Background and Objective. Tibetan medicine is one of the earliest-known traditional medicines. This study aimed to evaluate the antioxidant, cytotoxic, and antibacterial potential of ethanolic extracts of nine common Tibetan formulations. Materials and Methods. An open-ended and semistructured questionnaire was used for an ethnomedicinal survey of the Tibetan formulations practiced in four Tibetan refugee settlements in Gandaki Province, Nepal. Based on the ethnomedicinal survey data, commonly used nine formulations were selected (Aru-18, Basam, Dadue, Dashel, Mutik-25, Raab Ga Yangzin Tea, Serdok-11, Sugmel-10, and Yungwa-4) to test biological activities. Antioxidant activity was evaluated using the 2,2-diphenyl-1-picryl-hydrazyl (DPPH) radical scavenging method. The cytotoxicity was examined by using the Allium cepa L. root tip meristem model. Similarly, the antibacterial effect was assessed by using well diffusion and broth dilution methods. Results. An ethnomedicinal survey showed a total of 52 Tibetan formulations were generally used by respondents for common diseases such as stomach disorders, diabetes, and migraine. From the antioxidant activity test, Sugmel-10 showed the highest DPPH free-radical-scavenging activity (IC50 1.8 μg/ml) and Yungwa-4 showed the lowest activity (IC50 5.2 μg/ml). Also, from the cytotoxic activity, the A. cepa root meristem model exhibited significant dose- and time-dependent growth suppression in Basam, Dadue, Mutik-25, and Serdok-11 as compared with cyclophosphamide standard drug. Similarly, Basam showed a good antibacterial effect having MIC 20 mg/ml and MBC 100 mg/ml against Enterococci faecalis. Conclusion. Research showed that Tibetan people preferred Tibetan formulations for the treatment and mitigation of several diseases. The result of antioxidant, cytotoxic, and antibacterial activities experimentally justified the ethnomedicinal value of nine common formulations (Aru-18, Basam, Dadue, Dashel, Mutik-25, Raab Ga Yangzin Tea, Serdok-11, Sugmel-10, and Yungwa-4). To the best of our knowledge, this study was performed for the first time in Nepal. Results from this preliminary study open the door to the scientific world to perform extensive pharmacological studies for designing and developing new therapeutic agents.
Tibetan medicine (TM) is one of the earliest-known traditional medicines, and its history goes back approximately 2,500 years. In this system of medicine, a Tibetan doctor formulates an anticipative diagnosis and personalized treatment plan, where the treatment may last several months to years for chronic diseases . In TM, particular treatment is codified in the form of sacred texts or pharmacopeia elucidated with the Buddhist understanding of herbal remedies . There are more than 20 different dosage forms in traditional TM such as pills, powders, decoctions, lotions, ointments, and medicinal liquors . Dried raw materials are ground, mixed homogenously, and ultimately pressed into pills, powder, or decoctions. Mantras are chanted to enhance the potency of the remedy in the course of blending . In a particular formula, ingredients are blended as a dry powder where galenical forms are chiefly pills and medicinal powders in comparison to medicinal butter, plasters, and decoctions. An entire formula can be considered as a pharmacologically active entity with distinct pleiotropic effects . The dosage form of this system constitutes several ingredients, which is through combinations of up to 108 or more ingredients. The governing research concerning multi-ingredient formula came from the Padma, a Swiss pharmaceutical company that develops standardized herbal formulas that originated from Tibetan medical knowledge [6–9].
Along with TM, traditional Iranian medicine (Persian medicine), traditional Chinese medicine (TCM), and Ayurveda are very popular in Asian countries. All these traditional systems of medicine use herbal drugs or extracts, acupuncture, massage, diet therapy, physical activity, and exercise. Like TM, Persian medicine, TCM, and Ayurveda follow humoral theory. Historically, different formulations and potential herbs are used in the abovementioned traditional system of medicines as evidence-based therapy. The integration of their principles, techniques, medication, and knowledge with modern medical sciences is the field of tremendous ongoing efforts and interests to develop new therapeutic options in current medicine. The effective management as suggested by traditional medicines regarding the human body is based on maintaining balance in body fluids and temperament, along with therapeutic and supportive strategies [10–13].
Antioxidants antagonized the damaging effects of free radicals and helped to prevent or repair that deleterious phenomenon in living cells . Bioactive phytochemicals present in traditional medicines possess antimutagenic, anticancer, and antioxidant properties that provide a protective effect against various kinds of cellular injury. For instance, phenolic compounds (caffeic acid and ρ-coumaric acid) and flavonoids (kaempferol) are responsible for antioxidant activity; terpenoides (ρ-cymene and γ-terpinene) and essential oil (cuminaldehyde) are liable for antimicrobial effects [15, 16]. Also, natural antioxidants present in various plants decrease oxidative damage and help in inhibiting aging, mutagenesis, and carcinogenesis considering their radical scavenging activities . The A. cepa root tip meristem model has been extensively used for the assessment of antimitotic and cytotoxic properties [18–22] by utilizing the growing roots of A. cepa. The cell division in its meristematic cells resembles normal human cancer cell division. Thus, these meristematic cells can be used in the analysis of drugs with possible human anticancer activity . Using plant extract to treat infections is an ancient practice in traditional medicine. For this intent, humans have used natural products derived from plants, animals, and microbial sources for millennium either in the crude extracts or pure forms . Plant secondary metabolites are predominately liable for their antimicrobial activity . Likewise, many animals and their products have been used in traditional medicines across numerous medicinal systems [26–28]. In recent years, indiscriminate uses of antibiotics have generated the problem of antibiotic resistance. Similarly, the quest for new antimicrobial agents is a worldwide concern as herbal medicine from natural sources showed lesser side effects than synthetic medicines. Furthermore, some traditional TM may suggest promise in clinical treatments as plants, animals, trace elements, and minerals are the abundant sources used in such traditional medicines including biologically active substances and amino acids. Plants used are mostly cold and drought resistant, and they perform thorough photosynthesis .
Tibetan traditional medicines are being practiced more in the Himalayan region and are recommended by Amchis (Tibetan medicinal practitioners) in the northern belt of Nepal, the border of Tibet (autonomous region of China). Tibetan people residing in the country and families in the Tibetan refugee camp more preferred these formulations. This system of medicine is not normally assessed by the people in the central or southern parts of the country. This may be due to the limited number of general health practitioners available in that region or due to the insufficient scientific evidence of these traditional medicines.
Antioxidant, cytotoxic, and antibacterial studies of nine common Tibetan formulations (described briefly in Table 1) have not yet been studied and justified. Therefore, in this study, we performed the ethnomedicinal survey, and based on the survey data, we collected nine formulations and evaluate their effectiveness against antioxidant, cytotoxic, and antibacterial activity.
Aru-18 (myrobalan 18)
Carthamus tinctorius, Caesalpinia bonducella, Cupressus torulosa, Elettaria cardamomum, Eugenia jambolana, Malva verticillata, Mucuna prurita, Rubia cordifolia, Swertia chirata, Symplocos crataegoides, Terminalia chebula, Verbascum thapsus, vermilion, and crabshell
Inflammation of the kidney; kidney disorder giving rise to stooping; pain in hip and waist region; imbalance of kidney channels
2-3 g once daily in the afternoon with warm water
Basam (medicinal butter)
Angelica species, Asparagus spinosissimus, Emblica officinalis, Mirabilis himalaica, Polygonatum cirrhifolium, Terminalia belerica, Terminalia chebula, Tribulus terrestris, clarified butter, honey, and milk
Kidney diseases; leg cramps; tonify body, promotes longevity; pain in the bones, hip joint, and lower back muscles; male-specific diseases such as impotence, prostatitis, and decreased libido
1–4 g daily with warm water
Carthamus tinctorius, Crocus sativas, Dracocephalus tanguiticum, Inula racemosa, Saussurea lappa, calcium carbonate, iron powder, and mineral pitch
Liver disorder, gastric problem, food poisoning, indigestion, chronic fever, colic pain, eye problem, all kinds of chronic diseases, and general tonic
1-2 pills daily in the morning or evening with warm water
Aconitum heterophyllum, Aconitum orochryseum, Adhatoda vasica, Amomum subulatum, Bainbusa textilis, Beaumontia grandiflora, Carthamus tinctorius, Commiphora mukul, Corydalis stracheyi, Crocus sativus, Cynanchum thesioides, Dracocephalus tanguiticum, Elettaria cardamonium, Emblica officinalis, Eugenia caryophyllata, Cerinthe gymnandra, Herpetospermum pedunculosum, Inula racemosa, Lagotis kunawurensis, Meconopsis species, Moschus moschiferus, Myristica fragrans, Pedicularis oliveriana, Picrorhiza kurroa, Piper longum, Pterocarpus santalinus, Punica granatum, Rheum spiciforme, Santalum album, Saussurea costus, Saussurea lappa, Saxifi aga umhellulata, Strychnos nux-vomica, Syzigium aromaticum, Taraxacum officinale, Terminalia chebula, Ursus thibetanus, Vincetoxicum sibiricum, calcitum, mineral pitch, and smithsonite (calamine)
Gastrointestinal diseases; gastrointestinal tumor; cholecystitis; hepatitis; nausea; belching; constipation; chronic diseases of the stomach, liver, and skin; anti-inflammatory action in the gastrointestinal tract
1-2 g in the morning or evening with warm water
Mutik-25 (Pearl 25)
Amomum subulatum, Bambusa textilis, Bos taurus domesticus, Carthamus tinctorius, Cinnamomum zeylanicum, Cuminum cyminum, Emblica officinalis, Elettaria cardamomum, Eugenia caryophyllata, Gmelina arborea, Margaritum, Myristica fragrans, Nigella sativa, Piper longum, Polygonum aviculare, Potamom yunnanensi, Pterocarpus santalinus, Punica granatum, Saussurea lappa, Terminalia belerica, Terminalia chebula, Malva verticillata, Moschus moschiferus, Santalum album, and vermiculite
Hypertension, stroke neuralgia, hemiplegia, palpitation, tranquilizer, facial paralysis, unconsciousness, delirious mania
1-2 pills one hour before or after meals with warm water daily
Raab Ga Yangzin Tea (herbal tea)
Carthamus tinctorius, Elettaria, Rosa bronunii, Rubus hoffmeiteriannus, Symplocos paniculata, and Zingiber officinalis
Provides body energy; promotes skin health; appetite; relaxation; assists in cold, flu, and poor digestion
Pour hot water on the teabag, infuse for few minutes, then relish it purely or with salt or sugar
Carthamus tinctorius, Crocus sativa, Cuminum cyminum, Emblica officinalis, Herpetospermum caudgerum, Myristica fragrans, Punica granatum, Rosa bronunii, Saussurea lappa, Terminalia chebula, and mineral pitch
Acute and chronic diseases of the liver and gall bladder (cirrhosis, cholecystitis, and gallstone disease); protects liver cells; restores the function of the pancreas; normalizes metabolism
2 pills daily with hot water
Sugmel-10 (cardamom 10)
Caesalpinia bonducella, Elettaria cardamomum, Eugenia jambolana, Hedychium spicatum, Malva verticillata, Piper longum, crab shell, and sodium chloride
Kidney disorders, removes kidney stones, clears obstruction of the urinary tract, removes tumors and stones from the urinary bladder
2-3 g daily at night with hot or warm water
Yungwa-4 (decoctions of turmeric)
Berberis dictyophylla, Curcuma longa, Emblica officinalis, and Tribulus terrestris
Antipyretic, diuretic, inflammation of the urethra
3–5 g decocted to one-third water level and taken twice daily