A Comprehensive Update on the Bioactive Compounds from Seagrasses

Marine angiosperms produce a wide variety of secondary metabolites with unique structural features that have the potential to be developed as effective and potent drugs for various diseases. Recently, research trends in secondary metabolites have led to drug discovery with an emphasis on their pharmacological activity. Among marine angiosperms, seagrasses have been utilized for a variety of remedial purposes, such as treating fevers, mental disorders, wounds, skin diseases, muscle pain, and stomach problems. Hence, it is essential to study their bioactive metabolites, medical properties, and underlying mechanisms when considering their pharmacological activity. However, there is a scarcity of studies on the compilation of existing work on their pharmacological uses, pharmacological pathways, and bioactive compounds. This review aims to compile the pharmacological activities of numerous seagrass species, their secondary metabolites, pharmacological properties, and mechanism of action. In conclusion, this review highlights the potency of seagrasses as a promising source of natural therapeutical products for preventing or inhibiting human diseases.


Introduction
Marine natural products have attracted the attention of scientists worldwide for the last five decades. Marine organisms are an exceptional reservoir of new bioactive compounds that exhibit a greater variety of structural and chemical features than terrestrial natural products [1]. Several chemically unique compounds from marine organisms with different biological activities have been isolated, and some of them are under investigation for development as new pharmaceuticals [2]. In addition, some marine-derived drugs are approved by the European Medicines Agency (EMA) and/or the Food and Drug Administration (FDA) [3]. Although they are mostly anticancer agents, a number of viral infections, chronic pain relievers, and hypertriglyceridemia drugs have also been approved. However, no drugs isolated from seagrasses have been approved by the FDA or EMA.
Marine organisms, including seagrasses, are valuable sources of biologically active compounds for the treatment of human diseases. Seagrasses are one of the true marine flowering plants that belong to the group of angiosperms, which have 72 species worldwide, divided into four families (Zosteraceae, Hydrocharitaceae, Posidoniaceae, and Cymodoceaceae) [4,5]. Seagrasses are eukaryotic organisms found in shallow water areas of temperate, subtropical, and tropical seas, except in polar regions [6]. Seagrass beds are found in shallow coastal areas around the world, with distribution ranges around 50-90 m in depth [6]. Seagrass meadows can store carbon, improve water quality, provide food and habitat, and act as biological indicators [7]. Seagrasses are known to produce a wide variety of secondary metabolites that act as defense mechanisms under stress conditions. These active metabolites, such as polyphenols, terpenoids, and halogenated compounds, are produced by several species of the seagrass reported to have anticancer (antitumor), antifungal, anti-inflammatory, antimicrobial, antiviral, antidiabetic, antimalarial, antioxidant, anti-aging, and cytotoxic properties. These species are also effective in the prevention of human diseases [8]. Seagrasses have been used in folk medicine for a variety of remedial purposes, such as the treatment of fevers, stomach problems, muscle pain, wounds, and skin diseases; they are also used as a remedy for stings of different kinds of rays and as tranquilizers for babies [9]. In addition to their pharmacological activity, seagrasses have been utilized for making baskets, burned to obtain salt, mattress filling material, thatched roofs, fertilizer, paper materials to transport fragile items, and nitrocellulose, among other uses [10].
Based on the benefits discussed above, this review amalgamates a comprehensive compilation of the phytochemical composition and biomedical applications of seagrasses around the world.

Methods
Articles published from 2011 to 2022 were retrieved through several databases, namely PubMed, Springer, Elsevier, MDPI, and Google Scholar, for investigating the bioactive compounds and pharmacological activity of marine angiosperms (seagrasses), as well as the compounds isolated from seagrasses under the clinical trial. The search terms used were "Marine angiosperm" OR "Seagrasses" AND "bioactive compounds" OR "phytochemicals" OR "chemical compounds" OR "anticancer" OR "antioxidant" OR "anti-inflammatory" OR "antimicrobial" OR "antibacterial" OR "antifungal" OR "antiviral" OR "anti-dengue" OR "anti-hyperlipidemia" OR "lipid reducing" OR "antidiabetic" OR "hepatoprotective" OR "anti-aging" OR "mechanism of action" OR "underlying mechanism" OR "clinical trial." The search was restricted to articles published in English and Indonesian languages. From the 138 articles that reported the bioactive compounds, their pharmacological activity, clinical trial, and the underlying mechanism of marine angiosperms were found. A limitation of this study is the bioavailability and pharmacokinetic evaluation of the bioactive compounds, which were not well indicated or mentioned in the original articles reported.
The polyphenol compounds from T. ciliatum act as anticancer agents by mobilizing endogenous copper (and possibly chromatin-bound copper) and the result of prooxidant action [17]. Balls and leaves hydroethanolic extract from P. oceanica L. inhibit metastatic activity by decreasing the expression of MMP-2 and MMP-9 [25]. The aqueous extract and silver nanoparticles from C. serrulata induced the inactivation of replication and, reacting with sulfur-containing proteins, led to the inhibition of enzyme functions, which resulted in the loss of cell viability and cell death [26]. Hydrophilic extracts of P. oceanica L. inhibit HT1080 cell migration by decreasing the expression of MMP2 and MMP9 [27]. The chloroform fraction of the hydroethanolic extract from T. testudinum inhibits the proliferation and migration of A549 and EA.hy926 cells by decreasing the expression and activity of hypoxia-inducible factor 1 (HIF-1) [28]. Moreover, hydroethanolic extracts from T. testudinum suppress the angiogenesis of RKO, SW480, and CT26 by inhibiting bFGF-induced neovascularization, triggering ATF4-P53-NFκB-specific gene expression and autophagy stress pathways, and promoting antitumor immunogenic cell death (IFNγ, PD-1, and ZAP70) [29]. Hydroethanolic extracts from T. testudinum also induced cytotoxicity based on oxidative stress, nuclear damage, and sustained hypercalcemia in HepG2 cells [30]. Furthermore, Thalassiolin B from T. testudinum increased the production of ROS and induces pro-apoptotic effects on HCT15 and HT29 cells [23]. T. testudinum extract up to 1000 µg/mL did not induce significant toxicity effects in normal cells (hepatocyte, lymphocyte, CHO, VERO, 3T3, MDCK, and BHK-21), indicating that it is selective in cancer cells [30]. The anticancer mechanisms of various seagrasses are summarized in Figure 2 [31,32].

Antioxidant
Oxidative stress is caused by an imbalance between the free radicals and antioxidants in the body that can irreversibly damage several cellular structures. Oxidative stress has been recognized as being involved in the pathology of many age-related diseases, such as atherosclerosis, diabetes, neurodegenerative diseases, aging, and cancer [35]. Antioxidants from endogenous and exogenous sources may help to counteract the negative effects of oxidative stress. The most effective and widely used strategy to reduce oxidative stress is exogenous antioxidants supplementation [36]. In recent years, there have been concerns over the safety of synthetic antioxidants. Therefore, antioxidants from natural sources are attracting more attention. Natural products, such as carotenoids, tocopherols, and flavonoids, are well recognized as strong antioxidants with activity in scavenging free radicals and relieving cellular damage caused by oxidation [37]. Another group of naturally-derived chemicals, polysaccharides, has also attracted wide attention because of their promising in vitro and in vivo biological activity [38].
Marine organisms have been considered a promising source of nutrients and bioactive compounds. In recent years, many polysaccharides from marine organisms with antioxidant activity have been isolated and identified, but the characteristics of these polysaccharides have rarely been summarized, and their structure-activity relationships have been scarcely reported [38]. Seagrasses are known to produce secondary metabolites as defense mechanisms under stress conditions, and these compounds are found to be antioxidative ( Figure 3). The methanol extract of H. ovalis at 500 µg/mL had higher reducing power than ascorbic acid [39]. The superoxide dismutase activity of E. acoroides showed that the ethyl acetate extract was the most active, with an IC 50 value of 7 ppm; quercetin and catechin, as reference compounds, had IC 50 values of 5 and 13 ppm, respectively [40]. Old leaf extracts of H. stipulacea induced a 3.9-fold up-regulation of the CYBB gene and the down-regulation of EPHX2 (19-fold), EPX (2-fold), MBL2 (11.6-fold), MPO (6.9-fold), and SPINK1 (10-fold) genes in WI-38 cells treated with 10 mM of H 2 O 2 . It is indicated that the oxidative stress response was not activated when cells were treated with H. stipulacea [41]. Moreover, WI-38 cells pre-treated with old leaf extracts of H. stipulacea before an injury with 10 mM of H 2 O 2 exhibited an up-regulation of genes involved in the antioxidant cell response, such as glutathione peroxidase 5/GPX5 (2.3-fold), keratin 1/KRT1 (2.2-fold), lactoperoxidase/LPO (2.6-fold), metallothionein 3/MT3 (2.0-fold), NADPH oxidase 5/NOX5 (2.8-fold), and thyroid peroxidase/TPO (2.3-fold) [41]. A recent comparative study between Z. marina and Z. notei ethyl acetate extracts displayed the antioxidant activity of both extracts ascribed in Z. marina fraction to rosmarinic acid. On the other hand, the Z. noltei extract had the capacity to chelate copper and iron ions, suggesting its potential application to alleviate Alzheimer's disease (AD) symptoms [42] (Table 3).

Anti-Inflammatory Effects
Inflammation is a complex physiological response to various harmful stimuli characterized by the recruitment and activation of immune cells (innate and adaptive immunity), which rapidly manage the resolution and healing of damaged tissues [62]. An uncontrolled immune response can make inflammation a pathological condition. Inflammation leads to a reduction in the pain threshold, inducing pathological hypersensitivity and resulting in persistent pain [63]. The failure of a rapid resolution can evolve into chronic inflammation, which could determine the onset of inflammatory diseases or the development of cancer [64]. Macrophage cells play a significant role in immune responses and inflammatory processes, covering a wide variety of functions, such as the activation of inflammation and regulation of tissue repair [65]. Lipopolysaccharide (LPS) is one of the most widely used pro-inflammatory stimuli that can activate macrophages and trigger the inflammatory response [66]. The ethanolic extract of P. oceanica decreased the LPS-induced high levels of COX2, thus exhibiting an anti-inflammatory role associated with antioxidant effects [67]. This extract also exhibited a strong ability to inhibit oxidative stress by affecting the production of both ROS and NO radicals, as well as by reducing iNOS and COX-2 levels [67]. In addition, its anti-inflammatory role via inhibiting the NF-κB-signaling pathway through modulation of ERK1/2 and Akt intracellular cascades was evidenced [67].
Palmitoleic acid (36) isolated from S. filiforme and T. testudinum had potent antiinflammatory activity by inhibiting the LPS-induced release of TNF-α, IL-1β, IL-6, MIP-3α, and l-selectin [68]. Compound 21 revealed powerful anti-inflammatory properties through MAPK and NF-κB-signaling pathways [69]. Stearic acid (42) isolated from S. filiforme and T. testudinum mitigated the inflammatory response by inhibiting neutrophil migration, thereby reducing TNF-α and IL-1β [70]. Compound 24 acted as an anti-inflammatory agent by suppressing the expression of pro-inflammatory cytokines [71]. The administration of P. oceanica extract at 10-100 mg/kg in a dose-dependent manner increased the pain threshold; the higher dose was significantly effective between 15 and 45 min after treatment, completely blocking carrageenan-induced hypersensitivity [63]. The underlying mechanism is the reduction in the TNF-α and IL-1β levels that play an important role in inflammation pathways [63]. On the other hand, the methanolic extract of H. ovalis exhibited 50% inhibition of the proliferation of peripheral blood mononuclear cells (PBMCs) at 78.72 µg/mL [39]. Compound 14 isolated from T. ciliatum at 20 mg/kg has been found to possess significant anti-inflammatory activity according to the carrageenan-induced rat paw edema test [19]. The anti-inflammation pathway of the hydroethanolic extract of P. oceanica is shown in Figure 4 [72,73].
Mar. Drugs 2022, 20, x 15 of 41 leads to a reduction in the pain threshold, inducing pathological hypersensitivity and resulting in persistent pain [63]. The failure of a rapid resolution can evolve into chronic inflammation, which could determine the onset of inflammatory diseases or the development of cancer [64]. Macrophage cells play a significant role in immune responses and inflammatory processes, covering a wide variety of functions, such as the activation of inflammation and regulation of tissue repair [65]. Lipopolysaccharide (LPS) is one of the most widely used pro-inflammatory stimuli that can activate macrophages and trigger the inflammatory response [66]. The ethanolic extract of P. oceanica decreased the LPS-induced high levels of COX2, thus exhibiting an anti-inflammatory role associated with antioxidant effects [67]. This extract also exhibited a strong ability to inhibit oxidative stress by affecting the production of both ROS and NO radicals, as well as by reducing iNOS and COX-2 levels [67]. In addition, its anti-inflammatory role via inhibiting the NF-κBsignaling pathway through modulation of ERK1/2 and Akt intracellular cascades was evidenced [67]. Palmitoleic acid (36) isolated from S. filiforme and T. testudinum had potent anti-inflammatory activity by inhibiting the LPS-induced release of TNF-α, IL-1β, IL-6, MIP-3α, and l-selectin [68]. Compound 21 revealed powerful anti-inflammatory properties through MAPK and NF-κB-signaling pathways [69]. Stearic acid (42) isolated from S. filiforme and T. testudinum mitigated the inflammatory response by inhibiting neutrophil migration, thereby reducing TNF-α and IL-1β [70]. Compound 24 acted as an anti-inflammatory agent by suppressing the expression of pro-inflammatory cytokines [71]. The administration of P. oceanica extract at 10-100 mg/kg in a dose-dependent manner increased the pain threshold; the higher dose was significantly effective between 15 and 45 min after treatment, completely blocking carrageenan-induced hypersensitivity [63]. The underlying mechanism is the reduction in the TNF-α and IL-1β levels that play an important role in inflammation pathways [63]. On the other hand, the methanolic extract of H. ovalis exhibited 50% inhibition of the proliferation of peripheral blood mononuclear cells (PBMCs) at 78.72 μg/mL [39]. Compound 14 isolated from T. ciliatum at 20 mg/kg has been found to possess significant anti-inflammatory activity according to the carrageenan-induced rat paw edema test [19]. The anti-inflammation pathway of the hydroethanolic extract of P. oceanica is shown in Figure 4 [72,73].

Antibacterial Activity
Infectious diseases remain a major cause of death due to multidrug resistance. The emergence and spread of antibiotic-resistant pathogens are a great concern to the global health community. Annual deaths due to antimicrobial resistance in the world are projected to reach up to 10 million by the year 2050 [74]. Increasing pathogen resistance rates and the ineffectiveness of antibiotics have spurred research on other options. The effective treatment of a disease entails the development of new pharmaceuticals or potential sources of novel drugs. Commonly used medicinal plants could be an excellent source of drugs to resolve this problem [75]. According to the World Health Organization (WHO), medicinal plants could be the best source for obtaining a variety of drugs, as they produce several bioactive compounds with known therapeutic properties. Many plant extracts exhibit a good antibacterial activity towards different tested bacterial isolates, as indicated by their MIC values [76]. Antimicrobial agents are essential to reducing the global burden of infectious diseases. They act through several mechanisms, including damaging bacterial wall permeability, microsomes, and lysosomes; acting as DNA-intercalating agents; and inhibiting the reverse transcriptase and topoisomerase enzymes [77]. The biological activity of alkaloid compounds caused the presence of nitrogenous groups that react with amino acids and change their arrangement [77]. This process destroys the genetic balance in DNA so that bacterial DNA is damaged. DNA damage in the bacterial cell nucleus prevents the bacteria's metabolism, leading to cell lysis [77].
Seagrasses produce antimicrobial compounds that may reduce or control microbial growth, and many reports have described their antibacterial activity [78] (Figure 5).

Antibacterial Activity
Infectious diseases remain a major cause of death due to multidrug resistance. The emergence and spread of antibiotic-resistant pathogens are a great concern to the global health community. Annual deaths due to antimicrobial resistance in the world are projected to reach up to 10 million by the year 2050 [74]. Increasing pathogen resistance rates and the ineffectiveness of antibiotics have spurred research on other options. The effective treatment of a disease entails the development of new pharmaceuticals or potential sources of novel drugs. Commonly used medicinal plants could be an excellent source of drugs to resolve this problem [75]. According to the World Health Organization (WHO), medicinal plants could be the best source for obtaining a variety of drugs, as they produce several bioactive compounds with known therapeutic properties. Many plant extracts exhibit a good antibacterial activity towards different tested bacterial isolates, as indicated by their MIC values [76]. Antimicrobial agents are essential to reducing the global burden of infectious diseases. They act through several mechanisms, including damaging bacterial wall permeability, microsomes, and lysosomes; acting as DNA-intercalating agents; and inhibiting the reverse transcriptase and topoisomerase enzymes [77]. The biological activity of alkaloid compounds caused the presence of nitrogenous groups that react with amino acids and change their arrangement [77]. This process destroys the genetic balance in DNA so that bacterial DNA is damaged. DNA damage in the bacterial cell nucleus prevents the bacteria's metabolism, leading to cell lysis [77].

Antifungal Treatments
Fungal infections are caused by eukaryotic organisms, and it is more difficult to ascertain their presence and apply the appropriate therapeutic treatment compared to bacterial infections [98]. Over the last decades, control of pathogenic fungi has become a critical challenge due to an increase in the incidence of fungal infections and the emergence of antifungal-resistant strains. The onset and severity of the fungal infections depend on the inoculum charge, the host's immunological state, and resistance [99]. Patients who receive immunosuppressive agents such as cancer therapy and immunocompromised patients can be vulnerable to fungal infections [100]. Fungal diseases kill more than 1.5 million and affect over a billion people in the world. Since 2013, the Leading International Fungal Education (LIFE) portal estimated the burden of serious fungal infections for over 5.7 billion people (>80% of the world's population) [101].
Fungi cell walls are considered the prime target for selectively toxic antifungal agents because of their chitin structure, which is absent in human cells [102]. Fungal infection treatments are very limited when compared to bacterial infections. The rise in fungal infection incidence has exacerbated the urgency for new antifungal agents, as many available drugs have several side effects, are ineffective against new or re-emerging fungal strains, and lead to the rapid development of resistance [103]. Ideally, new antifungals should combine major aspects such as sustainability, high efficacy, limited toxicity, and low cost of production. Previous studies showed that seagrasses produced secondary metabolites with a defensive role against marine pathogens. Isoscutellarein (28) and its glycosylated derivatives (29,30) isolated from T. hemprichii showed antifungal activity against A. niger and C. albicans with MIC values between 5 and 8 µg/mL [80]. Flavone glycosides isolated from T. testudinum were reported to inhibit the growth of the thraustochytrid (zoosporic fungus) Schizochytrium aggregatum [104]. The antifungal activities of different seagrasses are summarized in Table 5. A. fumigates Zone of inhibition: 6 mm antifungal activity index: 50% [92] Fusarium Zone of inhibition: 8 mm antifungal activity index: 8% [92] T. hemprichii Hexane/ethanol (3:1) F. acuminatum Zone of inhibition: 2.5 mm [106] A. niger Zone of inhibition: 1.7 mm [106] P. expansum Zone of inhibition: 2.1 mm [106] A. terrus Zone of inhibition: 3.2 mm [106] A. fumigatus Zone of inhibition: 1.5 mm [106] 3.6. Antiviral Activity Infectious viral diseases remain a global problem. Viruses have been resistant to therapy or prophylaxis longer than any form of life because they completely depend on the cells they infect for their multiplication and survival. Currently, there are only a few drugs available to cure viral diseases, including acyclovir, the known antiherpetic drug modeled on a natural product parent. A number of life-threatening viruses, including human immunodeficiency virus (HIV), adenovirus (ADV), hepatitis virus (HAV, HBV, and HCV), herpes simplex virus (HSV), and influenza virus, have affected human health for a long time [107]. Many research efforts have been devoted to the discovery of new antiviral natural products to combat viruses that have devastating effects on humans, animals, insects, crop plants, fungi, and bacteria. Many recent studies have revealed the antiviral activity of various seagrasses; these are summarized in Table 6.

Anti-Dengue Activity
Dengue fever is a global arboviral infection caused by four antigenically distinct dengue virus serotypes: DENV-1, DENV-2, DENV-3, and DENV-4 [114]. They are classified into RNA viruses that belong to the Flavivirus genus/Flaviviridae family [114]. The infection is endemic in more than 100 countries, particularly in Southeast Asia, the western Pacific region, and the Americas [115]. It is considered one of the most prevalent and widely spread diseases transmitted by female mosquito vectors. The primary vectors for the spread of this infection are A. aegypti and A. albopictus, which are mostly found in tropical and subtropical regions [115].
Dengue infection causes several clinical manifestations such as fever, headache, myalgia, rash, leukopenia, thrombocytopenia, and increased liver function [114]. Severe infections cause severe thrombocytopenia, hemorrhaging, and plasma leakage [114]. Until now, there are no protective vaccines against this infection, and their progression is inhibited by abolishing the mosquito vectors. Most of the current mosquito abatement programs are based on the use of larval insecticides [116]. Currently, active endeavors have been directed to the natural extracts of the botanical origins as potent compounds for mosquito larvae. There is no specific antiviral therapy for dengue infection; in fact, dengue fever is usually maintained through adequate hydration and fluid replacement therapy. [114]. Seagrasses produce secondary metabolites that have insect growth inhibitory activity, which can kill mosquito larvae. The underlying mechanism may involve the inhibition of A. aegypti larvae alterations in the spiracular valves of the siphon and anal papillae [117]. Studies on larvicidal activities from several seagrass extracts are shown in Table 7.

Lipid-Reducing Activity
Obesity is a disease associated with poor mental health outcomes and reduced quality of life, and it affects around 600 million people in the world [118]. Obesity is a major risk factor for cardiovascular diseases, diabetes, musculoskeletal disorders, and forms of cancer [118]. Obesity is caused by several factors, such as physical inactivity, a poor diet, and genetic susceptibility, which leads to the accumulation of fat in various body regions [118]. Large quantities of fatty acids from the diet must be transported as triglycerides to protect the body against their toxicity. Elevations in plasma triglyceride are the result of overproduction and impaired clearance of very low-density lipoproteins (VLDL) and chylomicrons, as well as the reduced expression of high-density lipoproteins (HDL) [119]. Elevated levels of VLDL and triglycerides, together with the reduction in HDL levels, lead to hyperlipidemia.
HDL cholesterol levels were significantly reduced in untreated diabetic mice and enhanced significantly in T. hemprichii extract-treated animals [120]. T. hemprichii ethanolic extract decreased both LDL and VLDL cholesterol levels in alloxan-induced diabetic mice [120]. Furthermore, ethyl acetate and methanolic extracts of H. stipulacea were tested using the zebrafish Nile red fat metabolism assay and showed IC 50 values of 2.2 µg/mL and 1.2 µg/mL, respectively, after 48 h [21]. Its mechanism of action is through the inhibition of the acetyl-CoA carboxylase and PPARα agonists [21].

Antidiabetic Activity
Diabetes mellitus (DM) is a cluster of syndromes characterized by hyperglycemia; the altered metabolism of lipids, carbohydrates, and proteins; and an increased risk of complications from vascular diseases [121]. DM is a global health problem, and its incidence is increasing at an alarming rate throughout the world. Decreased physical activity, increasing obesity, stress, and changes in food consumption have been cited as reasons for the increasing diabetic prevalence in the past two decades [122]. The treatment of type 2 DM with oral hypoglycemic agents such as sulfonylurea and biguanides is associated with numerous side effects [121]. The major advantages of herbal medicine seem to be their good potential, low incidence of serious side effects, and low cost. Seagrasses contain many flavonoids and sterols/triterpenoids as their main constituents, which are known bioactive compounds for antidiabetic potential [123]. Flavonoids are also known to regenerate damaged β-cells in diabetic mice [123].
Compounds 52 and 53 isolated from S. filiforme and T. testudinum have potential for type 2 DM treatment by increasing GLUT4 translocation and expression [124,125]. H. stipulacea extracts at doses of 100 and 200 mg/kg/day demonstrated 9-and 13-fold increases in serum NO, respectively, compared to diabetic controls. Its mechanism of action was predicted as a result of the improvement of glucose uptake by the tissues through the restoration of liver GLUT-2 [126]. Moreover, H. stipulacea extracts ameliorated oxidative stress status generated by the free radicals and dyslipidemia under diabetic conditions [126]. Weight loss is one of the clinical features of DM due to adipocytes and muscle tissue degeneration, which make up for the energy lost from the body as the result of frequent urination and the conversion of glycogen to glucose. Alloxan-mediated bodyweight reduction was significantly reversed by the administration of T. hemprichii ethanolic extract for 15 days of treatment. The intraperitoneal administration of the extract resulted in a notable increase in body weight [120].
The methanolic extract of H. beccari exhibited a 50% inhibition of α-amylase and α-glucosidase at 270 µg/mL and 100 µg/mL, respectively [121]. It also regulated the glucose movement out of the cells and took up glucose by facilitating diffusion into the bloodstream, thus controlling post-postprandial glucose levels [121]. The same extract from H. uninervis reduced serum glucose levels in Streptozotocin-induced diabetic rats. The administration of 150 mg/kg H. uninervis extract decreased glucose levels by 24.8% after 6 h and exhibited a 52.5% reduction in glucose levels in the serum absorbed on the 18th day of administration at a dose of 150 mg/kg. Moreover, the administration of 250 mg/kg H. uninervis extract decreased glucose levels by 29.9% after 6 h and by 61.9% on the 18th day [123]. P. oceanica (L.) Delile hydroalcoholic leaves extracts exhibited strong in vitro activity against human serum albumin glycation, validating the recognized traditional antidiabetic role of these extracts. No advanced glycation end products were formed by incubating human serum albumin with glucose in the presence of 0.2 mg of dry extract for 72 h [127].

Hepatoprotective
The liver is a vital organ for survival, and it contributes to almost every metabolic function of the body, regulating homeostasis; it is also a frequent target for many toxicants [128]. In addition, the liver plays an important role in the storage of vitamins, iron, and copper, as well as in the detoxification of a large number of endogenous and exogenous substances [128]. Liver damage can include fatty liver, necrosis, cholestasis, hepatitis, and liver cirrhosis. Damage to the liver can be overcome by preventive (hepatoprotective) and curative (antihepatotoxic) efforts [129]. Conventional drug therapy for various liver damage diseases has limited efficacy and potentially adverse effects [130]. Treatment using extracts derived from natural resources is considered the best method to maintain liver function in the long term without significantly inducing toxic effects [130]. Liver cells containing various enzymes, such as SGOT and SGPT, are important for the diagnosis of liver damage because the enzyme is passed into the blood vessels [129]. Elevated enzyme activity may indicate the presence of liver disease [129].
The ethanolic extract of T. hemprichii exhibited hepatoprotective activity by lowering the levels of SGPT and SGOT in alloxan-induced diabetic mice [120]. Moreover, rats treated with higher doses of the H. uninervis methanolic extract (150 and 250 mg/kg) showed significant improvements in hepatic and renal function [123]. The administration of 280 mg/kg ethanolic extract of C. rotundata rhizome can significantly decrease SGPT and SGOT levels in paracetamol-induced rats [131]. The hydro-methanolic extract of T. ciliatum improved histopathological changes in the liver by inducing antioxidant defense enzymes superoxide dismutase (SOD); elevating GSH (non-enzymatic antioxidant glutathione); and reducing lipid peroxidation, nitric oxide (NO), alanine aminotransferase (ALT), and aspartate aminotransferase (AST) levels in thioacetamide (TAA)-induced liver failure [129]. The underlying mechanism is probably the stimulation of the Nrf2/ARE pathway, which results in the induction of antioxidant enzymes and the modulation of intracellular GSH-P in response to stress [129].

Anti-Aging Effects
Skin aging is a biochemical process resulting from intrinsic and extrinsic factors such as age, hormones, lifestyle, and exposure to UV [132]. The aging process takes effect in the epidermal and dermal layers, which are predominantly related to extracellular matrix (ECM) degradation. ECM consists of several enzymes, including matrix metalloproteinases (MMPs) and collagenase [132]. Collagen is the main constituent of the dermal matrix and is produced by fibroblasts. It is essential for skin tone and turgor, and it undergoes physiological turnover [133]. During skin aging, collagen degradation tends to overwhelm renewal, resulting in the formation of fine lines, wrinkles, and other alterations [133]. Hence, the maintenance of fibroblast function is a prerequisite for reducing skin aging. P. oceanica L. Delile ethanolic extract showed a significant increase in collagen production in fibroblasts exposed to 5 and 10 µg/mL and an increase in lipolysis in the concentration range of 10-200 µg/mL [133].
One of the manifestations of aging is hyperpigmentation, which occurs when the skin produces more melanin [132]. Tyrosinase is a rate-limiting enzyme for melanogenesis that converts tyrosine to melanin. Tyrosinase inhibitors play an important role as skinlightening agents [132]. These inhibitors specifically interact with melanogenic cells and do not lead to side effects compared with other melanogenesis inhibitors [134]. The ethanolic extract of P. oceanica L. Delile also induced 20% tyrosinase inhibition at 5 µg/mL and 45% inhibition at 1000 µg/mL [133].

Bioactive Compounds from Seagrass under the Clinical Trial
Seagrasses share most features of their primary and secondary metabolism with terrestrial plants, since they are derived from land plants, which have secondarily recolonized marine habitats. In this review, we report some bioactive compounds isolated from seagrasses that have been tested in clinical trials, such as rutin (2), ferulic acid (19), quercetin (20), gallic acid (23), azelaic acid (26), lauric acid (33), and rosmarinic acid (54). However, to the best of our knowledge, there are no drugs commercially available that are isolated from seagrasses.
A clinical trial of compound 2 for skin aging has been conducted by doing a doubleblind clinical study in 40 subjects aged 30-50 years and divided into control and experimental groups. Compound 2 increased the mRNA expression of collagen, type I, and alpha 1 (COL1A1) and decreased the mRNA expression of matrix metallopeptidase 1 (MMP1) in HDFs. It was verified that ROS scavenging activity was stimulated by rutin in a dose-dependent manner. In addition, compound 2 exerted protective effects under oxidative stress conditions and increased skin elasticity while decreasing the length, area, and number of wrinkles [134].
Another clinical trial of compound 2 was a controlled study conducted on 53 type 2 diabetes patients randomized into three groups: 20 participants received rutin with vitamin C (group A), 20 received vitamin C (group B), and 13 received antidiabetic treatment only (group C). After eight weeks, significant reductions were observed in the % change of fasting blood glucose (FBG) of groups A and B versus group C. Vitamin C supplementation alone or with compound 2 significantly reduced the % change of FBG compared to controls but had no effect on HbA1c, FBG, TC, fasting insulin, and HOMA-IR or oxidative stress in T2DM patients [135].
Ferulic acid's (19) ability to treat hyperlipidemia has been tested in a clinical trial. The study design was a randomized, double-blind, and placebo-controlled trial. Subjects with hyperlipidemia were randomly divided into two groups. The treatment group (n = 24) was given compound 19 (1000 mg daily), and the control group (n = 24) was provided with a placebo for six weeks. Compound 19 supplementation demonstrated a statistically significant decreases in total cholesterol (8.1%; p = 0.001), LDL-C (9.3%; p < 0.001), and triglyceride (12.1%; p = 0.049), as well as increased HDL-C (4.3%; p = 0.045) compared with the placebo. Compound 19 also significantly decreased the oxidative stress biomarker and the inflammatory markers [136].
Quercetin (20) has strong antioxidant, anti-inflammatory, immunomodulatory, and antiviral properties. It is also characterized by a very high safety profile and is exerted in animals and in humans. Like most other polyphenols, compound 20 has a very low rate of oral absorption, and its clinical use has been considered of modest utility. Compound 20 in a delivery-food grade system with sunflower phospholipids (Quercetin Phytosome ® , QP) increases its oral absorption up to 20-fold. In the reported clinical trial (a randomized, controlled, and open-label study), a daily dose of 1000 mg of QP was investigated for 30 days in 152 COVID-19 outpatients to disclose its adjuvant effect in treating the early symptoms of the disease and preventing severe outcomes. The results revealed reductions in the frequency and length of hospitalization, the need for non-invasive oxygen therapy, the progression to intensive care units, and deaths. The results also confirmed the very high safety profile of compound 20 and suggested possible anti-fatigue and pro-appetite properties. QP is a safe agent and, when used in combination with standard care during the early stages of viral infection, could aid in improving the early symptoms and prevent the severity of COVID-19. Further research is needed to confirm these results [137].
The combination of 4% niacinamide + 1% gallic acid (23) + 1% lauric acid (33) can be used as an alternative topical treatment for acne vulgaris, which is a chronic inflammatory skin disease. In addition, this combination could be used to prevent resistance to topical antibiotics and side effects that might be caused by other skin disease treatments [138].
Azelaic acid (26) is known as an antioxidant agent. However, compound 26 was reported in a clinical trial for its anti-inflammatory activity. The results suggested that this new non-irritating product represents a valid therapeutic option for mild/moderate inflammatory rosacea. Furthermore, the evaluation of erythema changes was clearly defined by the instrumental evaluation of erythema degree by erythema-directed digital photography (EDDP) [139].
Previously, rosmarinic acid (54) was reported as an antioxidative agent. However, the capacity of compound 54 to treat AD has been investigated in clinical trials. A randomized placebo-controlled double-blind 24-week trial using Melissa officinalis extract richly containing compound 54 was carried out on patients with mild dementia due to AD to examine the safety and tolerability of this compound (500 mg daily), its clinical effects, and disease-related biomarker changes. There were no significant differences in cognitive measures; however, the mean Neuropsychiatric Inventory Questionnaire (NPI-Q) score improved by 0.5 points in the M. officinalis group and worsened by 0.7 points in the placebo group between the baseline and 24-week visit, indicating a significant difference. No significant differences were apparent in disease-related biomarkers between the groups. M. officinalis extract containing 500 mg of compound 54, taken daily, was safe and welltolerated by patients with mild dementia due to AD. The results suggest that compound 54 helps prevent the worsening of AD-related neuropsychiatric symptoms [140].
In this context, the indication of the percentage of the bioactive compounds' abundance in the seagrasses identifies them as an alternative and valuable source of those compounds. However, to the best of our knowledge, there are no drugs commercially available that were isolated specifically from seagrasses. Moreover, a limitation of this study is the scarcity of reports about the bioavailability and pharmacokinetics of the bioactive compounds mentioned.

Conclusions
The objective of this review was to compile the most recent promising bioactivities of seagrass secondary metabolites and extracts on human health. The compounds described were organized on the basis of their bioactivities and, consequently, their chemical structure. The addressed biological compounds fall into four major classes: steroids, fatty acids, terpenes, and (mainly) polyphenols, including flavonoids, catechins, chalcones, phenyl-propanoids, and phenylethanoids. Our efforts were intended to create a comprehensive collection of more than 50 natural products isolated from seagrasses that have been reported in the last decade, thus shedding light on the promising bioactive potential such as anticancer, antioxidant, anti-inflammatory, antimicrobial, antifungal, antiviral, anti-dengue, anti-dyslipidemia, antidiabetic, hepatoprotective, and anti-aging properties. A deep investigation of these effects and the discovery and characterization of metabolites derived from seagrasses could enable the preparation of innovative drugs, food supplements, and nutraceuticals for the management of several diseases and metabolic syndromes. Despite the huge interest in their ecological role, documented by around 30% of publisher papers in the last decade, the scientific community has not sufficiently addressed interesting aspects of the bioactive properties of seagrasses' secondary metabolites. This overview provides the scientific bases for re-examining these metabolites with the latest available analytical techniques and the phytochemical investigation of the bioactive seagrass extracts, particularly with more applications for the isolation and structure elucidation. The advent of new analytical techniques, such as high-performance liquid chromatography coupled with tandem high-resolution mass spectrometry (UHPLC-HRMS/MS), can speed up the dereplication in comparative studies for the occurrence or absence of known compounds and improve efficiency in the discovery of new bioactive substances.
Furthermore, after the preliminary screening to identify promising natural products derived from seagrasses, compounds should be evaluated for their safety and efficacy in animal models before being tested in clinical trials and used to develop drugs. A limitation of this study is the scarcity of reports about the bioavailability and pharmacokinetics of the addressed bioactive compounds. In addition, to the best of our knowledge, there are no commercially available bioactive compounds isolated from seagrass. Therefore, future studies should include the physical and chemical properties, bioavailability, and pharmacokinetic aspects of seagrasses' secondary metabolites. Future studies should also determine the appropriate pharmaceutical form or drug delivery system to explore these marine angiosperms' potential pharmacological applications. It is clear that natural products still have much potential value in the field of marine angiosperms.