Abstract
The quality of 20 selected medicinal plants commonly used in traditional medicine in Jordan was evaluated. Crude and powdered plant materials of the selected species were subjected to macroscopic examination to confirm their identity and to detect adulterations. Macroscopic evaluation revealed that Melissa officinalis. L. leaves were substituted by Aloysia citriodora. Ort. leaves, Achillea fragrantissima. (Forskal) Sch. Bip herb was substituted by Artemisia. spp., and Anchusa officinalis. L. stems were substituted by Paronychia argentea. Lam. herb. Microscopic examination of some powdered plant samples (Curcuma longa. L. roots and Paronychia argentea. herb) indicated adulteration with dyes. This was confirmed by comparative thin-layer chromatography (TLC) analysis of extracts prepared from the purchased powdered and crude samples. Microscopic key elements for 11 of the selected medicinal plants, not previously reported, have been identified and proposed. These are Achillea fragrantissima. herb, Alhagi maurorum. Medic. roots, Anchusa officinalis. stems, Cyperus rotundus. L. roots, Aloysia citriodora. leaves, Moringa aptera. Garten. fruits, Paronychia argentea. herb, Prunus mahaleb. L. seeds, Rhus coriera.. fruits, Vitex agnus castus. L. stems and fruits, and Ziziphus spina christi. L. Desf. leaves.
Introduction
For many centuries, plants have been used for medicinal purposes and are still employed worldwide as home remedies and in modern health care services (Oliver-Bever, Citation1986; WHO, Citation1999). A World Health Organization (WHO) survey revealed that a large percentage of the world population relies on medicinal plants to alleviate their illnesses (WHO, Citation1999). In Jordan, medicinal plants purchased from herbalist's shops are commonly used in crude forms. Moreover, complementary medicine and plant-based products are gaining popularity (Fawzi & Qurran, Citation1988; Al-Khalil, Citation1995; Al-Eisawi, Citation1998; Oran & Al-Eisawi, Citation1998; Abu-Irmaileh & Afifi, Citation2000). The significant increase in the consumption of medicinal plants and their products necessitates close monitoring of their quality using simple and inexpensive techniques. In an attempt to establish a national record of the quality of marketed medicinal plants in different geographical regions throughout the kingdom, five samples of each of the selected medicinal plants commonly used were evaluated. Plant species evaluated during this study are listed in .
Table 1.. List of selected plants used in traditional medicine in Jordan.
The first step involved the establishment of the botanical identity of whole crude plant material. The second step involved the identification of their main microscopic characteristics in crude and powdered samples, and findings were compared with available reported data. The third stage was devoted to comparative Thin-layer-chromatography (TLC) for extracts from the whole and the powdered samples to detect the presence of adulterants. Plant extracts were screened for the presence of five major groups of secondary metabolites, including alkaloids, flavonoids, coumarins, saponins, and essential oils.
Materials and Methods
Plant material
Five samples representing the 20 selected plant species were purchased from five geographically different herbal shops, known locally as attareen. A voucher specimen from each plant was deposited, after giving the herbarium specimen number, in the Department of Pharmaceutical Sciences, Faculty of Pharmacy, University of Jordan.
Macroscopic identification
Macroscopic identification of the purchased whole samples including size, shape, nature of the outer and inner surfaces, type of the fracture, and organoleptic characteristics of whole and powdered samples including color, odor, and taste were carried out by comparing their morphological characteristics with those present at the Herbarium at the Faculty of Science, University of Jordan, and using reported published descriptions and photographs (Al-Eisawi, Citation1998; Montasir & Hassib, Citation1956; Rechinger, Citation1964; Migahid & Hammudah, Citation1974; Duke, Citation1985; Rizk, Citation1986; Bremness, Citation1988; WHO, Citation1989; BHP, Citation1993). Emphasis was given to the detection and recording of the possible adulterations.
Microscopic identification
Purchased crude and powdered samples were examined microscopically in chloral hydrate solution prepared according to the British Pharmacopoeia. (Citation1998). Each sample was examined in duplicate to identify all diagnostic elements and compared to published data, when available. Identified foreign microscopic elements were recorded.
Thin-layer chromatography analysis
Extracts from each sample were prepared according to Wagner and Bladt (Citation1996), applied to precoated TLC silica gel plates, and developed in the appropriate solvent systems. Chromatograms were examined before and after spraying under UV and daylight to detect the presence of alkaloids, coumarins, essential oils, flavonoids, and saponins. The major groups of active constituents are recorded in .
Table 2.. Thin-layer chromatography screening of the purchased samples for the detection of the reported active constituents.
Chromatographic material
Precoated TLC plates were used for the chromatographic analysis of the crude extract of the purchased samples (silica gel 60 F254, Fluka, Buchs, Switzerland). The following solvent systems were used: CHCl3:MeOH:ammonia (80:40:1) for alkaloids, EtOAc:HCO2H:glacial HOAc:H2O (100:11:11:26) for flavonoids, CHCl3:glacial HOAc:MeOH:H2O (64:32:12:8) for saponins, toluene:EtOAc (93:7) for essential oils, and diethyl ether:toluene (1:1 saturated with 10% HOAc) for coumarins.
Spraying reagents
Dragendroff's (alkaloids), iron (III) chloride reagent and natural product/polyethylene glycol (flavonoids), anisaldehyde/sulfuric acid (saponins), vanillin/sulfuric acid (essential oils), and potassium hydroxide (coumarins) reagents were prepared according to Wagner and Bladt (Citation1996) and used during this study.
Results and Discussion
Morphologic descriptions and microscopic characteristics of medicinal plants are still considered the preliminary step for the identification of medicinal plants and for the detection of adulterants as part of the quality control of plant-based medicines (WHO, 1998). In the current study, TLC analysis of the crude extracts was used to support the presence of adulterants detected during macroscopic and microscopic examination of the medicinal plants commonly used in Jordan.
The majority of the purchased samples were free from adulterants. Foreign botanical organs, soil particles, and small stones have been detected in some purchased samples but did not count for more than 2% w/w of the examined samples.
Adulterations observed during this study were attributed to the following reasons:
The use of the same common Arabic name for more than one species from the same genus resulted in species substitution. This was observed during the purchase of Origanum syriacum. L., known locally as zaater., Achillea fragrantissima. (Forskal) Sch. Bip., known locally as qaisoom, be'ethran., and both Ruta. spp., known locally as fijin. or southab., where all were substituted with any available species sharing the common name and/or organoleptic characteristics (Abu-Irmaileh & Afifi, Citation2000).
The use of the same or similar common Arabic names for different plant species or organs representing different genera. For example, it was observed that some shops replace Achillea fragrantissima. by Artemisia. spp. On the other hand, Anchusa officinalis. L. roots and stems, which are not well recognized by many herbalists, were occasionally substituted with Paronychia argentea. Lam., because they have similar Arabic names. A. officinalis. is known locally as saq-Al-hamamah. (“pigeon's leg”) and P. argentea. is commonly known as rejl-al-hamamah. (“pigeon's foot”). Fijin., fijim., sadab., and southab. are the commonly used Arabic names for both Ruta graveolens. L. and Ruta chalepensis. L. However, in all herbalist shops visited, only one species of Ruta. was available, which was identified as R. chalepensis.. The botanical identity of the sold Melissa officinalis. (known in Arabic as melissa.) was found to be Aloysia citriodora. Ort. (also known locally as melissa.).
Cost and profit consideration contributed to adulteration of expensive crude drugs. Foreign or other nonofficial plant parts were occasionally mixed with plant material to increase the weight and therefore the income. This was observed in the purchased samples of the leaves and fruits of Ziziphus spina christi. L., which were mixed with stems. In the case of Crocus sativus. L. style, the anthers and petals of the same plant as well as capsicum, ginger, or safflower were identified as adulterants and detected during macroscopic and microscopic examinations. Salt was detected during organoleptic examination of Rhus coriara. L. fruits to enhance the characteristic taste and to increase the weight. In cases where the plant color is an indication for the quality, coloring agents were occasionally added to intensify the color. This was recognized microscopically and was detected chromatographically in some samples of R coriara. L. fruits and Curcuma longa. L. roots.
No published data were located for the microscopic identification of 11 of the selected plant species for the current study. Therefore, the major microscopic key elements detected in all five examined samples of Achillea fragrantissima. herb, Alhagi maurorum. Medic. roots, Anchusa officinalis. stems, Cyperus rotundus. L. roots, Aloysia citriodora. leaves, Moringa aptera. Garten. fruits, Paronychia argentea. herb, Prunus mahaleb. L. seeds, Rhus coriara. fruits, Vitex agnus castus. L. stems and fruits, and Ziziphus spina christi. leaves were characterized and proposed as illustrated in .
Figure 1. Key elements for the microscopic identification of 11 medicinal plants.
TLC chromatograms of extracts from all five samples representing each plant species were examined for uniformity and reproducibility of the detected groups of secondary metabolites and the adulterants. Results of the TLC analysis are listed in . TLC chromatograms indicated the presence of alkaloids in eight of the selected plants with the highest contents detected in Z. spina christi. leaves and fruits and A. maurorum. roots. Variable quantities of flavonoids were detected in all examined plant extracts except in Anchusa officinalis., Cyperus rotundus., and Moringa aptera.. Fourteen plant extracts (Achillea fragrantissima, Alhagi marurorum, Anchusa officinalis, Cyperus rotundus, Aloysia citriodora, Moringa aptera, Oregano syriacum. L., Paroumaychia argentea, Prunus mahaleb, Ruta chalepensis, Urtica dioica. L., Viscum album. L., Zea mays. L. and Ziziphus spina christi.) tested positive for saponins, which was supported by results from frothing and hemolytic activity tests according to Brian's procedure (Brian & Turner, Citation1975). Essential oils were detected as major components in Achillea fragrantissima, Carthamus tinctoria. L., Crocus sativus, Curcuma longa, Cyperus rotundus, Aloysia citriodora, Moringa aptera, Origanum syriacum, Vitex agnus castus. and Zea mays.. Coumarins were detected in Ruta chalepensis., Prunus mahaleb., and Urtica dioica.. Adulterated samples identified during macroscopic and microscopic evaluation failed to reproduce consistent chromatograms compared to the pure samples.
Conclusions
Our study revealed that microscopic examination supported by TLC analysis of medicinal plants can be considered as a quick, in expensive, and valuable quality-control procedure. The majority of the purchased samples of the selected 20 medicinal plant species were pure. Adulterants were mainly detected in powdered plant samples identified during microscopic examination. Further studies are needed for the quality assessment of other medicinal plants available in the Jordanian market, as findings of such studies could be the basis to draft legislation to control this continually growing market.
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