A Comprehensive Review on Phytochemical, Pharmacological and Future Prospective of Dietary Medicinal Plant Cinnamomum osmophloeum Kanehira

Background : The plants belonging to the genus Cinnamomum traditionally used as an ethnomedicine in Asia, Europe, and North America. Cinnamomum osmophloeum Kanehira, is an endemic and economic medicinal plant of Taiwan. It is traditionally used as an antibacterial, antifungal, anti-termite, antidiabetic, anti-hyperuricemia, antiinflammatory, and antioxidant agent. Despite these attributes, C. osmophloeum is not enough explored scientifically. This review updated the essential oils and other secondary metabolites, and the pharmacological activities of C. osmophloeum , as well as its other economic benefits. Methods: The information in the review is extracted from the major scientific databases, such as PubMed, BioMed Central, Google scholar, Elsevier, ACS publications, MDPI, Taylor and Francis, Wiley Online Library, Scopus, Springer, and Web of Science, using journals, dissertations, books and/or chapters, and conference proceedings. Results : Various secondary metabolites including essential oil components, flavonoids, lignans, and benzenoids are reported from the extracts of C. osmophloeum . The review established a wide range of pharmacological properties including, antibacterial, antidiabetic, anti-fungal, anti-inflammatory, antioxidant, antitermitic, anti-tyrosinase, anti-xanthine oxidase, anxiolytic, cytotoxic, hepatoprotective, and mosquito larvicidal properties of extracts, as well as essential oils and other secondary metabolites of C. osmophloeum . Conclusions : The present review provides a scientific basis for future studies and necessary information for the development of C. osmophloeum based therapeutic agents.


Introduction
The genus Cinnamomum belongs to the plant family of Lauraceae. It comprises about 250 species, which are distributed in tropical and subtropical Asia, Australia, Pacific islands [1]. The inner bark of the Cinnamomum trees is known as cinnamon [1]. sidewalk trees, and used in traditional medicine, timber, as well as edible fruits. Importantly, cinnamon is commonly used as a spice in food to give aroma taste and flavor, and to act as a preservative [1]. Other folk uses of cinnamon include applying its essential oil as a fragrance in cosmetics, perfumes, and cigarettes [1].
Although people used cinnamon for quite a long time, however, the hepatotoxic compound, coumarin is found in cinnamon in various amounts [2]. Coumarin is a natural flavoring molecule used as an ingredient in foods, alcoholic beverages, tobaccos, toothpastes, and detergents. In this connection, it is interesting to note that the coumarin contents of Cassia cinnamons are generally higher (~40-12180 mg/kg), than that of Ceylon cinnamon (~0−486 mg/kg) [3,4]. However, the use of coumarin as a food flavoring agent is prohibited in the 1950s due to its hepatotoxicity. In this connection, it is necessary to found the alternative source for the safer cinnamons with low coumarin content, which can be beneficial to the global spice market.
Cinnamomum osmophloeum Kanehira is a native tree species in Taiwan, commonly known as pseudo cinnamomum or indigenous cinnamon [5]. Eight of 14 Cinnamomum species in Taiwan are endemic, including C. osmophloeum [5]. It is a small evergreen tree that grows in the mountainous area of Po-Li, Taiwan [6]. The plant C. osmophloeum grows to 12 m in height and about 40 cm in diameter, and normally inhabits in Taiwan's natural hardwood forests at elevations between 400 and 1500 m [6]. The leaves of C. osmophloeum are traditionally used in Taiwanese folk medicines as an antibacterial, antifungal, antitermite, antidiabetic, antihyperuricemia, antiinflammatory, and antioxidant agent [5]. Additionally, C. osmophloeum leaves are used in food, flavoring agent, spices, beverages, medical products, and perfumes. The leaves of C. osmophloeum has nine chemotypes with various secondary metabolite profiles, which are discussed later of this review. It is interesting to note that the leaf essential oil of C. osmophloeum is similar to those of commercial C. cassia bark essential oil [2]. However, the Cassia cinnamon bark samples contain higher level of coumarin, whereas the C. osmophloeum leaf samples comparatively contains the lower levels of coumarin [2]. Taste-wise, C. osmophloeum leaves are milder than ceylon cinnamon, and less heavy on the spicy notes with wafts of vanilla. The plant C. osmophloeum is cultivated in the large areas in Taiwan. A recent review reported that the potential use of C. osmophloeum in the alleviation of oral mucositis [7]. However, the chemical structures of the compounds and their complete pharmacological activities are not fully displayed. Therefore, the present review reported the phytochemical constituents and their potential pharmacological activities, analyses methods, as well as other economic benefits of C. osmophloeum.

Literature Methodology
Relevant information about C. osmophloeum is obtained from ancient books, records, doctoral and master's theses, and scientific search engines including, PubMed, SciFinder, Web of Science, Science Direct, Google Scholar, and so on. The literature search is carried out to gather all relevant information about the traditional uses, phytochemicals and pharmacological activities, and underlying mechanism of action, toxicological and safety considerations of C. osmophloeum. All chemical structures were drawn using ChemDraw 17.0 software.

Specific Identification of C. osmophloeum
Many Cinnamomum plants are morphologically similar [1]. The hepatotoxic adulterant Cinnamomum species, such as C. burmannii, C. loureiroi, and C. cassia, are easily confused with that of non-hepatotoxic C. osmophloeum. Therefore, specific differentiation of C. osmophloeum is critical to avoid toxic issues associated with fraudulent adulteration. In this connection, it is reported that the genetic variation and taxonomic relationship of C. osmophloeum, C. macrostemon and C. insulari [8]. The linalool synthase (LIS) genes are isolated from different provenances of C. osmophloeum [9], and the cinnamaldehyde is increased in 4-coumarate: coenzyme A ligase 1 and 4 (Co4CL1 and Co4CL4), and cinnamoyl-CoA reductase (CoCCR) transgenic plants [10]. Further, the Cinnamomum species, C. burmannii (Nees & T. Nees) Blume, and C. insularimontanum Hayata are morphologically similar with C. osmophloeum [11,12]. However, the leaves of C. burmannii and C. insularimontanum contains lower amount of cinnamaldehyde as compared with the leaves of C. osmophloeum [12]. Therefore, quantitative determination of cinnamaldehyde is an optional method for the identification of C. osmophloeum from C. burmannii and C. insularimontanum [12]. On the other hand, a novel method using leaf images and deep convolutional neural networks (CNN), is reported for the distinction of C. burmannii, C. insularimontanum and C. osmophloeum [12]. To continue, a novel DNA sequence comparisons of internal transcribed spacer 2 (ITS2) method is also reported for the identification of gene resources, genetic diversity, and nucleotide sequence polymorphisms for 73 geographical strains of C. osmophloeum [13]. Recently, Yang et al., developed a polymerase chain reaction based restriction fragment length polymorphism (PCR-RFLP) method for rapid identification C. osmophloeum from adulterant Cinnamomum species by DNA polymorphism analysis [14].

Essential oil components
Essential oils (EOs) are colorless volatile liquids with a characteristic feature of strong odor. Hence, they are widely used in aromatherapy and cosmetics industry [15]. The EOs comprising the aromatic and volatile compounds naturally present in all parts of the plants including seeds, flowers, peel, stem, bark and whole plants [15]. EOs are freely soluble in various solvents such as alcohol, ether, and fixed oils, but insoluble in water [15]. In general, EOs display similar chemical composition and biological activities when obtained from a single plant species grown under similar climate, edaphic conditions and common harvest season. However, the quality and quantity of EOs are vary depends on plant organ, age of trees, chemotypes, growing season, methods of preparation, soil type and climatic conditions [15]. The GC and GC/MS analyses methods are widely used to identify the leaf EO components of C. osmophloeum. The chemical components of leaf EOs are different from various C. osmophloeum clones found in different regions in Taiwan [16][17][18][19][20]. It is interesting to note that the chemical constituents of C. osmophloeum leaf EOs are similar to those of C. cassia bark oil with cinnamaldehyde as the major component [21]. C. cassia bark oil has commercial value, and generally used in food and beverages.
Hu et al., (1985) [22], established an indigenous cinnamon clonal orchard with cuttings of trees from 13 natural populations from central, eastern and southern regions of Taiwan, and analyzed the composition of the EOs of C. osmophloeum leaves [22]. They found that C. osmophloeum leaves from certain provenances contain cinnamaldehyde as the major constituent, whereas linalool is a major compound in some other provenances. Based on the abundances of each individual constituent, it is classified the C. osmophloeum leaf EOs into nine types: cassia type, cinnamaldehyde type, coumarin type, linalool type, eugenol type, camphor type, 4-terpineol type, linalool/terpineol type, and mixed type [22]. The GC/MS analysis of volatile oil obtained from the steam distillation of C. osmophloeum leaves, resulted in the identification of EOs components, such as α-pinene (EO14), camphene (EO15), benzaldehyde (EO1), etc. (Figure 1, Table,  These above previous reports indicates that the leaf EOs of C. osmophloeum contains numerous volatile compounds, including monoterpenes, sesquiterpenes, and their oxygenated derivatives and, alcohols, phenols, aldehydes, ketones, esters, acids, and other miscellaneous compounds ( Figure 1, Table 1). It is also observed that the major components of C. osmophloeum leaf EOs are, transcinnamaldehyde, cinnamyl acetate, linalool and eugenol (Table 1).
On the other hand, EOs from the twigs of C. osmophloeum constituents various components including trans-cinnamaldehyde (EO6) ( Table 1) [41]. The thermal stability results of C. osmophloeum leaf EOs indicated that trans-cinnamaldehyde content in eugenol-free EO is affected by high temperatures, however, the stability of EO improved by adding appropriate amounts of eugenol [33]. The identified chemical structures of the EOs from the C. osmophloeum are presented as in Figure 1, and the components names are listed in Table 1  Cinnamon (C. cassia) is a common spice with sweet, spicy, and special flavor. It has been widely used in bakeries, drinks, desserts, and cuisines. The main constituent of essential oil from cinnamon bark is transcinnamaldehyde (EO06). The leaf EOs of indigenous cinnamon (C. osmophloeum) contains higher amount of EO06 as compared with Cinnamon (C. cassia) [2]. In particular, the C. osmophloeum leaf EOs contains ~80% (w/w) of trans-cinnamaldehyde (EO06), and these values ranged from 769 to 809 g/kg of EOs, which correspond to about 8.9−26.1 g/ kg of sample [2]. Additionally, the cinnamaldehyde content of the cinnamon bark EOs is ~325 g/kg, which is much lower than the cinnamaldehyde contents of the C. osmophloeum leaf EOs (769 to 809 g/kg). Therefore, C. osmophloeum leaves can be considered a good quality and has a potential cinnamon substitute source to replace commercial bark cinnamons [2]. Further, a recent report indicates that the relative content of transcinnamaldehyde in the leaves of C. osmophloeum ct. cinnamaldehyde has the seasonal variation, which is relatively lower (32.2%) in the month of May as compared with the rest of the months (>76.3%) [42]. On the other hand, the leaf EOs of C. osmophloeum contains comparatively lower level of coumarin (0.29−13.99 mg/kg), as compared with the EOs of Cassia cinnamon (26.8−97.4 mg/kg) [2]. Therefore, it is reasonable to suggest that the C. osmophloeum as a safer spice substitute for C. cassia.

Other (Non-essential oil) metabolites of C. osmophloeum
Flavonoids are a class of secondary metabolites that consist of more than 7000 structures with fifteen carbon atoms. This class of compound have a wide-range of bioactive properties, including antioxidant, protective against inflammatory processes, hypertension, arthritis and AIDS, and so on. The flavonoid glycosides compounds, kaempferitrin (F1) and kaempferol-7-O-α-rhamnoside (F10) along with coumarin, fumaric acid are reported from the leaves of C. osmophloeum ( Figure 2, Table 2) [43]. Chemical examination of the 80% methanolic extract from C. osmophloeum leaves, resulted in the isolation of highly sweet constituent, trans-cinnamaldehyde (in 1.03% yield, w/w) [23]. Phytochemical investigation on the n-butanol fraction of methanol extract from C. osmophloeum leaves, resulted in the isolation of four kaempferol glycosides (F1 -F4), including a novel one, kaempferol Table 2  On the other hand, lignans are an important part of the secondary metabolites of Cinnamomum species, which have high content and abundant structural types. The phytochemical investigations of the ethanol extracts of C. osmophloeum heartwood and roots, resulted in the isolation of various lignans including three novel structurally related lignan esters, one secolignan ester (L3) and two cyclolignan (or aryltetralin lignan) esters (L4 and L5) ( Figure 2, Table 2) [48]. Chemical examination of the n-butanol soluble fraction of 70% acetone extract from C. osmophloeum twig extracts, resulted in the isolation and structure identification of proanthocyanidins, cinnamtannin B1 (A1) and parameritannin A1 (A2) (Figure 2, Table 2 [1,3] dioxol-5-yl)cyclopropoxy)-2,6-dimethoxyphenol (B5) is reported from the stems of C. osmophloeum ( Figure 2, Table 2

Pharmacological potential of essential oils (EOs)/components
Most of the chemical components in EOs of C. osmophloeum are low-molecular weight compounds, which can easily diffuse across cell membranes to induce biological reactions [7]. A couple of studies have proposed cinnamaldehyde to be a major functional compound for the antidiabetic activity of cinnamon [1]. The antibacterial activities of the EOs from leaves of two C. osmophloeum clones (A and B) were examined against nine strains of bacteria. The results showed that the MICs (minimum inhibitory concentrations) of the B leaf oil were 500 µg/ml against both Klebsiella pneumoniae and Salmonella sp. and 250 µg/ml against the other 7 strains of bacteria (  (Table 3) [25]. The compound transcinnamaldehyde showed potent inhibitory activity against Jurkat (IC 50 =0.057µM) and U937 (IC 50 =0.076µM) cell viability, without affecting the viability of primary purified T cells and macrophages (Table 3)   Leaf essential oil Mosquito larvicidal activity against larvae of Aedes aegypti. LC 50 for cinnamaldehyde type and cinnamaldehyde/cinnamyl acetate type in 24 h were 36 ppm (LC 90 =79 ppm) and 44 ppm (LC 90 =85ppm), respectively. [19] Leaf essential oil Anti-inflammatory-60 µg/mL, inhibited IL-1β and IL-6 but not for TNF-α in LPS-treated J774A. Leaf essential oil In vivo antioxidant activity against juglone-induced oxidative stress in Caenorhabditis elegans.

Enhanced of antioxidant-genes, SOD-3, GST-4 [29]
Leaf essential oil In vivo antidiabetic activity in STZ-induced rats. 12.5 mg/(kg bw)-reduced fasting blood glucose, fructosamine and, elevated plasma and pancreatic insulin levels. However, 25 and 50 mg/(kg bw) shown to be less effective than that of 12.5 mg/(kg bw). Ameliorated oxidative stress and proinflammatory environment in the pancreas. [31]

Pharmacological activities of C. osmophloeum crude extracts
Studies are reported that C. osmophloeum crude extracts showed various pharmacological activities such as antioxidant, anti-inflammatory, anti-tyrosinase, anti-obesity, and anti-diabetic, and wound-healing effects. Diabetes mellitus (DM) is a chronic disease that affects about 7% of the world's people and it is expected to increase by 5.5% in 2025 [69]. DM type 2 (T2DM) accounts for 85-90% of all diagnosed diabetic patients with high medical and social costs [69]. Cinnamon also has a long history of therapeutic use for various health problems including diabetes [1]. However, it was not until the past decade that the possible antidiabetic role of cinnamon in humans and in experimental animals has been investigated scientifically [1]. The results of the antidiabetic effect of cinnamon are inconsistent [1]. Although several clinical studies and a few meta-analyses have confirmed the usefulness of cinnamon as an antidiabetic agent, the results of other clinical studies and meta-analysis have shown cinnamon to be ineffective in oral glucose tolerance, insulin sensitivity, fasting blood glucose, glycated hemoglobin, lipid profile, or peripheral insulin levels in type 2 diabetes patients [1]. Rao et al., (2007) [70] reported that the chloroform and methanol extracts of C. osmophloeum bark showed antiinflammatory and anti-cancer properties through the reduced inflammatory mediators (NO, TNF-α and IL12) production in activated macrophages, and tumor cells proliferation, respectively [70]. Oral administration of C. osmophloeum leaves hot-water extracts, reduced the total cholesterol (TC), triglyceride (TG) and low-density lipoprotein (LDL-C) levels in hyperlipidemic hamsters [71]. The phenolic content of C. osmophloeum water extracts is 160.9 mg/g, which showed a potential antioxidant activity with an IC 50 values of 10.3 and 16.9 μg/mL, for DPPH and superoxide radical scavenging assays, respectively (Table  4) [46]. The ethanolic extract of C. osmophloeum leaves, dosedependently (10, 25, 50, 100, and 200 g/mL) reduced the cell viability, tyrosinase activity and melanin content in melanoma B16-F10 cells (Table 4) [72]. Additionally, the ethanolic extract also showed in vivo wound-healing activity [72]. To continue, the ethanolic extract from the leaves showed skinwhitening and protective properties through decreased tyrosinase activity and melanin content in IBMX-induced B16-F10 cells [73]. The proanthocyanidin-rich n-butanol soluble fractions of 70% acetone extract from C. osmophloeum twig extracts, showed antihyperglycemic effects through reduced α-glucosidase, α-amylase and protein tyrosine phosphatase 1B [49,74]. Additionally, the twig extracts showed better α-glucosidase and αamylase activities than leaf, 2-cm branch and 5-cm branch extracts [74]. Furthermore, the proanthocyanidin-rich n-butanol soluble fractions of 70% acetone extract also showed antihyperglycemic activity in high-fat diet and streptozotocin-induced hyperglycemic rats [75]. The ethanolic extract from the C. osmophloeum leaves showed liver protective property through induced the ghrelin gene variant 1 but not variant 3, mRNA and ghrelin hormone expression in D-ribose-treated HepG2 cells [76]. Recently, the water extract of C. osmophloeum (COK) leaves are confirmed to be useful to treat hair loss [77]. The in vitro bioassays suggested that COK water extract significantly promoted the proliferation of human hair dermal papilla cells (hDPCs) via up-regulating mRNA levels of some hair growth-related factors covering vascular endothelial growth factor, keratinocyte growth factor (KGF) and transforming growth factor-β2 [77]. Besides, the in vivo assays showed that COK leaf extract promoted the anagen phase in the hair growth cycle in hair removal C57BL/6 mouse model [77]. The hydrosol obtained from the steam distillation of C. osmophloeum leaves, reduced oxidative stress and melanogenesis in B16F10 melanoma cells and protect against DNA damage [78]. A recent in vivo study indicated that 95% ethanolic extract from C. osmophloeum leaves had a therapeutic effect against 5-fluororacil-induced oral mucositis in rats [79]. On the other hand, the water-soluble fractions from ground wood of C. osmophloeum, enhanced the cultured mycelia of Antrodia camphorata and its anti-inflammatory potential through reduced ROS production in human leukocytes [80]. The reported pharmacological activities of C. osmophloeum crude extracts are presented as in Table 4.

Tested Sample / Extract
Reported Activity Ref.

Water-soluble fractions from ground wood
Enhanced the cultured mycelia of Antrodia camphorata and its anti-inflammatory potential through reduced ROS production in human leukocytes Leaves / water extract Promote hair growth in vitro and in vivo C57BL/6 mice. Hair growth-related factors-vascular endothelial growth factor, keratinocyte growth factor (KGF), and transforming growth factor-β2 increased in the cultured human hair dermal papilla cells (hDPCs) [77] Leaves/ hydrosol from Decreased melanin synthesis in B16-F10 melanoma cells, antioxidant, anti-tyrosinase, steam distillation antimelanogenesis, and DNA protective activities. [78]

Pharmacological activities of non-essential oil secondary metabolites
Lignan esters (L3, L4, L5) showed cytotoxicities against human liver carcinoma cells HepG2, Hep3B, and Ca9-22 cancer cells [48] ( Table 5). It is reported that kaempferitrin (F1) showed anti-inflammatory activity through reduced pro-inflammatory mediators such as nitric oxide, TNF-α and IL-12 in activated-macrophages [44] ( Table 5). Diabetes mellitus is characterized by an altered metabolism (of carbohydrates, lipids, and lipoproteins) and chronic hyperglycemia resulting from pancreatic β-cell dysfunction, insulin production deficiency, insulin resistance in key target tissues and impaired glycemic index control [69]. These alterations cause severe complications in the functioning of the cardiovascular system, as well as hypertension and dyslipidemia that are risk factors for stroke and myocardial infarction [69]. The major constituent of C. osmophloeum leaves, kaempferitrin activates the insulin signaling pathway and stimulates secretion of adiponectin in 3T3-L1 adipocytes [82] ( Table 5). The compounds F2 and F3 showed insulin-like anti-diabetic activity in mouse 3T3-L1 adipocytes, through enhanced adiponectin secretion, activation of insulin signaling pathway, GLUT4 translocation activity, phosphorylation of IR and activation of phosphatidylinositol-3 kinase (PI3K) [83] ( Table 5).

NO.
Reported activity a Ref.

Analyses of Secondary Metabolites, and Formulations of C. osmophloeum
A simple and accurate reversed-phase high-performance liquid chromatographic (RP-HPLC) separation method is developed for the determination of bioactive flavonol glycosides, namely kaempferol-3,7-O-α-Ldirhamnoside (kaempferitrin,

Conclusions/Future Prospects
The major compounds of C. osmophloeum are essential oils, and flavonoids. Based on a recent report, Cinnamomum essential oil containing cinnamaldehyde might be useful as a food preservative [88]. The self-life of C. osmophloeum essential oil is comparatively longer than that of C. cassia bark oil due to the existence of both eugenol and cinnamaldehyde in C. osmophloeum. Therefore, C. osmophloeum essential oil might useful as a natural food products preservative. The different clones (cultivars) may provide different amount of essential oil and flavonoids as well as the biological activities. Thus, the cultivars of C. osmophloeum need to take into account for the future medical and pharmaceutical research studies. It is interesting to mention that the C. osmophloeum leaves are rich source for the flavonoid compound, kaempferitrin (kaempferol 3,7-dirhamnoside, 0.2428%, w/w) [44]. Kaempferitrin (F1) has various interesting biological activities such as insulin-mimetic in glucose homeostasis, anti-inflammatory, anti-convulsant, antidepressant, anthelmintic, and osteoporosis [89]. Kaempferitrin also improves the meat quality of broiler chickens [89]. On the other hand, the C. osmophloeum leaves compounds, kaempferol 3-O-β-D-glucopyranosyl-(1→4)-α-L-rhamnopyranosyl-7-O-α-Lrhamnopyranoside (F2) and kaempferol 3-O-β-D-apiofuranosyl-(1→2)-α-L-arabinofuranosyl-7-O-α-Lrhamnopyranoside (F3) had an important role on the insulin signaling pathway through insulinmimetic potential, stimulate the glucose transporter-4 (GLUT4), phosphorylation of IRβ, and activation of PI3-K, which are important for the treatment of diabetes and insulin resistance [89]. Therefore, C. osmophloeum extract enriched these compounds has the potential to be used as a new drug, or as a lead to develop novel therapeutic antidiabetic drugs. Although, various pharmacological activities are reported from the C. osmophloeum leaves, however, little is known about the specific active substances and their pharmacological action mechanisms. Therefore, systematic in vivo studies of can be done to better interpret the traditional usage of C. osmophloeum for diabetes, and to develop as a dietary supplement. Additionally, further in vivo studies are need to investigate the bioavailability, distribution and metabolism of C. osmophloeum.