50 10 275 (57.7%) 60 (22%)
VIC 21 0 0 0 21 (4.4%) 0 (0%)
TAS 43 0 3 0 46 (9.6%) 3 (6.5%)
QLD 22 1 1 4 28 (5.9%) 5 (18%)
SA 14 1 0 0 15 (3.2%) 0 (0%)
WA 88 4 0 0 92 (19.2%) 0 (0%)
Sum 397
(83%)
12
(2.6%)
54
(11.4%)
14
(3.0%)
477
(100%)
68
(14.3%)
Non-peroxide activity<5 5 – 10 10 – 20 >20 Sum
(% of country)
Sum >10(% of state)
NSW 209 6 50 10 275 (57.7%) 60 (22%)
VIC 21 0 0 0 21 (4.4%) 0 (0%)
TAS 43 0 3 0 46 (9.6%) 3 (6.5%)
QLD 22 1 1 4 28 (5.9%) 5 (18%)
SA 14 1 0 0 15 (3.2%) 0 (0%)
WA 88 4 0 0 92 (19.2%) 0 (0%)
Sum 397
(83%)
12
(2.6%)
54
(11.4%)
14
(3.0%)
477
(100%)
68
(14.3%)
17% of all samples had non-peroxide activity
14.3% of all samples had therapeutically beneficial non-peroxide activity
Distribution of non-peroxide activity
0
50
100
150
200
250
300
350
400
<5 5-10 10-15 15-20 20-25 25-30
Antibacterial activity (% (w/v) phenol equivalent)
Nu
mb
er o
f sa
mp
les
Of the honeys that have non-peroxide activity, most are in the therapeutically beneficial range
High non-peroxide activity is rare
0
5
10
15
20
25
30
5-10 10-15 15-20 20-25 25-30
Antibacterial activity (% (w/v) phenol equivalent)
Nu
mb
er o
f sa
mp
les
Non-peroxide activity
Of the 80 honeys with non-peroxide activity, 62 (77.5%) were from Leptospermum, or contained Leptospermum
Mean non-peroxide activity 17.2 4.1 (9.8 – 25.9)
Some species/areas more reliable than others L. polygalifolium (Jelly bush): 29 samples, 28 had non-
peroxide activity L. scoparium (Manuka): 11 samples, none had non-
peroxide activity, some inactive Most active Leptospermum honeys from NSW-QLD border
Non-peroxide activity
18 non-Leptospermum samples had non-peroxide activity
Mean non-peroxide activity 10.1 1.7 (8.1 – 15.9)
Spotted gum (3/6 samples) South west Tasmanian wildflowers (3/5 samples) Stonefruit orchard (1/2 samples) Clover (1/5 samples) Melaleuca (2/28 samples) Moort (1/1 samples) Forest red gum (1/2 samples) Messmate (1/6 samples) Unspecified mixed flora (5/64 samples)
Floral sources 477 honey samples from 142 different floral
sources, including mixed flora 78% of honeys were from native
Australian flora, another 17% were likely to contain natives
38% of honeys contained eucalypts
24% of honeys contained Leptospermum (likely to be biased)
Eastern Australia
Inactive
Hydrogen peroxide activity
Non-peroxide activity
Eastern Australia
Highest total activity: 34.3 (Apple box/Red stringybark)
Highest non-peroxide activity: 25.9 (Jelly bush)
High non-peroxide activity (>20% phenol equivalent) in Leptospermum samples from QLD and northern NSW
12 non-Leptospermum honeys
had non-peroxide activity
Western Australia
Inactive
Hydrogen peroxide activity
Non-peroxide activity
Western Australia Highest total activity: 31.9 (Stonefruit) Highest non-peroxide activity: 9.7 (Melaleuca) 31 honeys with total activity >20
(many from flora endemic to WA)
Jarrah: 19 samples Activity range 17.7 to 31.4 1 sample was inactive
Karri: 3 samples Activity range 18.5 to 29.6
Marri: 9 samples Activity range 18 to 29.7 1 sample was inactive
Important message
It is impossible to predict the activity of a given honey, must test every batch 32 samples, same beekeeper, same floral source,
same time, different hives 31 samples had total activity between 11 and 19 1 sample was inactive
30% of Leptospermum honeys were inactive,
15% had hydrogen peroxide activity only Activity relates to floral source to an extent, but there
are clearly many other factors involved Environmental conditions, soil type, soil and plant
microbiology, bee health…
Stingless bee honey
26 samples of Trigona spp. honey from QLD
Mixed flora, various times of year
23 samples had total activity >20 (12.4 – 32.1)
24 samples had non-peroxide activity >10 (11.5 – 23.7)
Conclusions Australian honeys show a wide range of
antibacterial activities Many have potential for therapeutic use Non-peroxide activity exists in honeys from
various floral sources Leptospermum spp. remain the most
reliable source of high non-peroxide activity Statistical analysis is ongoing Identified areas to focus on in
future studies Role of MGO?
Acknowledgments
The 124 beekeepers who supplied honey samples for the survey
Rob Manning, Dept of Agriculture, WA Tim Heard, CSIRO Entomology, QLD Comvita, New Zealand RIRDC