Home Archive Vol 36, No.2, 2010 For practitioner Researches Upon the Heavy Metals Content of Sambucus nigra L. (Caprifoliaceae)

Researches Upon the Heavy Metals Content of Sambucus nigra L. (Caprifoliaceae)

 TANIA POPA(1), MARIA-VIORICA BUBULICĂ(1), L. CHIRIGIU(1), G. D. MOGOŞANU(1), R. POPESCU(2), H. POPESCU(1,3)

(1)Faculty of Pharmacy, University of Medicine and Pharmacy of Craiova; (2)Faculty of Dental Medicine, University of Medicine and Pharmacy of Craiov; (3)Faculty of Pharmacy, “Iuliu Haţieganu” University of Medicine and Pharmacy, Cluj-Napoca

ABSTRACT The analysis of heavy metals in the flowers and leaves of Sambucus nigra species has been performed. The mean levels of heavy metals in vegetal dried samples were in normal limits. Differences depend on the presence of organic compounds with ligand character and on the environmental conditions. The results indicate that there is not any real danger by utilization of pharmaceutical preparations of Sambuci flos.

KEY WORDS Sambucus nigra, heavy metals, content.

 


Introduction

Sambucus nigra L., elder, elderberry, European elder (Caprifoliaceae), is a species native to most of Europe, northwest Africa and southwest Asia. It is a deciduous shrub or small tree of 4–6 m tall. It grows in a variety of conditions including both wet and dry fertile soils, primarily in sunny locations, forests, riversides [1–5].

In the specialty papers, there are very few data on the chemical composition and pharmacological action of the S. nigra leaves.

From the phytochemical point of view, S. nigra species contain: cyanide glycosides (S­sambuni-grin), flavonosides (rutin), anthocyanins (in fruits), saponins, catechic tannin, mucilages, volatile oil (in flowers), polyphenolic acids (caffeic and chlo-rogenic acid), sugars, organic acids, vitamins (ascorbic acid), lipids, mineral salts. Sickening smell of fresh flowers is due to some aliphatic amines: ethylamine, n-propylamine, i­propyl-amine, buthylamine, etc. Through enzymatic hydrolysis, sambunigrin released hydrocyanic acid, benzaldehyde and glucose [5–13].

Elder flowers have the following pharmacolo-gical actions: diuretic and diaforetic (flavonosides, saponins), antitussive (sambunigrin), expectorant (saponins), softeners and immunostimulatory (mucilages), slightly laxative, antirheumatic. It is used to treat influenza and other respiratory illnesses (virosis) accompanied by fever. Due to easy action laxatives and diuretics, Sambuci flos is used in diets. Elder flowers are given external as hot baths or poultices, for the treatment of fu-runculosis, abscesses, burns, blisters, rheumatism [4–9, 14–20].

The concentration of heavy metals represents one of the quality parameters in the analysis of vegetal medicinal products, because the producing plants are often harvested from different areas, in different environmental conditions. For example, the availability of heavy metal cations to plants is influenced by the soil properties such as salinity, pH (acidity), humidity, and/or existence of metal-rich minerals [21].

The analysis of heavy metals content in the flowers and leaves of S. nigra (Caprifoliaceae) species harvested from the Romanian flora was the aim of our research

Material and Methods

Sampling

For an accurate analysis, the vegetal samples must be harvested in suitable climate conditions, from areas with unpolluted soil, water and air, in a certain time of day, and from a sufficient number of plants [21].

From the S. nigra species, twenty vegetal samples were collected at the flowering, in June 2009, from the surroundings of Dobreşti village, Dolj County.

Preparation of plant tissue for analysis

Sample preparation is critical in obtaining accurate data and reliable interpretation of plant analysis results.

The vegetal products should be processed during decontamination, drying, particle-size reduction, storage and organic matter destruction.

Plant material must be cleaned and free of extraneous substances, including soil and dust particles that may influence analytical results.

The decontamination process must be thorough while still preserving sample integrity. Deconta-mination procedures involving washing and rinsing with deionized water and 0.2% detergent solution (non-phosphate), should only be used for fresh, fully turgid plant samples. After decontami-nation, water is removed from plant tissue, at temperatures under 600C, to stop the enzymatic reactions and to stabilize the samples.

Plant tissue samples are reduced to 0.5 to 1.0 mm particle size to ensure homogeneity and to facilitate organic matter destruction [21].

Gravimetric determination of ash

The ash represents the residue obtained through the dry ashing of a matter, being made by inorganic compounds. Dry-ashing is conducted in a muffle furnace at temperature 500 to 5500C for four to eight hours. For tissues high in carbo-hydrates and oils, ashing aids may be required to achieve complete decomposition of organic matter [21].

At the end of the ashing period, the vessel is removed from the muffle furnace, cooled, and the ash is dissolved in nitric acid. The final solution is diluted as needed to meet the range requirements of the analytical procedure or instrument utilized [21].

Weigh 0.5 to 1.0 g dried plant material that has been ground and homogenized into a high-form, 30 mL porcelain crucible.

Samples were placed in a cool muffle furnace. Temperature control of the furnace was set to allow gradual increase (two hours) in the ashing temperature and maintain for four to eight hours. After that, the furnace was turned off to allow samples to cool (in one hour). Then, the ash is weighing on analytical balance nearest 0.1 mg.

High temperature oxidation. Heavy metals detection

This method prepares plant tissue for the quantitative determination of the content of Ca2+, Zn2+, Fe2+/3+, Mn2+, Ni2+, Pb2+, Cr3+, by atomic absorption spectrometry (AAS), utilizing high-temperature dry oxidation of the organic matter and dissolution of the ash with 4% nitric acid. The method detection limit is approximately 0.04%. The method is generally reproducible within ±7% [21].

The tissue samples (leaves, stems) were prepared in the above-mentioned manner.

Heavy metals content was determined using analytical balance, porcelain crucibles, muffle furnace, volumetric labware, deionized water, standard calibration solutions, and an AAS–30 Carl Zeiss Jena (Germany) spectrometer with Photron & Narva cathode (Table 1).

Five standard calibration dilutions (0.001 mg/L to 2 mg/L) were prepared starting from 5 mg/L reference solutions diluted with 4% nitric acid.

Table 1 – Experimental data

Parameters

Mo

Pb

Zn

Ni

Ca

Cr

Fe

Wavelength 
[nm]

313.3

283.3

213.9

232.0

422.7

357.9

248.3

Lamp current [mA]

15

9

13

12

9

10

11

Air flow 
[L/hr]

590

620

710

630

680

690

Acetylene flow [L/hr]

150

65

65

69

65

65

69

N2O flow 
[L/hr]

620

Flame type

Red.

Ox.

Ox.

Ox.

Ox.

Ox.

Ox.

Red. – reducing; Ox. – oxidizing.

Results and Discussion

The results of AAS analysis are given as mean and standard deviation (Table 2, Figure 1).

Table 2 – Heavy metals content of S. nigra samples (flowers and leaves)

Heavy metals

Sambuci flos

Sambuci folium

[mg%]

[mg%]

Ca2+

0.6398 ± 0.02

1.1230 ± 0.2

Zn2+

1.5678 ± 0.2

2.1604 ± 0.2

Mn2+

0.0936 ± 0.002

0.1117 ± 0.02

Fe2+/3+

0.3081 ± 0.02

0.9233 ± 0.02

Ni2+

0.2722 ± 0.02

0.0692 ± 0.002

Pb2+

0.8940 ± 0.02

3.2755 ± 0.2

Cr3+

0.0943 ± 0.001

0.4323 ± 0.01

sample [g]

0.8592 ± 0.0002

1.2167 ± 0.0002

dried sample [g]

0.7818 ± 0.0002

1.1089 ± 0.0002

ash [g]

0.0674 ± 0.0002

0.0963 ± 0.0002

% dry ashing residue

8.621± 0.5

8.684 ± 0.5

The maximum limits of heavy metals allowed in plants for adequate growing and development are the followings: 300 mg% Ca2+, 10 mg% Zn2+, 5 mg% Mn2+, 100 mg% Fe2+/3+, 0.80 mg% Ni2+, 50 mg% Pb2+ and Cr3+ [21].

In the vegetal products obtained from S. nigra species, the content of the heavy metals can be considered as normal.

All samples contain Pb2+ and Cr3+, markers for soil and air pollution. The levels of Pb2+ (normal limits) in vegetal tissues samples appear because the plants collected were relative closely to roads (cars circulation).

Generally, for most plants the concentrations of some heavy metals are higher in the roots then in the aboveground parts.

This is an important finding, because only the flowers and fruits of S. nigra species are usually used as medicinal products.

High levels of Ca2+, Zn2+, Mn2+, Fe2+/3+, Pb2+and Cr3+, but in normal limits, have been determined in leaves, because their intake from soil or aerial parts.

Figure 1 – Maximum limits allowed and heavy metals content of S. nigra samples 
(flowers and leaves).

The flowers contain high levels of Ni2+ pro-bably because of the bioinorganic mechanisms during the flowering period and of the soil acidity.

AAS analysis confirmed indirectly that the samples were collected from mature S. nigra species, during the flowering period, when appear complex combinations with divalent (Ca2+, Zn2+, Mn2+, Fe2+, Pb2+) or trivalent (Fe3+, Cr3+) metallic cations.

For the internal use of medicinal products as vegetal powders and extracts, the soil analysis and the collecting of samples from unpolluted zones are strictly recommended. Therefore, the quality control of the raw materials before their further use is the only safe way to prevent the high heavy metals intake during the administration of Sambuci flos pharmaceutical preparations. High contents of heavy metals in the raw product (aerial parts) could be induced by the low pH soils especially from heavy metals mineral-carriers. Thus, medicinal plants harvesting is not recommended in these conditions.

The analysis of plants mineral content is very important to determine the quality of medicinal products taking into account some main factors, which are interactive: soil, geographic area, water, climate, growth stage, genetic susceptibility and physiological stage of plants, genotype–phenotype correlations, time of sampling [21].

Conclusions

The content of some heavy metals (Ca2+, Zn2+, Mn2+, Fe2+/3+, Ni2+, Pb2+, Cr3+) of the flowers and leaves of S. nigra species has been established using atomic absorption spectrometry analysis.

The content of the heavy metals can be considered in normal limits, even for Pb2+ and Cr3+ markers for soil and air pollution.

High levels of Ni2+ have been determined in the flowers then in the leaves.

The samples have been collected during the flowering period, when appearing complex combinations with divalent (Ca2+, Zn2+, Mn2+, Fe2+, Pb2+) or trivalent (Fe3+, Cr3+) metallic cations.

The results indicate that there is not any real danger by utilization of pharmaceutical preparations of Sambuci flos.

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Corresponding address: Assistant Professor George Dan Mogoşanu, Department of Pharmacognosy & Phytotherapy, Faculty of Pharmacy, University of Medicine and Pharmacy of Craiova, 2–4 Petru Rareş Street, 200349 Craiova, Romania; Phone/Fax +40251–523 929, e-mail: mogosanu@umfcv.ro, mogosanu2006@yahoo.com


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