Article Information

Kenneth C. Ugoeze¹, Nora Amadi¹, Ngozi A. Okoronkwo², Sunday O. Abali¹, Kennedy E. Oluigbo³, Bruno C. Chinko^{*, 4}

¹Department of Pharmaceutics & Pharmaceutical Technology, Faculty of Pharmaceutical Sciences, University of Port Harcourt, Port Harcourt, Nigeria.

²Department of Zoology & Environmental Biology, Michael Okpara University of Agriculture, Umudike, Abia State, Nigeria.

³Department of Clinical Pharmacy and Biopharmaceutics, Faculty of Pharmaceutical Sciences, Enugu State University of Science and Technology, Agbani City, Enugu, Nigeria.  

⁴Department of Human Physiology, Faculty of Basic Medical Sciences, University of Port Harcourt, Port Harcourt, Nigeria.

*Corresponding author: Bruno C. Chinko, Department of Human Physiology, Faculty of Basic Medical Sciences, University of Port Harcourt, Port Harcourt, Nigeria

Received: 23 December 2022; Accepted: 05 January 2023; Published: 14 January 2023

Citation: Kenneth C. Ugoeze, Nora Amadi, Ngozi A. Okoronkwo, Sunday O. Abali, Kennedy E. Oluigbo, Bruno C. Chinko. GC-FID guided Identification and Quantification of detectable Phytochemicals in selected Commercial Chamomile Herbal Tea. International Journal of Applied Biology and Pharmaceutical Technology 14 (2023): 01-11.

Abstract

Keywords

GC-FID identification and quantification, Phytochemical, Chamomile, Herbal tea

Article Details

1. Introduction

Phytochemicals are considered plant-based bioactive chemicals with various therapeutic and nutritious benefits [1-3]. They are classified as primary (PM) or secondary metabolites (SM). The PM comprises the common sugars, amino acids, proteins, purines and pyrimidines of nucleic acids, chlorophylls, etc. whereas the SM are the plant chemicals such as alkaloids, terpenes, flavonoids, lignans, plant steroids, saponins, phenolics, glycosides, etc. [1].   Due to their antimicrobial, anti-inflammatory, anthelmintic, anti-carcinogenic, anti-genotoxic, anti-proliferative, anti-mutagenic, anti-allergic, immune-modulatory and anti-oxidative properties, the SM could play protective roles against pathogens or damaging disorders [4-9]. There are over 8000 natural flavonoids [5] with their health-based biotic actions ascribed to their antioxidant influence [6, 7]. The antioxidant and anti-inflammatory actions of the flavonoids aid in toxin-mediated stress and protracted infection inhibition [10]. Flavonoid-rich foods include all nourishments of plant basis, predominantly tea, fruit, vegetables, grains, legumes, nuts, and wine [11, 12].

Tea is processed from the vegetative parts of Camellia sinensis [13]. They are used as a medicinal drink by two-thirds of the globe [14] and have been designated as “safe” by the US Food and Drug Administration's (FDA) list of compounds generally recognized as safe (GRAS) [15]. Normally, Tea refers to dried leaves processed from leaves of Camellia sinensis (true tea).  Other infusions got from parts of other suitable plants are also noted as tea, but, are called tisanes or herbal tea, hence, there are true or traditional tea and herbal tea [12, 13]. True tea is grouped as green, black/dark, white, yellow and oolong which is established on the extent of oxidation of the leaves [16]. Green tea is a non-fermented tea [17], they are processed from freshly harvested vegetation of the tea plant without fermentation after withering, steaming or pan firing, drying and grading to retain its content of polyphenols [15, 16, 18]. On the other hand, black tea is wholly fermented tea, processed with polyphenol oxidase converting its polyphenols into a series of new products like theaflavins, theaflagallins and thearubigens obtainable in black tea  [16, 18]. Oolong tea is a semi-fermented tea gotten when leaves are wilted in the sun and moderately battered to attain brief partial oxidation resulting in an intermediate product, the oolong tea [18-20].

Herbal tea or tisanes implies any infusions taken as a drink derived from other parts of vegetation other than Camellia sinensis [6, 15]. They are often caffeine free compared to traditional teas. It could be gotten from plants like chamomile, rose hip, ginger, turmeric, valerian, hibiscus, peppermint, etc. [21]. Chamomile is one of the early therapeutic herbs with standardized forms prepared from dried flowers of Matricaria species and of the daisy family, Asteraceae/ Compositae [22]. Two common varieties are the German Chamomile (Matricaria chamomilla) and Roman Chamomile (Chamaemelum nobile) [23]. Their preparations are used to manage several ailments like hay fever, inflammation, muscle spasms, menstrual disorders, insomnia, ulcers, wounds, gastrointestinal disorders, rheumatic pain and haemorrhoids while their essential oils are applied widely in cosmetics and aromatherapy [24, 25]. Their therapeutic influence is largely based on their dried flowers' terpenoid and flavonoid content [24]. Chamomile herbal tea is essentially caffeine-free and is believed to be a calming and soothing effect which promotes sleep and reduces anxiety [26, 27]. They have also been found to be effective in treating injuries, ulcers, eczema, gout, skin irritations, bruises, burns, canker sores, neuralgia, sciatica, rheumatic pain, haemorrhoids and mastitis [25, 28]. Aside from their use as tea, and tincture, they are used as poultices and tincture [29]. Some of the flavonoids found in chamomile are apigenin, quercetin, and patuletin [30, 31]. Apigenin is known to bind to benzodiazepine receptors in the brain to enhance sleep and diminish insomnia [32, 33]. Similarly, their essential oils are used in aromatherapy to enhance sleep and relieve anxiety [34].

Despite the aforementioned benefits and safety margin of chamomile, there has been no confirmation of their reaction to other medications or their safety among children and pregnant women. Therefore, the present study aims to identify and quantify the phytochemicals in selected commercial chamomile herbal tea using Gas Chromatography with flame ionization detection (GC-FID) methods.

2. Materials and Methods

2.1 Procurement of Samples

Five brands of chamomile herbal tea (CHT) were randomly sourced from retail outlets in Port Harcourt, Nigeria and coded CHT-A, CHT-B, CHT-C, CHT-D and CHT-E. Also, a crude unpackaged dry chamomile flower was sourced from a retail outlet in Abuja and coded as CHT-F while its oil extract was coded as CHT-G.

2.1 Preparation of samples and extraction of phytoconstituents

A teabag of each CHT was macerated in 200 ml of ethanol (Honeywell, Germany) for 48 hours. It was filtered and evaporated to obtain the extract. A 0.1 g of the extract was re-extracted in 25 ml of ethanol in a test tube immersed in a water bath (60 ^o C) for 90 mins. It was transferred to a separatory funnel. The test tube was washed in steps with 20 ml of ethanol, 10 ml of cold water, 10 ml of hot water and 3 ml of n-hexane (BDH, England) respectively and transferred into the separatory funnel. The extracts were pooled and washed thrice with 10 ml of 10 % v/v ethanol-aqueous solution. The solution was dried with anhydrous sodium sulphate (Sigma-Aldrich, USA) and the solvent was evaporated. The sample was solubilized in 1000 ul of pyridine (Sigma-Aldrich, USA), of which 200 ul was transferred to a vial for analysis [35, 36].

2.2 Quantification of the phytoconstituents by GC-FID

The analysis of phytoconstituents was carried out on a BUCK M910 Gas Chromatography fitted with an HP-5MS column (30 m in length × 250 μm in diameter × 0.25 μm in thickness of film). Spectroscopic detection by GC–FID involved an electron ionization system which used high-energy electrons (70 eV). Pure helium gas (99.995 %) was employed as the carrier gas with a flow rate of 1 mL/min. The early temperature was set at 50 –150 °C with an increasing rate of 3 °C/min and a holding time of about 10 min. Later the temperature was amplified to 300 °C at 10 °C/min. A 1µl of the prepared 1% of the extracts diluted with respective solvents was injected in a splitless mode. The relative quantity of the chemical compounds present in each of the extracts was expressed as a percentage based on the peak area produced in the chromatogram [36, 37]. Bioactive compounds extracted from the respective batches of extracts were identified based on the GC retention time on the HP-5MS column and matching the spectra with the computer software standards data (Replib and Mainlab data of GC–FID systems).

3. Results

3.1 GC-FID Quantified Phytoconstituents of selected Chamomile Herbal Tea

Table 1: Composition of the phytochemical constituents detected in CHT-A

Table 2: Composition of the phytochemical constituents detected in CHT-B

Table 3: Composition of the phytochemical constituents detected in CHT – C

Table 4: Composition of the phytochemical constituents detected in CHT-D

Table 5: Composition of phytochemical constituents detected in CHT-E

Table 6: Composition of the phytochemical constituents detected in CHT-F

Table 7: Composition of the phytochemical constituents detected in CHT-G

4. Discussions

The results obtained from the GC-FID of selected chamomile herbal tea and dry chamomile flower and essential oils are summarised in Tables 1 – 7. These results are discussed in this section under flavonoids, alkaloids, glycosides, saponins, tannins, steroids, anti-nutrients and other phenols such as resveratrol.

4.1 Flavonoids

The level of flavonoids detected in the various CHT were as CHT-B > CHT-E > CHT-G > CHT-A > CHT-C > CHT-C > CHT-D > CHT-F. The types of flavonoids identified include proanthocyanin, anthocyanin, flavan-3-ol, naringenin, rutin, flavanones, kaempferol, flavone, catechin, epicatechin, etc. (Tables 1-7). The variation in the composition of these flavonoids from one CHT to the other may be attributed to differences in environment or influences like latitude, longitude, rainfall, temperature and soil quality [38]. Given these differences, and with CHT-B presenting the highest concentration of flavonoids displayed its subgroup of flavonoids as Rutin   > flavonone > flavone > anthocyanin > epicatechin > kaempferol > naringenin > proanthocyanin, etc. (Table 2). Flavonoids have been known to possess immense pharmacological benefits such as anti-oxidative, anti-mutagenic, anti-inflammatory, anti-carcinogenic, antitumour, anti-HIV, antidiarrhoeal, antihepatotoxic, antifungal, antilipolytic, vasodilator, immunostimulant and anti-ulcerogenic properties and enzyme modulatory functions [39, 40]. Rutins have been found in fresh leaves, red wine and tea [12, 41, 42]

4.2 Alkaloids

The composition of alkaloids detected was CHT-A > CHT-D > CHT-G > CHT-E > CHT-B > CHT-F > CHT- C. The various forms of alkaloids detected were ribalinidine, lunamarin, spartein, ephedrine, etc., though, ephedrine was not detected in CHT-A and CHT-G.  Alkaloids are natural products that have heterocyclic nitrogen atoms [43-45]. They found use in the ancient preparation of spices, drugs and poisons. Lunamarin retains anticancer, immunomodulatory, anti-estrogenic and anti-amoebic activities [46] while ribalinidine possesses a radical scavenging influence.

4.3 Glycosides

The composition of glycosides identified in the various CHT were CHT-F > CHT-B > CHT-C > CHT-D > CHT-E > CHT-G > CHT-A with cardiac and cyanogenic glycosides prevailing (Tables 1-7). With the highest level of glycosides detected in CHT-F, the extent of cardiac and cyanogenic glycosides was 4.37 and 14.33 % respectively.  In CHT-B, they were 3.15 and 15.34 % correspondingly whereas it was 3.69 and 11.72 % in CHT-C. Further, it was 3.59 and 9.81% in CHT-D while in CHT-E it was 4.07 and 8.63 %. Only cardiac glycoside was detected in CHT-A and CHT-G at 2.11 and 2.34 % respectively. The cyanogenic glycosides retained higher concentrations where it was detected. Glycosides are plant-based substances comprising of a glucose unit confined to an aglycone like alcohol, phenol or steroid nucleus through a glycosidic bond [47] with potent antibacterial, antifungal, anti-inflammatory, antioxidant, antiviral and anticancer activities [48, 49]. Cardiac glycosides are used in the treatment of cardiac insufficiency [48, 50] by increasing the output force of the heart and decreasing its rate of contractions by inhibiting the cellular Sodium-Potassium-ATPase pump [50]. However, their relative toxicity prevents their extensive application [51]. On the other hand, cyanogenic glycosides which are mostly found in foods including linamarin, amygdalin and prunasin [52] are known to release hydrogen cyanide when chewed or digested [53] resulting in significant cyanide poisoning. However, processing methods, such as peeling, drying, grinding, soaking and fermentation, boiling or cooking have been reported to cause a significant reduction in the cyanogenic glycosides of processed foods [54].

4.4 Saponins

The saponin detected was sapogenin which occurred in these CHTs as CHT-C (4.90 %) > CHT-D (4.17 %) > CHT-E (2.45 %) > CHT-B (2.16 %) (Tables 1-7). Saponins constitute a vast group of glycosides occurring in many plants and are characterized by their surfactant properties. They are grouped as triterpenoid and steroid saponins [55].  The steroidal saponins are essential precursors for steroid drugs, comprising anti-inflammatory agents, androgens, oestrogens and progestins [56]  while triterpene saponins exhibit various pharmacological activities, including anti-inflammatory, molluscicidal, antitussive, expectorant, analgesic and cytotoxic influences and include the ginsenosides, which are responsible for some of the pharmacological activity of ginseng and the active triterpenoid saponins from liquorice [57, 58].

4.5 Tannins

Tannins were detected only in these CHTs as CHT-A (2.54 %) > CHT-G (2.48 %) > CHT-F (2.22 %). The existence of tannins in tea leaves accounts for the bitter and dry sensation felt when tea is tasted. Tannins are higher in black tea than in oolong, green and white teas [59]. Tannins also occur in red wine, coffee, grapes, apple juice, strawberry, raspberry, blackberry, pomegranate, plums, walnuts, olives, chickpeas, lentils, chocolate and cocoa [60].  Foods rich in tannins have been considered to be of low nutritional value since tannins have been reflected as an anti-nutrient, due to their ability to decrease the efficiency in converting the absorbed nutrients to relevant substances [61, 62]. Tea polyphenols and several components of tannin have been suggested as anti-carcinogenic and many tannin molecules have also been shown to reduce the mutagenic activity of several mutagens. These properties have been attributed to their anti-oxidative properties which enable them to defend against oxidative impairment [61].

4.6 Anti-nutrients

The ‘anti-nutrients’, comprise lectins, oxalates, phytates, phytoestrogens and tannins [63]. Phytates and oxalates were detected in the CHTs as CHT-F > CHT-A > CHT-G > CHT-C > CHT-D > CHT-E (Tables 1, 3-7). Oxalate was higher than phytate in CHT-F (16.77 %), CHT-A (12.76 %) and CHT-G (12.71 %) whereas phytate was higher than oxalate in CHT-C (13.51 %), CHT-D (11.53 %) and CHT-E (4.57 %). Anti-nutrients limit the bioavailability of vital nutrients by binding to vital micronutrients which prevents the body from absorbing them or hindering the peak effects of some digestive enzymes, thereby, inhibiting the appropriate breakdown of food [63]. For example, oxalates are known to affect calcium absorption and use by forming calcium oxalate crystals which could lead to kidney stones. They also irritate and cause swelling in the mouth and throat, and are capable of forming tissue crystals leading to indications of arthritis [64]. Some of the health benefits derivable from dietary phytate include anti-cancer, anti-calcification, antioxidant, antihyperglycaemic and hypolipidaemic activities [63, 65]. They can bind to harmful trace elements like lead and cadmium thereby reducing their bioavailability. It has been associated with certain health benefits, including blood glucose – and lipid-lowering effects, anticancer activity, antioxidant properties, and anti-calcification. The ability of phytate to bind toxic trace elements such as cadmium and lead and reduce their bioavailability has been documented [66].

4.7 Steroids

Steroids were detected in the CHTs, but, due to the limitations of the study, the type of steroid identified was not identified. However, steroids were detected in the order CHT-G (13.12 %) > CHT-A (12.95 %) > CHT-C (10.05 %) > CHT-D (8.93 %) > CHT-F (7.83 %) > CHT-E (6.27 %) > CHT-B (6.23 %) (Tables 1-7). Amongst the plant-based steroids, phytosterols are the most abundant [67]. They are known to reduce blood cholesterol by inhibiting intestinal absorption of cholesterol thereby reducing the risk of heart attack and stroke [67]. They have also been demonstrated to slow the in vitro development and progression of various cancers [68].

4.8 Resveratrol

Resveratrol, a non-flavonoid polyphenol was detected in low levels in CHT-D (2.28 %) > CHT-E (1.70 %) > CHT-B (1.57 %). It is a polyphenolic phytoalexin formed by plants like grapes, peanuts and berries and retains anti-inflammatory, antioxidant, antiplatelet, anticancer and anti-diabetic activities [69, 70]. In vitro investigations have also revealed its ability to prevent all phases of carcinogenesis comprising initiation, promotion and progression at lower doses [71]. However, at higher doses, resveratrol acts as a pro-apoptotic compound which signals the death of cancer cells. They are also able to depress cardiac function [71].

5. Conclusion

The GC-FID-guided phytochemical identification and quantification of selected CHTs showed that they contain mostly detectable flavonoids, alkaloids, glycosides, saponins, tannins, steroids, anti-nutrients and other phenols such as resveratrol. This present study corroborates the literature on the abundant phytochemical constituents of chamomile which serves as the basis for the numerous health benefits ascribed to chamomile herbal tea.

References

1. Forni C, Facchiano F, Bartoli M, Pieretti S, Facchiano A, D’Arcangelo D, et al. Beneficial Role of Phytochemicals on Oxidative Stress and Age-Related Diseases. BioMed research international (2019).
2. Craig WJ. Phytochemicals: Guardians of Our Health. Journal of the American Dietetic Association 97 (1997): S199-S204.
3. Ugoeze KC. Phytopharmaceuticals for Treating Sexually Transmitted Diseases. In: Sindhu RK, Singh I, Shirkhedkar AA, Panichayupakaranant P, eds. Herbal Drugs for the Management of Infectious Disease. USA: John Wiley & Sons (2022).
4. Velu G, Palanichamy V, Rajan AP. Phytochemical and Pharmacological Importance of Plant Secondary Metabolites in Modern Medicine. Bioorganic Phase in Natural Food: An Overview: Springer (2018): 135-56.
5. Croft KD. The Chemistry and Biological Effects of Flavonoids and Phenolic Acids A. Annals of the New York Academy of Sciences 854 (1998): 435-42.
6. Catoni C, Schaefer HM, Peters A. Fruit for Health: The Effect of Flavonoids on Humoral Immune Response and Food Selection in a Frugivorous Bird. Functional Ecology (2008): 649-54.
7. Kris-Etherton PM, Harris WS, Appel LJ. Omega-3 Fatty Acids and Cardiovascular Disease: New Recommendations from the American Heart Association. Arteriosclerosis, Thrombosis and Vascular Biology 23 (2003): 151-2.
8. Shukitt-Hale B, Galli RL, Meterko V, Carey A, Bielinski DF, McGhie T, et al. Dietary Supplementation with Fruit Polyphenolics Ameliorates Age-Related Deficits in Behavior and Neuronal Markers of Inflammation and Oxidative Stress. Age 27 (2005): 49-57.
9. Saravanan D, Thirumalai D, Asharani IV. Anti-HIV Flavonoids from Natural Products: A Systematic Review. International Journal of Research in Pharmaceutical Sciences 6 (2015):248-255.
10. Middleton E. Biological Properties of Plant Flavonoids: An Overview. International Journal of Pharmacognosy 34 (1996): 344-8.
11. Harborne JB, Williams CA. Advances in Flavonoid Research since 1992. Phytochemistry 55 (2000): 481-504.
12. Liu RH. Health-Promoting Components of Fruits and Vegetables in the Diet. Advances in nutrition 4 (2013): 384S-92S.
13. Martin LC. Tea: The Drink That Changed the World. USA: Tuttle Publishing (2007).
14. Hernández Figueroa TT, Rodríguez-Rodríguez E, Sánchez-Muniz FJ. El Té Verde¿ Una Buena Elección Para La Prevención De Enfermedades Cardiovasculares? Archivos Latinoamericanos de Nutrición 54 (2004): 380-94.
15. US Food Drug Administration. Generally Recognized as Safe. Silver Spring (2019). https://www.fda.gov/food/food-ingredients-packaging/generally-recognized-safe-gras. Retrieved on 12^th November 2022.
16. Khan N, Mukhtar H. Tea Polyphenols in Promotion of Human Health. Nutrients 11 (2018): 39.
17. Ma C, Chen L. Research Progress on Isolation and Cloning of Functional Genes in Tea Plants. Frontiers of Agriculture in China 1 (2007): 449-55.
18. Khan N, Mukhtar H. Tea, and Health: Studies in Humans. Current Pharmaceutical Design 19 (2013): 6141-7.
19. Weerawatanakorn M, Hung W-L, Pan M-H, Li S, Li D, Wan X, et al. Chemistry and Health Beneficial Effects of Oolong Tea and Theasinensins. Food Science and Human Wellness 4 (2015): 133-46.
20. He R-r, Chen L, Lin B-h, Matsui Y, Yao X-s, Kurihara H. Beneficial Effects of Oolong Tea Consumption on Diet-Induced Overweight and Obese Subjects. Chinese Journal of Integrative Medicine 15 (2009): 34-41.
21. Owuor PO, Obaga SO, Othieno CO. The Effects of Altitude on the Chemical Composition of Black Tea. Journal of the Science of Food and Agriculture 50 (1990): 9-17.
22. Astin JA, Pelletier KR, Marie A, Haskell WL. Complementary and Alternative Medicine Use among Elderly Persons: One-Year Analysis. J Gerontol Med Sci 55 (2000): M4-9.
23. Hansen HV, Christensen KI. The Common Chamomile and the Scentless Mayweed Revisited. Taxon 58 (2009): 261-4.
24. Singh O, Khanam Z, Misra N, Srivastava MK. Chamomile (Matricaria chamomilla): An Overview. Pharmacognosy Reviews 5 (2011): 82.
25. Srivastava JK, Shankar E, Gupta S. Chamomile: A Herbal Medicine of the Past with a Bright Future. Molecular Medicine Reports 3 (2010): 895-901.
26. Mikstas C. All About Herbal Tea. WebMD LLC (2022). Available from https://www.webmd.com/food-recipes/ss/slideshow-herbal-tea. Retrieved on 12^th November 2022.
27. Center GM. Different Types of Tea and Caffeine Content (2022). Available from https://www.garfieldmedicalcenter.com/GMC-Blog/2016/October/Different-Types-of-Tea-and-Caffeine-Content.aspx. Retrieved on 12^th November 2022.
28. Newall CA, Anderson LA, Phillipson JD. Herbal Medicines. A Guide for Health-Care Professionals: The Pharmaceutical Press (1996).
29. Hamon N. Herbal Medicine. The Chamomiles. Can Pharm J 612: (1989).
30. Redaelli C, Formentini L, Santaniello E. Reversed-Phase High-Performance Liquid Chromatography Analysis of Apigenin and Its Glucosides in Flowers of Matricaria chamomilla and Chamomile Extracts. Planta Medica 42 (1981): 288-92.
31. Avallone R, Zanoli P, Puia G, Kleinschnitz M, Schreier P, Baraldi M. Pharmacological Profile of Apigenin, a Flavonoid Isolated from Matricaria chamomilla. Biochemical Pharmacology 59 (2000): 1387-94.
32. Avallone R, Zanoli P, Corsi L, Cannazza G, Baraldi M. Benzodiazepine-like Compounds and Gaba in Flower Heads of Matricaria chamomilla. Phytotherapy Research (United Kingdom) (1996): s177-s179.
33. Salehi B, Venditti A, Sharifi-Rad M, Kregiel D, Sharifi-Rad J, Durazzo A, et al. The Therapeutic Potential of Apigenin. International Journal of Molecular Sciences 20 (2019): 1305.
34. Anderson C, Lis-Balchin M, Kirk-Smith M. Evaluation of Massage with Essential Oils on Childhood Atopic Eczema. Phytotherapy Research: An International Journal Devoted to Pharmacological and Toxicological Evaluation of Natural Product Derivatives 14 (2000): 452-6.
35. Buss AD, Butler MS. Natural Product Chemistry for Drug Discovery: Royal Society of Chemistry (2010).
36. Ugoeze KC, Oluigbo KE, Chinko BC. Phytomedicinal and Nutraceutical Benefits of the GC-FID Quantified Phytocomponents of the Aqueous Extract of Azadirachta indica Journal of Pharmacy and Pharmacology Research 4 (2020): 149-63.
37. Bezerra KdS, Antoniosi Filho NR. Characterization and Quantification by Gas Chromatography of Free Steroids in Unsaponifiable Matter of Vegetable Oils. Journal of the Brazilian Chemical Society 25 (2014): 238-45.
38. Liu W, Yin D, Li N, Hou X, Wang D, Li D, Liu J. Influence of Environmental Factors on the Active Substance Production and Antioxidant Activity in Potentilla Fruticosa L. And Its Quality Assessment. Scientific Reports 6 (2016): 1-18.
39. Del Rio D, Rodriguez-Mateos A, Spencer JP, Tognolini M, Borges G, Crozier A. Dietary (Poly) Phenolics in Human Health: Structures, Bioavailability, and Evidence of Protective Effects against Chronic Diseases. Antioxidants & Redox Signaling 18 (2013): 1818-92.
40. Panche AN, Diwan AD, Chandra SR. Flavonoids: An Overview. Journal of nutritional science 5 (2016).
41. Atanassova M, Bagdassarian V. Rutin Content in Plant Products. Journal of the University of Chemical Technology and Metallurgy 44 (2009): 201-3.
42. Cheol-Ho P, Kim Y, Choi Y, Heo K, Kim S. Rutin Content in Food Products Processed from Groats, Leaves, and Flowers of Buckwheat. Fagopyrum 17 (2000): 63-6.
43. Yang L, Stöckigt J. Trends for Diverse Production Strategies of Plant Medicinal Alkaloids. Natural Product Reports 27 (2010): 1469-79.
44. Alves de Almeida AC, de-Faria FM, Dunder RJ, Manzo LPB, Souza-Brito ARM, Luiz-Ferreira A. Recent Trends in Pharmacological Activity of Alkaloids in Animal Colitis: Potential Use for Inflammatory Bowel Disease. Evidence-Based Complementary and Alternative Medicine 2017 (2017).
45. Jadhav SJ, Sharma RP, Salunkhe DK. Naturally Occurring Toxic Alkaloids in Foods. CRC Critical Reviews in Toxicology 9 (1981): 21-104.
46. Manu KA, Kuttan G. Immunomodulatory Activities of Punarnavine, an Alkaloid from Boerhaavia diffusa. Immunopharmacology and Immunotoxicology 31 (2009): 377-87.
47. Morsy N. Cardiac Glycosides in Medicinal Plants. Aromatic and medicinal plants–back to nature. London: Intechopen (2017): 29-45.
48. Hollman A. Plants and Cardiac Glycosides. British Heart Journal 54 (1985): 258.
49. Shah A, Varma C, Patankar S, Kadam V. Plant Glycosides and Aglycones Displaying Antiproliferative and Antitumour Activities–a Review. Current Bioactive Compounds 9 (2013): 288-305.
50. Kelly RA. Cardiac Glycosides and Congestive Heart Failure. The American Journal of Cardiology 6 5(1990): E10-E6.
51. Ambrosy AP, Butler J, Ahmed A, Vaduganathan M, Van Veldhuisen DJ, Colucci WS, et al. The Use of Digoxin in Patients with Worsening Chronic Heart Failure: Reconsidering an Old Drug to Reduce Hospital Admissions. Journal of the American College of Cardiology 63 (2014): 1823-32.
52. Heinrich M. Plant Resources of South-East Asia No. 12 (1). Medicinal and Poisonous Plants 1-Ls De Padua, N. Bunyapraphatsara and Rhmj Lemmens. Backhuys, Leiden, 1999, Pp. 711, Numerous Botanical and Some Chemical Line Drawings, Bibliography, Indexes (Compounds, Pharmaceutical Terms, Scientific Plant Names, Vernacular Plant Names), Hardcover, Isbn 90 5782 042 0 (350 Dutch Guilders-Ca Us $180). Phytochemistry 53 (2000): 619-20.
53. Kwok J. Cyanide Poisoning and Cassava. Food Safety Focus 19 (2008). Available from https://www.cfs.gov.hk/english/multimedia/multimedia_pub/multimedia_pub_fsf_19_01.html. Retrieved on 12^th November 2022.
54. Bolarinwa IF, Oke MO, Olaniyan SA, Ajala AS. A Review of Cyanogenic Glycosides in Edible Plants. In: Soloneski S, Larramendy ML, editors. Toxicology - New Aspects to This Scientific Conundrum. London: IntechOpen; 2016.
55. El Aziz MMA, Ashour AS, Melad ASG. A Review on Saponins from Medicinal Plants: Chemistry, Isolation, and Determination. J. Nanomed Res 8 (2019): 282-8.
56. Shao B, Guo H, Cui Y, Ye M, Han J, Guo D. Steroidal Saponins from Smilax China and Their Anti-Inflammatory Activities. Phytochemistry 68 (2007): 623-30.
57. Dinda B, Debnath S, Mohanta BC, Harigaya Y. Naturally Occurring Triterpenoid Saponins. Chemistry & Biodiversity 7 (2010): 2327-580.
58. Biswas T, Dwivedi UN. Plant Triterpenoid Saponins: Biosynthesis, in Vitro Production, and Pharmacological Relevance. Protoplasma 256 (2019): 1463-86.
59. Khasnabis J, Rai C, Roy A. Determination of Tannin Content by Titrimetric Method from Different Types of Tea. Journal of Chemical and Pharmaceutical Research 7 (2015): 238-41.
60. Perez-Jimenez J, Neveu V, Vos F, Scalbert A. Systematic Analysis of the Content of 502 Polyphenols in 452 Foods and Beverages: An Application of the Phenol-Explorer Database. Journal of Agricultural and Food Chemistry 58 (2010): 4959-69.
61. Chung K-T, Wong TY, Wei C-I, Huang Y-W, Lin Y. Tannins and Human Health: A Review. Critical Reviews in Food Science and Nutrition 38 (1998): 421-64.
62. Ozcan T, Akpinar-Bayizit A, Yilmaz-Ersan L, Delikanli B. Phenolics in Human Health. International Journal of Chemical Engineering and Applications 5 (2014): 393.
63. Gibson RS, Raboy V, King JC. Implications of Phytate in Plant-Based Foods for Iron and Zinc Bioavailability, Setting Dietary Requirements and Formulating Programs and Policies. Nutrition Reviews 76 (2018): 793-804.
64. Samtiya M, Aluko RE, Dhewa T. Plant Food Anti-Nutritional Factors and Their Reduction Strategies: An Overview. Food Production, Processing and Nutrition 2 (2020): 1-14.
65. Schlemmer U, Frølich W, Prieto RM, Grases F. Phytate in Foods and Significance for Humans: Food Sources, Intake, Processing, Bioavailability, Protective Role and Analysis. Molecular Nutrition & Food Research 53 (2009): S330-S75.
66. Kumar V, Sinha AK, Makkar HP, Becker K. Dietary Roles of Phytate and Phytase in Human Nutrition: A Review. Food Chemistry 120 (2010): 945-59.
67. Gunaherath GMKB, Gunatilaka AAL. Plant Steroids: Occurrence, Biological Significance and Their Analysis. Encyclopedia of Analytical Chemistry: Applications, Theory and Instrumentation (2006): 1-26.
68. Woyengo T, Ramprasath V, Jones P. Anticancer Effects of Phytosterols. European Journal of Clinical Nutrition 63 (2009): 813-20.
69. Koushki M, Amiri-Dashatan N, Ahmadi N, Abbaszadeh HA, Rezaei-Tavirani M. Resveratrol: A Miraculous Natural Compound for Diseases Treatment. Food Science & Nutrition 6 (2018): 2473-90.
70. Hosseini H, Koushki M, Khodabandehloo H, Fathi M, Panahi G, Teimouri M, Majidi Z, Meshkani R. The Effect of Resveratrol Supplementation on C-Reactive Protein (Crp) in Type 2 Diabetic Patients: Results from a Systematic Review and Meta-Analysis of Randomized Controlled Trials. Complementary Therapies in Medicine 49 (2020): 102251.
71. Mukherjee S, Dudley JI, Das DK. Dose-Dependency of Resveratrol in Providing Health Benefits. Dose-Response 8 (2010): 09-015.