Kangaroo Island Propolis Types and Their Distribution

Colin Duke, Faculty of Pharmacy Rujee Duke, Faculty of Medicine Van Tran, Faculty of Pharmacy

Abdallah Abu-Mellal, Faculty of Pharmacy Douglas King, Faculty of Pharmacy

Tran VH, Duke RK, Abu-Mellal A and Duke CC. (2012) Propolis with high flavonoid content collected by honey bees from Acacia paradoxa.

Phytochemistry 81, 126-132.

Kangaroo thorn, Acacia paradoxa

Abu-Mellal A, Koolaji N, Duke RK, Tran VH and Duke CC. (2012) Prenylated cinnamate and stilbenes from Kangaroo Island propolis and their antioxidant activity.

Phytochemistry 77, 251-259.

Lepidosperma sp. ( Cyperaceae)

OCH3

O

OOH

OCH3

O

OO

OH

methyl (E)-4-(4'-hydroxy-3'-methylbut-(E)-2'-enyloxy)cinnamate

methyl (E)-4-(3'-carboxybut-(E)-2'-enyloxy)cinnamate

Sedge Type 1 Propolis

OH

OHOH

OH

OHO

OH

OH

OH

OHOH

OH

OH

O

OH

OH

OH

O

OH

O

OH

OHOH

O

OH

OOH

O

cinnamates

O-prenylstilbenes

C-prenylstilbenes

Hive S25, 4 December 20091/13/2010 2:14:24 PM

P-082_Proton_02

13.5 13.0 12.5 12.0 11.5 11.0 10.5 10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0Chemical Shift (ppm)

12.274

86

7.6724

6 7.4581

07.2

6038

7.0166

56.8

5613

6.6314

9

6.3265

9

5.6693

15.5

0046 4.5

0599

3.9519

8

3.4072

6 2.0780

3

1.7579

6

0.8799

6

0.0000

0

Hive S22, 4 December 20091/20/2010 11:07:34 AM

P-080_Proton_01

13.5 13.0 12.5 12.0 11.5 11.0 10.5 10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0Chemical Shift (ppm)

13.544

88 13.199

84

11.501

1011.

36652

7.8868

2 7.6107

97.4

3216

7.2608

7

6.5590

6 6.2037

46.0

1336

5.7921

5

5.3487

5

3.9294

6

2.0241

9 0.8804

50.7

5614

0.0000

0

Kangaroo thorn propolis Proton NMR spectrum

Sedge type 1 propolis Proton NMR spectrum

2/26/2010 4:19:15 PM

dcl3_Proton_01

8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0Chemical Shift (ppm)

7.6249

9 7.4566

37.3

0002

7.0499

3

6.8463

4

6.5908

7 6.3510

6

5.7716

0 5.4999

7

5.2861

05.1

6718

4.6219

84.4

8886

3.9446

33.7

7775

3.3896

4 2.0447

5

1.7990

71.7

5110

1.6850

3

1.2538

6

0.8799

6

0.0000

0

100222-31 Lepidosperma sp.

Type 1

7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0Chemical Shift (ppm)

7.6279

2 7.4556

5

7.27

6.8996

8 6.8497

6

6.5752

1

6.3471

4

5.7706

2 5.4994

9

4.6332

34.5

0696

3.9446

33.8

2424

3.3955

2

2.3002

2

1.7985

8 1.7496

4

0.8799

6

0.00

P-129 Sedge type 1 propolis w w

s

s

Lepidosperma sp. Type 2

2/26/2010 4:10:11 PM

100222_03_cdcl3_Proton_01

8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0Chemical Shift (ppm)

8.20

787

8.16

872 7.71

210

7.67

197

7.40

084

6.47

977

5.35

560

5.18

088

3.74

691

3.42

733

3.34

853

1.81

326

1.69

238

1.19

269

1.15

207

0.00

000

100222-03

8.0 7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0Chemical Shift (ppm)

8.1912

38.1

5208 7.7

1308

7.6729

57.5

0557

7.3959

4

6.9373

76.8

9724

6.6833

6

6.4787

96.4

3866

6.0138

5 5.3472

8

4.1301

2 4.0537

7

3.4121

6

3.1982

9 2.2845

62.2

1311

2.0452

41.8

1375

1.7007

01.6

1652

0.8

0.0000

0

P-042

s

w

w w

s

Sedge type 2 propolis

5/24/2010 2:27:21 PM

100501-01_Proton_01

7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0Chemical Shift (ppm)

7.62890

7.45369

7.27359

6.84928

6.59380

6.34763

5.77453

5.48236

5.28806

5.16669

5.10355 4.634

21 4.48935

4.08558

3.94317

3.82375

3.41167

1.82011

1.74866 1.254

35

0.88045

0.00000

5/28/2010 6:17:08 PM

P-250_Proton_01

7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0Chemical Shift (ppm)

7.6670

8 7.4576

1

7.2789

8

6.8987

1

6.5918

5

6.3598

7

5.7750

2 5.5024

2

4.6214

9 4.4962

0

4.0831

43.9

4757

3.8242

4

3.4106

9

2.5860

4

1.7990

7 1.7501

3

1.4266

31.2

5435

0.8799

60.7

1356

0.0000

0

5/28/2010 6:51:16 PM

P-260_Proton_01

7.5 7.0 6.5 6.0 5.5 5.0 4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0Chemical Shift (ppm)

7.66

854

7.45

565

7.26

919

6.86

151

6.62

757

6.36

378

5.77

062

5.50

536

4.63

519

4.49

571

4.08

852

3.94

512

3.82

375

3.41

118 2.21

360

1.79

956

1.74

964

1.25

387

0.87

996

0.00

000PLANT RESIN

BEE HIND LEGS

PROPOLIS FROM HIVE

F S2 S1 K C S1 P

Propolis from 64 Apiary Sites Analysed by 1H-NMR

Propolis Type # Sites Purity # Samples %TOTAL

Sedge Type 1 33 80-100% 157 48.3

70 to 90% 25 7.7

Sedge Type 2 2 80-100% 1 0.3

70 to 90% 1 0.3

Kangaroo Thorn 8 80-100% 16 4.9

70 to 90% 3 0.9

"Flavanone" 8 80-100% 9 2.8

70 to 90% 4 1.2

"C-Prenyl" 3 80-100% 6 1.8

70 to 90% 1 0.3

"Purple spot" 2 80-100% 2 0.6

70 to 90% 1 0.3

Mixed propolis 43 99 30.5

TOTAL 325 100.0

Distribution of 6 Main Types of Propolis

K

K

K K

K K

F F

N

20 km Sedge type 1 propolis

Kangaroo thorn propolis

“Flavanone” propolis

S2 Sedge type 2 propolis

S1

K

F

“C-prenyl” propolis C

“Purple spot” propolis

S1 S1 S1

S1 S1 S1

S1 S1 S1 S1

S1

S1 S1 S1

S1

S1 S1 S1

S1 S2

P

P P C

C K

K

S1

S1

S1

S1 S1 S1

S1

S2

Further Research on Propolis

Statistical Evaluation of NMR profiles • Clustering or scoring of match from comparison of

NMR spectra of propolis using computer software • Statistical evaluation for propolis types based on

cluster/match with selected NMR spectral profiles

Quantitative HPLC Analyses • Major and minor constituent profiles • Accurate and precise determination of amounts of

key constituents • Statistical evaluation for defining single plant source

propolis types based on specific constituents and their quantities

Botanical characterisation of the Lepidosperma sp. Sedge Type 1 and Sedge Type 2 through collaboration • George Plunkett and Jeremy Bruhl, University of New

England, NSW, AUSTRALIA • Karen Wilson, Royal Botanic Gardens, Sydney, NSW,

AUSTRALIA

Medicinal evaluation of extracts and constituents • Indigenous medicinal use of Lepidosperma species • In vitro and in vivo biological testing • Antioxidant assays

Betty McAdam for collection of propolis samples and ongoing research collaboration (JH & E McAdam, Hog Bay Apiary, Penneshaw, Kangaroo Island) Provision of propolis samples for research from beekeepers, on Kangaroo Island, in particular Graham Carmichael Major funding support from a National Health and Medical Research Council Complementary and Alternative Medicine Grant, 2008-2011

ACKNOWLEDGEMENTS

.