Baeckea frutescens L.

 

MALAYSIAN HERBAL MONOGRAPH

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Cucur atap Leaves

 

Baeckea frutescens L.

Myrtaceae 

 fig1a  fig1c
(a)  (b)
fig1b
(c)
Figure 1Baeckea frutescens. (a) Whole plant; (b) flowers; (c) flowering twig. (Photos courtesy of FRIM, 2013)

   

DEFINITION

Cucur atap leaves consist of the powder of dried leaves of Baeckea frutescens L. (Myrtaceae).

SYNONYM

Baeckea chinensis Gaertner, Baeckea cumingiana Schauer, Baeckea cochinchinensis Blume, Baeckea ericoides Schltdl, Baeckea stenophylla F.Muell, Baeckea sumatrana Blume [1, 2].

VERNACULAR NAMES

False rue, shrubby baeckea (English); Cucur atap, chuchur atap, cucuran atap, hujong atap, rempah gunong, timor tasek, ujan atap (Malay); Gang son (Chinese) [1, 2].

CHARACTER

Colour            :           Greenish brown (powder)

Odour            :           Characteristic

Taste             :           Bitter

IDENTIFICATION

Plant Morphology

B. frutescens is an evergreen, heather-like shrub or small tree up to 8 m tall. Bark greyish brown, fissured and flaky; branches upright, then spreading and drooping, with wiry ends. Leaves opposite, seemingly in clusters at condensed nodes, needle-like, 6–15 mm long x 0.4–0.8 mm wide, base narrowly cuneate, apex obtuse or acute, margin entire, resinous aromatic when crushed; petiole 0.5 mm long. Inflorescence axillary, 1-flowered; peduncle absent or very short; pedicel 0.8–1.7 mm long; bracteoles 2, early caducous. Flowers bisexual, 5-merous, hypanthium obconical to campanulate, 1.5–2.2 mm long, partly fused to the ovary; sepals semiorbicular, 0.4–0.9 mm long x 0.6–1.1 mm wide; corolla up to 5 mm across, petals orbicular, 1.1–1.8 mm across, white, oil glands present; stamens 7–13 in groups of 1–3 opposite each hypanthium lobe, filament 0.5–0.8 mm long, anther about 0.3 mm long; ovary 2–3 locular, 12–18 ovules per locule, style terete, about 1.2 mm long. Fruit a hemispherical to campanulate capsule opening by 3–4 longitudinal slits. Seedling with epigeal germination; cotyledons small, green; hypocotyl elongated. [1]

Microscopy

Powdered material consists of abundant adaxial and abaxial epidermis cells with straight to wavy anticlinal wall, tetracytic stomata  present only in abaxial epidermis. Brown coloured secretory cells found in group, sometimes associated with hypodermal tissue, brachysclereid cells, mesophyll palisade and parenchyma cells. The fragment of fibres found in groups or usually associated with vessels. Abundant of vessels are mostly of longitudinal pitted thickening and thick reticulated vessels. The very few starch granules are mostly simple, round to oval-shaped, usually found in group. Calcium oxalates consist of druses and prismatic crystals. Abundant brachysclereid cells are found in groups, very rarely isolated.

 

fig2a fig2b fig2c
(a) (b) (c)
fig2d fig2e fig2f
(d)  (e)  (f)
fig2g fig2h fig2i
(g) (h) (i)
fig2j fig2k fig2l
(j) (k) (l)
Figure 2 : Microscopic characters of Baeckea frutescens leaves powder of 0.355 mm size. (a) Epidermis cells (magnification 20x); (b) mesophyll palisade (magnification 10x); (c) starch granules (magnification 20x); (d) secretory cells (brown coloured) (magnification 20x); (e) adaxial epidermis cells attached with tetracytic stomata (arrow) (magnification 20x); (f) brachysclereid cells (arrow) (magnification 20x); (g) thick reticulated vessel (magnification 20x); (h) fragment of pitted vessel (magnification 20x); (i) mesophyll palisade (magnification 20x); (j) fragment of fibres (magnification 20x); (k) druse crystal [under polarizing filter] (magnification 10x); (l) prismatic crystal (magnification 20x). [Scale bars: a–l = 20 µm]

 

Colour Tests 

Observed colour of solution after treatment with various reagents:

 

HCl (conc.) :        Yellow
NaOH (5%) :        Brown

Thin Layer Chromatography (TLC)

Test solution :

Weigh about 0.5 g of B. frutescens dried leaves powder in a screw cap conical flask and add 5 mL of ethanol. Sonicate for 15 minutes and filter the solution through filter paper. Evaporate the filtrate to dryness. Reconstitute with 5 mL methanol and use as a test solution. 

Standard solution :

Dissolve myricitrin standard [CAS no.: 17912-87-7] in methanol to produce 1.0 mg/mL solution.

Stationary phase :

HPTLC Glass Silica Gel 60 F254, 10 x 10 cm

Mobile phase :

Chloroform : methanol : water;  (6 : 4 : 1) (v/v/v), (with saturation pad) 

Application :

(a) Myricitrin standard solution (S); 8 µL, 8 mm as a band

(b) Ethanol extract of B. frutescens dried leaves powder (L); 8 µL, 8 mm as a band

Development distance : 8 cm
Drying :

Air drying

Detection :

(a) Visible light before derivatization with 10% v/v sulphuric acid;

(b) UV 254 nm before derivatization with 10% v/v sulphuric acid;

(c) UV 366 nm before derivatization with 10% v/v sulphuric acid;

(d) UV 366 nm after derivatization with 10% v/v sulphuric acid in water, heat at 105°C for 10 min.

 

fig3
Figure 3 : TLC profiles of myricitrin standard (S) and ethanol extract of Baeckea frutescens dried  leaves powder (L) observed under (a) visible light before derivatization, (b) UV at 254 nm before derivatization, (c) UV at 366 nm before derivatization and (d) UV at 366 nm after derivatization.

High Performance Liquid Chromatography (HPLC)

Test solution :

Weigh about 0.5 g of B. frutescens dried leaves powder in a screw cap conical flask and add 5 mL of ethanol. Sonicate for 15 minutes. Filter through a 0.45 µm syringe filter and inject the filtrate into the HPLC column.

Standard solution : Dissolve myricitrin standard [CAS no.: 17912-87-7] in methanol to produce 1.0 mg/mL solution.
Chromatographic system  :

Detector: UV 350 nm

Column: C18 column (5 µm, 4.6 mm I.D x 250 mm) (Zorbax Eclipse Plus C18 if necessary) 

Column oven temperature: 25°C

Flow rate: 1.0 mL/min

Injection volume: 2 µL

Mobile phase (gradient mode) :
 Run Time (min)

A-
0.1% formic acid in water

(%)

B-
Acetonitrile

(%)

 0.0  90  10
10.0  70  30 
25.0  60  40 
30.0 0 100
45.0 0 100
System suitability requirements :

Perform at least five replicate injections of  myricitrin (1.0 mg/mL) The requirements of the system suitability parameters are as follow:

  1. Symmetry factor (As) is not more than 1.5.
  2. Percentage of relative standard deviation (RSD) of the retention time (tr) for myricitrin standard is not more than 2.0%.
Acceptance criteria :
  1. Retention time (tr) of myricitrin standard in the test solution is similar to the tof the standard solution.
  2. The ultraviolet (UV) spectrum of myricitrin standard in the test solution is similar to the UV spectrum of the standard solution (optional supportive data).

  

fig4a 
(a)
 fig4b
(b)
Figure 4 : Whole HPLC chromatogram of (a) myricitrin standard solution (1.0 mg/mL) at tr  = 12.889 min and (b) ethanol extract of Baeckea fructescens dried leaves powder showing peak corresponding to myricitrin standard solution at tr  = 12.937 min. 

  

 fig5a
(a)
 fig5b
(b)
Figure 5 : HPLC chromatogram highlighting the elution region of myricitrin in (a) myricitrin standard solution (1.0 mg/mL) at tr  = 12.889 min and (b) ethanol extract of Baeckea fructescens dried leaves powder showing peak corresponding to myricitrin standard solution at tr  = 12.937 min. 

 

 fig6
 Figure 6 : UV spectrum of myricitrin standard solution (1.0 mg/mL) and ethanol extracts of Baeckea frutescens dried leaves powder.

PURITY TESTS

The purity tests are based on B. frustescens dried leaves powder of 0.355 mm particle size.

Foreign Matter

Not more than 2%

 

Ash Contents
Total ash : Not more than 3%
Acid-insoluble ash : Not more than 1%

 

Loss on Drying

Not more than 10%

 

Extractive Values
Water-soluble extracts
Hot Method                 : Not less than 13%         
Cold Method : Not less than 9%
Ethanol-soluble extracts
Hot Method : Not less than 19%
Cold Method : Not less than 16%

SAFETY TESTS

Heavy Metals
Arsenic                      : Not more than 5.0 mg/kg
Mercury : Not more than 0.5 mg/kg
Lead : Not more than 10.0 mg/kg
Cadmium : Not more than 0.3 mg/kg

 

   Microbial Limits
 Total aerobic microbial count  :  Not more than 10cfu/g
 Total yeast and mould count  :  Not more than 10cfu/g
 Bile-tolerant gram negative bacteria  :  Not more than 10cfu/g

 

Specific Pathogens
 Salmonella spp. : Absent in 25 g 
 Escherichia coli  : Absent in 1 g
 Staphylococcus aureus  : Absent in 1 g
Pseudomonas aeruginosa  : Absent in 1 g

 

CHEMICAL CONSTITUENTS

Methanol extract of B. frutescens leaves was found to contain chromones (e.g. 6β-C-glucopyranosyl-5,7-dihydroxy-2-methylchromone, 8β-C-glucopyranosyl-5,7-dihydroxy-2-methylchromone, 6β-C-(2′-galloylglucopyranosyl)-5,7-dihydroxy-2-methylchromone, 8β-C-(2′-galloylglucopyranosyl)-5,7-dihydroxy-2-methylchromone, 6β-C-glucopyranosyl-5,7-dihydroxy-2-isopropylchromone, 8β-C-glucopyranosyl-5,7-dihydroxy-2-isopropylchromone, 6β-C-(2′-galloylglucopyranosyl)-5,7-dihydroxy-2-isopropylchromone, 8β-C-(2′-galloylglucopyranosyl)-5,7-dihydroxy-2-isopropylchromone) and flavonoids (e.g. myricetin 3-O-α-L-rhamnopyranoside  or myricitrin, 3-O-α-L-rhamnopyranosylmyricetinyl-(I-2″,II-2″)-3-O-α-L-rhamnopyranosylmyricetin) [3, 4].

Ethanol extractof B. frutescens dried leaveswas found to contain phloroglucinol derivatives (baeckeol) and flavanones [5, 6].

Dichloromethane extract of B. frutescens leaves was found to contain chromones (e.g. 5-hydroxy-7-methoxy-2-isopropylchromone, 5-hydroxyl-7-methoxy-2-isopropyl-8-methylchromone and 5 hydroxy-7-methoxy-2-iso-propyl-6-methylchromone) and chromanones (e.g. 2,5-dihydroxy-7-methoxy-2-isopropylchromanone, 2,5-dihydroxy-7-methoxy-2-isopropyl-8-methylchromanone, 2,5-dihydroxy-7-methoxy-2-isopropyl-6-methylchromanone, 2,5-dihydroxy-7-methoxy-2,8-dimethylchromanone and 2,5-dihydroxy-7-methoxy-2,6-dimethylchromanone) [7].

Essential oils of B. frutescens leaves was found to contain monoterpenes (e.g. 1,8-cineole, p-cymene, limonene, linalool, β-pinene, α-terpineol, terpinen-4-ol, γ-terpinene and α-thujene) and sesquiterpenes (e.g. β-caryophyllene, clovane-2,9-diol, α-eudesmol, β-eudesmol, γ-eudesmol and humulene epoxide) [8, 9, 10].

MEDICINAL USES

Uses described in folk medicine, not supported by experimental or clinical data

Traditionally, the leaves of B. frutescens are made into a herbal tea and used to encourage menstrual flow and also for rubbing on the body during childbirth [1, 11]. It is also traditionally used for fevers and as energy booster [11].

Biological and pharmacological activities supported by experimental data

Cytotoxic activity

Methanol extract from the dried leaves B. frutescens which produced phloroglucinol derivatives (BF-2), showed strong cytotoxic activity on leukemia cells (L1210) with inhibition concentration at 50% (IC50 = 5.0 µm/mL) in vitro [9].

Ethanol extract from the dried leaves B. frutescens which produced flavanones (BF-4 and BF-5), showed strong cytotoxic activity on leukemia cells (L1210) with inhibition concentration at 50% (IC50 = 0.2 and 0.5 µm/mL) in vitro [10].

Antibacterial activity

Ethanol extract of B. frutescens leaves (10 – 100 mg/mL) showed antibacterial effect to methicillin-resistant Staphylococcus aureus (MRSA), Staphylococcus aureus, Bacillus sp. and Escherichia coli with inhibition zones ranging from 7.0 to 14.5 mm compared to vancomycin using paper disc diffusion method [12].

Anticariogenic activity

Methanol extract (75%) of B. frutescens (1 – 20 mg/mL) leaves showed anticariogenic effect to Streptococcus mutans with microbial inhibition zones ranging from 9.0 to 14.0 mm using the well diffusion method [13].

Clinical studies

Information and data have not been established. 

SAFETY INFORMATION

Preclinical studies (Toxicology studies)

Information and data have not been established. 

Others (Adverse reaction, contraindication, side effect, warning, precaution)

Information and data have not been established. 

DOSAGE

Information and data have not been established. 

STORAGE

Store below 30°C. Protect from light and moisture.

REFERENCES

  1. Yusuf UK. Baeckea frutescens L.In: van Valkenburg JLCH, Bunyapraphatsara N. (Editors). Plant Resources of South-East Asia No. 12(2): Medicinal and poisonous plants 2. Backhuys Publisher, Leiden, The Netherlands. 2001. 96-98.
  2. Umberto Quattrocchi FLS. CRC World Dictionary of Medicinal and Poisonous Plants; common names scientific names, eponyms, synonyms and etymology. CRC Press. New York. 2012. pp. 517.
  3. Kamiya K, Satake T. Chemical constituents of Baeckea frutescens leaves inhibit copper-induced low-density lipoprotein oxidation. Fitoterapia. 2010;81:185-189.
  4. Tsui WY, Brown GD. Chromones and Chromanones from Baeckea frutescens. Phytochemistry. 1996;43(4):871-876.
  5. Fujimoto Y, Usui S, Makino M, Sumatra M. Phloroglucinolsfrom Baeckea frutescens. Phytochemistry 1996;41:923-925.
  6. Makino M, Fujimoto Y. Flavanonesfrom Baeckea frutescens. Phytochemistry. 1999;50:273-277.
  7. Satake T, Kamiya K, Saiki Y, Hama T, Fujimoto Y, Endang H. Chromone C-glycosides from Baeckea frutescens. Phytochemistry. 1999;50:303-306.
  8. Tsui WY, Brown GD. Sesquiterpenes from Baeckea frutescens. Journal of Natural Products. 1996;59:1084-1086.
  9. Ibrahim J, Abu Said A, Siti Aishah AB, Abdul Rashih A, Trockenbrodt M, Chak CV. Constituents of the essential oil of Baeckea frutescens L. from Malaysia. Flavour and Fragrance Journal. 1998;13:245-247.
  10. N’Guyen TT, Duong TT, Ange B, Vincent C, Alain M, Pascal R, Joseph C. Baeckea frutescens leaf oil from Vietnam: composition and chemical variability. Flavour and Fragrance Journal. 2004;19:217-220.
  11. Burkill IH. A Dictionary of the Economic Products of the Malay Peninsula. Vol. (1). Edited by: Ministry of Agriculture (Malaysia). Crown agents for the colonies, London. 1966. pp. 284-285.
  12. Somayeh R, Mahmood AA, Salmah I, Pouya H. Antibacterial activity of leaf extracts of Baeckea frutescens against Methicillin-Resistant Staphylococcus aureus. BioMed Research International. 2014:1-5.
  13. Hwang JK, Shim JS, Chung JY.  Anticariogenic activity of some tropical medicinal plants against Streptococcus mutans. Fitoterapia. 2004;75:596-598.