Abstract: Eurycoma longifolia which is known as Tongkat Ali is commonly found in Asian countries such as Malaysia, Indonesia, Thailand, Myanmar and Cambodia. This plant is famously known for its various pharmacological activities. The plant is also reported to consist of various types of important bioactive compounds such as quassinoids, canthine-6-one alkaloids, triterpenes, squalene derivatives, $-carboline alkaloids etc which are mostly found in the root part. The presence of these important phytochemicals contributes to their different types of therapeutic effects more especially in terms of aphrodisiac properties which have resulted in a massive increase in demand and production of their Herbal Medicinal Products (HMP). These situations have resulted in the production of E. longifolia HMPs whose quality are questionable, which might be as a result of restricted of sources that might lead to some unethical activities carried out by suppliers and manufacturers in order to gain more profit. Therefore, this review focused on adulteration issues such as contamination and substitution of E. longifolia HMP. The review also includes the possible solutions on how to improve the quality of these HMP so as they can be safe for consumption. Embracing pharmacovigilance in the preparation of the HMP, proper implementation of agricultural practices such as Good Agricultural and Collection Practices (GACP) and Good Manufacturing Practices (GMP) together with the establishment of effective regulatory bodies would undoubtedly improve the quality of E. longifolia HMP sold in the market. The detailed knowledge about the main composition of the E. longifolia HMP will help to ascertain their quality, efficacy and safety as these are very important toward quality control.
INTRODUCTION
Eurycoma longifolia is a common tropical medicinal plant which belongs to the family Simaroubaceae and is distributed in several parts of Southern Asia including Malaysia. In Malaysia, Eurycoma longifolia is known for its aphrodisiacs properties mostly, to improve libido and energy1. This plant is also known as Tongkat Ali by the Malays ethnic group due to resemblances of walking stick with twisted roots. The aphrodisiac activity of this plant is mostly contributed by root part2. The plant is also used traditionally by local folks for the treatment of other ailments such as bleeding, cough, fever, anti malarial treatment, ulcer, high blood pressure, fevers, etc. and all these unique activities performed by E. longifolia is as a result of the presence of very important bioactive compounds known as quassinoid3,4.
Eurycoma longifolia have been confirmed in several studies to exhibit different types of pharmacological effects such as anti malarial5-7, aphrodisiac8-10, anti cancer11-13, ergogenic14,15 and toxicity effects16. This plant species are also reported to consists of several classes of metabolites and the major among them are the quassinoids, canthin-6-one and beta-carboline alkaloids, squalene derivatives, tirucallane-type triterpenes, biphenylneolignans17. Other varieties of secondary metabolites presence in this plants species include tannins, polysaccharides, glycoprotein and mucopolysaccharides18. This study tends to highlight the important bioactive compounds present in E. longifolia and the risk associated with the HMP consumption and the various possible solutions on how to improve their quality so that they can be safe for consumption.
TAXONOMY, CULTIVATION AND ETHNOBOTANICAL USES
The family of Simaroubaceae consists of about 30 genera and more than 200 species including Eurycoma longifolia19. Most of the plants in this family generally have bitter taste caused by the presence of quassinoid which contributes most of their pharmaceutical properties20. This plant family is generally distributed around tropical Africa, region of Australia, West Africa, Madagascar and Asia (mostly Malaysia)2. Only 8 genera and 10 species are found in Malaysia with Eurycoma longifolia been the abundant19. The scientific classification of Eurycoma longifolia plants is shown in Table 1.
There are different species of plants which are referred to as locally Tongkat Ali and these includes; Entomophthora apiculata, Polyalthia bullata, Goniothalamus sp. and E. longifolia. Among these four species, E. longifolia is the most used17.
| Table 1: | Systemic classification (taxonomy) of Eurycoma longifolia |
In Asia, due to the richness of biodiversity and suitable climate, E. longifolia plant grows naturally. They are also being called by different names depending on the countries such as Tongkat Ali in Malaysia and Singapore, Pasak bumi and Tung saw in Indonesia and Thailand respectively or Cay Ba Binh in Vietnam21.
Among all the common names used to describe this plant species worldwide, Tongkat Ali or Long Jack are the most recognized names both locally and internationally. Presently, there are over 200 commercialize HMP available and most of them emphasizing on the aphrodisiac properties21. Due to modern lifestyle and health conscious, many Tongkat Ali products have been sold in the market usually formulated into tea, capsule, supplements, coffee and carbonated drinks, which are generally recommended to improve the health condition and libido21,22.
Eurycoma longifolia is an evergreen slow growing plant which can attain a maximum height of 15-18 m long and start to develop fruit at an approximate age of 2-3 years. This plant species is generally believed that it might take up to 25 years before it attains its complete maturation but for commercial use or purposes, the roots are harvested at the age of four years old. The type of leaves is 10-15 cm pinnate type and consisted of about 10-30 ovate-lanceolate leaflets in spirally arranged (Fig. 1a)23. The fruit of Eurycoma longifolia is green when unripe and turns red after ripening (Fig. 1b)24. The flowers are dioecious in nature (male and female reproductive parts are borne or produce on different trees) and produced large panicles.
Traditionally, almost all the parts of this plant are used for various therapeutic purposes but the root part is the most valuable components that contained the highest concentration of quassinoid25. The quassinoid form the major bioactive which are found in this plant species26. Apart from the quassinoids, flavonoids, which are present in some medicinal plants such as Melastoma decemfidum27, Justicia gendarussa28, Orthosiphon stamineus29 have also been reported to be present in Eurycoma longifolia30. Flavonoids are very important phytochemical or secondary metabolites which have strong antioxidant, anti-allergic, anti inflammatory and anti carcinogenic properties31.
| Fig. 1(a-b): | (a) Ovate-lanceolate leaflets spirally arranged and (b) Foliage and cluster of ripe fruits24 |
The valuable components are used for the treatment of various ailments such as sexual insufficiency, dysentery, aches, glandular swelling, persistent fever3. Traditionally, the Tongkat Ali root can be consumed as a tea by boiling the root. This practice is not always easy due to the bitter taste, therefore it is suggested to mix with additives such as honey, sugar crystal and dates in order to make more palatable. According to experience traditional healers, the bitter taste of this plant species is the one responsible for their efficacy and this due to the presence of secondary metabolite3.
CHEMICAL COMPOSITION
Numerous types of compounds have been isolated from E. longifolia roots such as quassinoid, coumarins, anthraquinone, triterpenes, alkanoids, flavonoid, steroids and other types of secondary metabolites2. Eurycomanone and eyrycomanol, two highly oxygenated quassinoid are the most abundant bioactive compounds present in root and were first isolated by Darise et al.32 (Fig. 2). Other than that, three types of squalene-type triterpenes which are eurylene, 14-deacetyl eurylene and longilene peroxide were isolated from the root and determined by spectroscopic, X-ray and chemical analysis33.
Kuo et al.34 demonstrated the isolation of 3 new compounds (n-pentyl â-carboline- 1-propionate, 1-hydroxy-9-methoxycanthin-6-one and 5-hydroxymethyl-9-methoxycanthin-6-one) and 19 known β-caroline alkaloids from the roots of E. longifolia and characterized them by 1D and 2D NMR and by mass spectral data. Out of the three β-caroline lkaloids compound isolated, two of them (1-hydroxy-9-methoxycanthin-6-one and 1-hydroxy- 9-methoxycanthin-6-one) were found to exhibit a significant cytotoxity effect against human lungs and breast cancer (MCF-7). Morita et al.35 also isolated two types of Biphenylneoligans (2-hydroxy-3,2’,6’-trimethoxy-4’-(2,3-epoxy-1-hydroxypropyl)-5-(3-hydroxy-l-propenyl)-biphenyl and 2-hydroxy-3,2’-dimethoxy-4’-(2,3-epoxy-1-hydroxypropyl)-5-(3-hydroxy-1-propenyl)biphenyl) from roots part.
Three form of quassinoid (Eurycolactone D, E and F) were also isolated from the root part as reported by Ang et al.36. The isolated chemicals were determined by spectroscopic method and further confirmed using X-ray crystallography. Miyake et al.37 also isolated 24 types of quassinoid from the stem part and discover 10 new compounds. The new types of quassinoid compounds found include two eurycomanone-type C20, one C19, one klaineanone-type C20 and six eurycomalactone-type C19 quassinoids. Epoxy group type of eurycomalactone quassinoid was first observed in this study. Evaluation of all the compounds isolated have shown to contain cytotoxicity properties against HT-1080 human fibrosarcoma cell line and the result showed that the entire compound isolated possesses cytotoxicity activity.
The use of various types of chromatographic separation, different forms of quassinoid (pasakbumin-C, 13α, 21-epoxyeurycomanone and eurylactone A) have been detected in roots of E. longifolia as reported by Huyen et al.38. Following the characterization by 1D-NMR and 2D-NMR, a new C19-quassinoid-type glycoside was also encountered for the first time as reported by Bedir et al.39. This new C19-quassinoid type (Eurycomaoside) which possessed the C (1)-glycosidation site was isolated from the roots part.
| Fig. 2: | Chemical structures of some of biological active compounds of E. longifolia |
The structure of this compound was determined using a combination of 1 and 2 dimensional NMR technique which includes 1H-13C-heteronuclear correlation spectroscopy (HMQC), 1H-1H-correlation spectroscopy (COSY) and 1H-13C-heteronuclear multiple-bond correlation spectroscopy (HMBC) as well as HR-ESI-FT-MS (High Resolution Electro Spray Ionization Fourier Transformation Mass Spectrometry) data.
The GC-MC analysis of methanol and water extract using solid-phase extraction showed a total number of 13 different types of volatile compounds present in 12 E. longifolia samples which were either in methanol and water-freeze or spray dried40. The results showed that higher number of 83 volatiles were extracted using a freeze dried extract technique compared to 28 volatiles using spray dried methods40. The quantification of volatile compounds detected were analysed by correlating the sensor response with the volatile compounds. The result showed that a total number of 8 volatiles are common in all the samples and these were used as predictable variables. The correlation between the GC-MS and QCM showed the presence of different types of compounds, including 3-phenoxy-1-propanol, 2-phenoxy-ethanol, 4-ethynyl-4-hydroxy-3, 5, 5-trimethyl-2-cyclohex-1-enone, benzoic acid, acetic acid and octanoic acid. They concluded that this novel technique (Nose approach) is very simple when compared to the GC-MS traditional method which is expensive. This approach has been shown to be very sensitive to the extent that it can detect small changes in the volatiles; therefore it can be useful for rapid detection and evaluation of herbal volatiles. The summary of the most important chemical constituent found in E. longifolia plant with their various activities is shown in Table 2 while their chemical structures are shown in Fig. 2.
QUALITY CONTROL OF E. longifolia HMP
The numerous therapeutic effects of E. longifolia have resulted in the development of various types of it HMP in the market, thus becoming a lucrative business. Among the different types of pharmacological properties exhibited by E. longifolia, the aphrodisiac properties such as the ability to improve Erectile Dysfunction (ED), male libido and sexual prowess21,69 are among the driving force that made their HMP to be highly demanding and patronized by consumers. Different types of clinical10,70,71 and animal studies68,72,73 have shown that E. longifolia extract plays a very vital role in enhancing sexual reproductive activities. Still on the aspect of clinical studies, it was recently reported that E. longifolia can be used to restore testosterone levels, therefore making it to be a natural alternative for the treatment of Testosterone Deficiency Syndrome (TDS)74. The TDS is a deficiency which arises as a result of decline in serum testosterone level leading to fatigue, low libido, osteoporosis or erectile dysfunction and more than 80% of people suffering from this deficiency are as a result of old age.
The HMP are generally made up of varieties of self-prescribed preparation of plant origin which can be categorised as either food and beverages or dietary supplements75,76. As such, they existed in a form of either mono herbal component or polyherbal (mixed with other components such as in coffee and beverages).
In Malaysia, it is reported that there are more than 140 licensed manufacturers of E. longifolia products and these produces over 200 products together with it neighbouring countries, Indonesia and Thailand77. Due to the massive and rapid increased in the consumption of E. longifolia HMP, there are over 300 popular coffees, tonic, candies and beverages premixed with root extract commercially available in Malaysia alone to improve health and libido24. The popularity and consumption of E. longifolia HMP are increasing tremendously therefore, various safety-related issues to the public is of major concern and needs to be addressed. The common perception by people which is sometimes portrayed by mass media in marketing and health advertisement that natural or plant based product are completely safe are untrue and misleading. It is a complete error in the sense that besides their adverse effects, they can sometimes produce life-threatening consequences. This common assumption of "natural" is "harmless" has resulted in a massive increase in the patronizing of these HMPs in developed countries in recent decades75. The safety information about most of the constituents of medicinal plants responsible for their pharmacological and possible toxic effects is mostly scanty. This is due to the fact that out the humongous number of medicinal plants that existed as herbal remedies, only few numbers of them have been subjected to in-depth analytical, pharmacological and clinical studies.
The misconception of tagging everything natural as being safe or harmless has resulted in less regulation control in the market of such products78. Consequently, quality control issues such as contamination by pathogenic microbials or natural toxins, misidentification, adulteration with toxic compounds, standardization of dose, methods of processing and product uniformity are becoming major problems for medicinal plants and their HMP78,79.
Several literatures80-82 have shown that contamination of HMP with certain toxic substances can result to a wide range of adverse or unexpected side effects and this can result in serious injuries and even death.
| Table 2: | Summary of chemical compounds and pharmacological activities of different parts of E. longifolia |
The contamination or adulteration of HMP with heavy metals such as lead, arsenic, mercury, cadmium and copper are always of great concern. However, some of the E. longifolia HMP which are sold in the market have been reported to contain some of these common heavy metals.
In the recent years, there has been a big rise in herbal-based product consumptions due to their claimed health benefits both locally and globally. However, due to the challenging financial climate, adulteration of HMP with closely related species can be risky as this can result to unexpected side effects82. The HMP available to consumers in the marketplace substituted with these undeclared ingredients on the labels might pose a major threat to consumer safety. Adulteration in some cases can be lethal if it is substituted with toxic adulterants.
Thus, consumers have become increasingly concerned about the HMP authenticity and the quality of the purchased. High incidence of mislabelling reports had also increased the customer awareness towards this problem. The consumer may have the misconception that natural or plant base products are safe or harmless has resulted in less regulation control in the market sample of such products73. Consequently, quality control such as contamination by pathogenic microbes or natural toxins, incorrect species used, adulteration with toxic compounds/adulterants, standardization of dose, methods of processing and product uniformity are becoming major problems for medicinal for HMP73, 74. The HMP contamination with heavy metals is often common, especially in Asian countries83. Analytical investigation has shown that contamination of HMP with heavy metals can occur because of (1) Environmental condition where the plant are grown, (2) During drying and processing step, (3) Improper storage and transportation condition and finally manufacturing process which the HMP are produced. Several literatures75,77 have shown that consumption HMP with certain toxic substance can result to a wide range of adverse or unexpected side effects and this can result in serious health problems, even death.
Ang et al.84 reported the presence of lead in a large number of E. longifolia HMP consisting of both Malaysian Registered Products (MRP) and Malaysian Unregistered Products (MUP) obtained from the various pharmaceuticals store, health store using atomic absorption spectrophotometer. Among of the 100 HMP analyzed, 8% of sample contained lead between 12.24-20.72 ppm and this does not complied with the quality requirement for traditional medicine in Malaysia which should not exceed 5 ppm as stipulated by the Drug Control Agency (DCA). The further disturbing issue, none of these HMP was registered with DCA. The high amount of lead is toxic to the human body as it may affect some vital tissues such as kidney, nervous system, liver, bone, heart and reproductive system85.
One year later, Ang et al.86 reported another heavy metal contamination, mercury in a number of E. longifolia HMP. From their reports, 36% of the over-the-counter HMP sold possess higher (0.52-5.30 ppm) mercury content and this was not in conformity with the standard quality requirement of Malaysia which should not exceed 0.5 ppm. The report of the study also shows that 5% out of the 36% HMPs were not registered with DCA. High concentration of mercury in human body is detrimental as it can cause many damages such as nervous tissue and brain damage, damages of chromosomes and damages of the reproductive functionality85.
Therefore, the quality of E. longifolia HMP sold in the market needs to be regulated from manufacturing processes until production; hence protecting the consumer. This can be effectively conducted by firstly regulating the correct use of raw material used in the production87. This will ensure that the customer gets the best of their purchased. From plant cultivation to the point of production of the HMP, there are several factors which can influence the quality of these HMP. Because of the high demand of E. longifolia HMP in the market, unscrupulous suppliers may perform some unethical activities. Adulteration is the addition of inferior, improper or extraneous ingredient to make an impure HMP and this has generated a lot of questions about their efficacy and safety. This is due to the fact that the quality assurances of any HMP solely relied on the proper authentication and correct use of raw plant materials (ingredients). The breakage of this essential rule as a result of adulteration can lead to undesirable or adverse effect after consumption. According to the WHO, this activity is not only a fraud but a threat to the consumer health88.
The different types of adulteration which are commonly observed in E. longifolia HMP include; (1) Use of stem instead of the roots as most of the active compounds are found in roots part, (2) Addition of the high amount of fillers such as rice and wheat and finally and (3) Include less concentration of E. longifolia extract/raw material to gain more profit89,90 . In some cases, E. longifolia HMP sold in the market may not even contained any bioactive compounds which are responsible for the therapeutic properties of the plant91. Therefore, authentication of E. longifolia HMP is the most basic quality step to ensure that they have met the required pharmacopeia standard as this is very crucial towards quality control.
In a study conducted by Norhidayah et al.90, investigation of the authenticity from 41 E. longifolia HMP showed that 12 were classified as international products (IR), 16 are Malaysian Registered Product (MRP) and 13 Malaysian Unregistered Product (MUP) using HPLC analysis. The results from Norhidayah et al.90 study demonstrated that eurycomanone was only detected in 58.9% of the total HMP. The Malaysian standard (2011) stated that the level of eurycomanone in E. longifolia should be approximately 0.8-1.5 % (w/v)76,90. Out of the 24 HMP which contained the standard, only 11 (0.84-8.48% w/v) fulfilled the Malaysian standard criteria while 9 (1.6-8.48 % w/v) of the HMP is above the set criteria. Norhidayah et al.90 also proved that out of the eight premixed coffee samples allegedly contain E. longifolia in their label, eurycomanone was not detected.
In the same year, Han et al.92 developed liquid chromatography with tandem mass spectrometry method to validate or confirmed the presence of bioactive compound in E. longifolia HMP (capsule and tablets) using six major quassinoids (eurycomanone, 13, 21-dihydroeurycomanone, 13α (21)-epoxyeurycomanone 14,15β-dihydroxyklaineanone, eurycomalactone and longilactone). It is interesting to note that out of the seven HMP sample tested in their study, the target bioactive compounds in two of the samples (TB2 and TB3) were either present below the detection or detected with only trace amount while other contain great amount of the quassinoids. This inconsistency or variation in the amount of bioactive compounds present among the product is alarming due to the fact that E. longifolia extract was reported to be toxic in male rats93. Therefore, for standardization purpose, consistency of the bioactive compounds needs to be ensured.
The absent or present of poor quassinoid content in some of the E. longifolia HMP have resulted in the addition of other unlisted illegal drugs compounds. The detection of synthetic phosphodiesterase-5 inhibitors (PDE-5i) such as sildenafil,vardenafil and tadalafil in HMP that are tagged or labelled as natural products that enhances sexual libido and increase testosterone level is becoming interestingly common94-96. A recent survey by Gilard et al.97 confirmed that 61% from 150 HMP advertised to enhance sexual libido were found to be adulterated with synthetic PDE-5i, this finding represents an alarming risk to the consumers health.
However, even though the uses of some synthetic PDE-5i were approved in the treatment of erectile dysfunction but their usage must be under medical supervision. This is due to the fact that their interaction with certain drugs can cause negative effects. Hyperlipidemia, hypertension and ischemic heart diseases are mostly treated with drugs containing nitrates, therefore patient suffering from such ailment should not take synthetic PDE-5i as this may drastically lower their blood pressure98. Therefore, the unknowingly consumption of such HMP that contain undeclared synthetic PDE-5i may pose a serious health threat to the patient.
As a result of the growing trend towards intentional adulteration nowadays, some studies have revealed the adulteration of E. longifolia HMP with certain type’s synthetic drugs. Said et al.99 reported detection of sildenafil-like compound in some of the E. longifolia HMP purchased online. Champagne and Emmel100 also reported such kind of deliberate adulteration and this type of situation is not only a fraud but a threat to public health.
Therefore the misconception among the consumers regarding natural terms that are safe should be the utmost priority. The reality is that "safety" and "natural" are not synonymous101. However, to overcome these hurdles despite the facts that each country has its own unique regulatory body, the government regulatory authorities involved should enact new policies in addition to the existing ones so as to ensure that Eurycoma longifolia HMP, which are introduced into the market are safe for public consumption. As quality control or quality assurance relied on the use of authentic raw materials, the effort of government in addition to research institutes and herbal industries should voluntarily establish the use of advance techniques as quality control standard. A reliable molecular approach which utilizes the use of DNA barcoding in addition to other techniques can also be used for the authentication of raw materials used in the manufacturing of Eurycoma longifolia HMP as most of them are sourced from the wild102.
As suggested by Sahoo et al.103, quality issue of the HMP arises from cultivation until to the manufacturing and finally circulation. The quality assurance of the HMP can also be controlled through the implementation of the various agricultural practices recommended by WHO to ensure their efficacy and safety103. Agricultural practice such as Good Agricultural and Collection Practices (GACP) is a very critical in the collection of herbal materials and inadequate of quality control can be overcome if properly adopted. External issues which involved misidentification of plant species, contamination by heavy metals and other toxic substances can be effectively minimized through the adoption of GACP practice by ensuring that the product collected are safe for consumption. Another great advantage of implementing GACP is that it will help to assist in ensuring that the quality of the starting herbal materials is reproducible104.
Since after the issue of GACP guideline by WHO in 2003 for medicinal plants, other countries such as Japan, China and European unions have also developed their national and regional GACP in order to effectively improve that quality of HMP which are disseminated to the public105. Their developed guidelines ensures that the soil and water which they will use for the cultivation of the medicinal plants are within the standard limit or free from toxic substances such as heavy metals, pesticides, microbes or mycotoxins. Implementation of Good Manufacturing Practices (GMP) together with post marketing quality assurance surveillance can also improve the quality of the E. longifolia HMP sold in the market. The initiation of pharmacovigilance practice which is a very essential and common tool used for assessing the safety of synthetic drugs can also be developed for HMP with little modifications. The pharmacovigilance will assist in providing reliable information about the safety or risk of using E. longifolia HMP. This will promote safe use of E. longifolia HMP and protect the public from the adverse effect of the HMP. The establishment of precise systemic methodologies and clinical trials by researchers, manufacturers and various regulating bodies as reported by Mukherjee et al.78 can assist in the production of high quality E. longifolia HMP which will be are introduced to the market. This will help to increase the consumers confident in the consumption of E. longifolia HMP.
CONCLUSION
The proper tightening and implementation of the existing regulation in the various countries must be the way forward by ensuring that E. longifolia HMP that circulates in the market are of good quality, safe, efficacious and sustainable. Furthermore, the use of pharmacovigilance to provide reliable information about the safety and risk of using E. longifolia HMP coupled with the proper implementation of various agricultural practices, DNA-based and metabolite-based method will help to minimize problems associated with adulteration, contamination and substitution. This will also make the E. longifolia HMP more credible, reliable and acceptable, therefore increasing consumer’s confidence.
SIGNIFICANCE STATEMENT
This study highlighted the importance of different bioactive compounds present in Eurycoma longifolia and the consequences that might arise as a result of compromising the quality of their HMP sold in the market. This study proposed the need to have stringent regulatory measures on such HMPs, implementation of various agricultural practices and the use of multidisciplinary approaches to their quality assurance and quality control. This will tremendously help to reduce the exposure of consumers to various health risks.
ACKNOWLEDGMENTS
The authors would like to thank the Ministry of Education Grant Tier 1; Q.J130000.2545.05H97, Fundamental Research Grant Scheme (R.J130000.7845.4F893) and all staff members of Plant Biotechnology Laboratory, Faculty of Biosciences and Medical Engineering, UTM for supporting.