Cataract Preventive Role of Isolated Phytoconstituents: Findings from a Decade of Research

Cataract is an eye disease with clouding of the eye lens leading to disrupted vision, which often develops slowly and causes blurriness of the eyesight. Although the restoration of the vision in people with cataract is conducted through surgery, the costs and risks remain an issue. Botanical drugs have been evaluated for their potential efficacies in reducing cataract formation decades ago and major active phytoconstituents were isolated from the plant extracts. The aim of this review is to find effective phytoconstituents in cataract treatments in vitro, ex vivo, and in vivo. A literature search was synthesized from the databases of Pubmed, Science Direct, Google Scholar, Web of Science, and Scopus using different combinations of keywords. Selection of all manuscripts were based on inclusion and exclusion criteria together with analysis of publication year, plant species, isolated phytoconstituents, and evaluated cataract activities. Scientists have focused their attention not only for anti-cataract activity in vitro, but also in ex vivo and in vivo from the review of active phytoconstituents in medicinal plants. In our present review, we identified 58 active phytoconstituents with strong anti-cataract effects at in vitro and ex vivo with lack of in vivo studies. Considering the benefits of anti-cataract activities require critical evaluation, more in vivo and clinical trials need to be conducted to increase our understanding on the possible mechanisms of action and the therapeutic effects.


Introduction
The ocular lens is located at the anterior segment of the eye that, together with the cornea, provides the refractive power of the eye. The mature lens is composed of a core of primary lens fiber cells, layers of secondary lens fiber cells, and one layer of anterior lens epithelial cells, which covers the anterior surface of the lens [1]. The major function of the lens is to maintain transparency so that the light can be properly focused on the retina. Unfortunately, the delicate balance required for lens transparency can be easily disturbed by oxidative stress, aging, and UV radiation, and cataracts develop as a result [1].
Cataracts are the most common cause of vision loss in people over the age of 40 and are the leading cause of blindness in the world [2]. Cataracts are defined as lens opacification that prevents a sharply defined image from reaching the retina. As a result, cataract patients have clouded, blurred, or dim visions, which significantly affect their daily life. According to a report from the World Health Organization, nearly 40 million people are blind worldwide, almost half of them are due to cataract [3]. Although cataract-related vision loss can be corrected by replacement with synthetic lenses, cataract surgery is a costly procedure and may develop complications like infectious endophthalmitis, posterior capsule rupture during surgery, post-operative macular edema, and posterior capsule opacity (also called posterior capsule opacification). In developing countries, many cataract patients cannot have their vision restored due to financial concerns or lack of medical resources. Therefore, identifying a safe compound that can reduce the incidence or delay the onset of cataract is an important step in finding new treatments for cataract.
There have been many compounds evaluated for their potential efficacies in reducing cataract formation. In this article, we focus on active ingredients derived from plants. Phytoconstituents are a trove of often structurally complicated compounds with interesting biological functions. They themselves or their derivatives have always been important sources of pharmacologically active agents. To provide a comprehensive review of potentially useful anti-cataract phytoconstituents, we searched, selected, and extracted the appropriate information from published literature according to the following procedures. We feel that, by listing the comprehensive collection of phytocontituents in one place, this manuscript serves as an overview and perhaps an inspiration to prompt additional studies in this important research area. Collaborative efforts between phytochemists and cataract researchers are promisingly fruitful.

Literature Search
Literature search of articles published from January 2008 to December 2017 was performed. We searched the databases of Pubmed, Science direct, Google Scholar, Web of Science and Scopus using different combinations of keywords: lens epithelial cells, sodium selenite-, ultraviolet radiation-, steroid induced, oxygen-, H 2 O 2 -induced opacity/cataract, congenital/juvenile cataract, transgenic/knockout mice with cataract, diabetic cataract, spontaneous cataract, isolated phytoconstituents, medicinal plants.

Data Extraction
All the selected manuscripts were analyzed for year of publication, plant species, family, part of plant, solvent extraction, isolation method, isolated phytoconstituents, anti-cataract activities (in vitro, ex vivo or in vivo), route of administration (in vivo), dose or concentration for IC50, treatment duration (in vivo) and isolated phytoconstituent(s) with the strongest activity(s), as well as their structural formula. The extracted data are presented in Tables 1 and 2 throughout this article.

Experimental Cataract Models
There are a large number of in vitro and in vivo models that mimic certain aspects of the pathophysiological features of human cataracts. They have been used to demonstrate the potential therapeutic effects of phytochemicals. In this section, we describe the most commonly used models in order to aid the understanding and appraisal of results. During our literature search, most of the phytochemicals were tested in in vitro or ex vivo models only. Only a dozen or so were assessed in in vivo cataract models. Nevertheless, for completion's sake, we list both in vitro and in vivo models. In vitro models discussed are hydrogen peroxide (H2O2)-, xylose-, galactose-induced lens opacity, aldose reductase (AR) activity assay, and advanced glycation end products (AGE) formation. In vivo models include sodium selenite-, ultraviolet (UV) radiation-, and steroid-induced cataracts. These models have been widely used to study the mechanisms of cataract and serve as the screening platform of anti-cataract therapies with the long-term goal to treat cataract in humans.

H2O2-Induced Cataract
It is widely accepted that oxidative stress is the major factor for the development of cataracts. Hydrogen peroxide (H2O2) is the major reactive oxygen species (ROS). H2O2 is mainly generated in vivo by the detoxification of superoxide (O2 − ) radical by superoxide dismutase (SOD) through the dismutation reaction [4,5]. Alternatively, H2O2 can be produced by a number of oxidase enzymes including monoamine oxidases and peroxisomal pathway for β-oxidation of fatty acids. In the lens,

Data Extraction
All the selected manuscripts were analyzed for year of publication, plant species, family, part of plant, solvent extraction, isolation method, isolated phytoconstituents, anti-cataract activities (in vitro, ex vivo or in vivo), route of administration (in vivo), dose or concentration for IC 50 , treatment duration (in vivo) and isolated phytoconstituent(s) with the strongest activity(s), as well as their structural formula. The extracted data are presented in Tables 1 and 2 throughout this article.

Experimental Cataract Models
There are a large number of in vitro and in vivo models that mimic certain aspects of the pathophysiological features of human cataracts. They have been used to demonstrate the potential therapeutic effects of phytochemicals. In this section, we describe the most commonly used models in order to aid the understanding and appraisal of results. During our literature search, most of the phytochemicals were tested in in vitro or ex vivo models only. Only a dozen or so were assessed in in vivo cataract models. Nevertheless, for completion's sake, we list both in vitro and in vivo models. In vitro models discussed are hydrogen peroxide (H 2 O 2 )-, xylose-, galactose-induced lens opacity, aldose reductase (AR) activity assay, and advanced glycation end products (AGE) formation. In vivo models include sodium selenite-, ultraviolet (UV) radiation-, and steroid-induced cataracts. These models have been widely used to study the mechanisms of cataract and serve as the screening platform of anti-cataract therapies with the long-term goal to treat cataract in humans.

Oxidative Stress Model H 2 O 2 -Induced Cataract
It is widely accepted that oxidative stress is the major factor for the development of cataracts. Hydrogen peroxide (H 2 O 2 ) is the major reactive oxygen species (ROS). H 2 O 2 is mainly generated in vivo by the detoxification of superoxide (O 2 − ) radical by superoxide dismutase (SOD) through the Compared with glucose, galactose has higher affinity with AR and its reduction product galactitol is more difficult to be metabolized by sorbitol dehydrogenase than sorbitol. Therefore, high galactose is more likely to induce sugar cataract than high glucose itself [13]. There are several methods available to establish galactosemic cataract. For example, rat galactosemic cataract can be induced by 30% or 50% galactose diet. Glactose-induced lens opacity can also be achieved by daily intraperitoneal injection of 30-50% galactose solution or daily retrobulbar injection of 20% galactose solution. Another cost efficient way to induce galactosemic cataract is to feed rat 10% galactose solution for 18 days. For in vitro lens culture, 30 mM galactose is added in the culture medium for 72 h incubation [13].

Formation of Advanced Glycation End (AGE) Products
Another important factor that is involved with the pathogenesis of diabetic cataract is the formation of AGEs. In diabetic patients with cataract, the elevated glucose starts forming covalent adducts with the lens proteins through a non-enzymatic process called glycation [14,15]. This process is known as one of the most important forms of post-translational modification of proteins under hyperglycemic conditions. Many studies have shown that protein glycation-induced AGEs play a pivotal role in diabetic cataract formation. Therefore, AGE formation assay is used to examine the potential anti-cataract potential of tested compounds. To determine the amount of AGEs, a reaction mixture containing 10 mg/mL of bovine serum albumin and 0.5 M fructose and glucose are mixed with tested compounds. After 15 days of incubation, the fluorescent intensity is measured using a spectrofluorometric detector with an excitation wavelength of 350 nm and an emission wavelength of 450 nm [16,17].

In Vivo Models
Commonly used in vivo models represent specific pathogenesis aspects of human cataract. For example, the diabetic cataract rodent model focuses on mechanisms involved in diabetes-related cataract; selenite-induced cataract addresses oxidative damage-induced cataract; the UV-and steroid-induced models represent their respective associated pathological changes. In various studies, drug effects in these in vivo models correlate well with the pharmacodynamics properties shown in appropriate in vitro models.

Diabetic Cataract
The in vivo diabetic cataract model can be established by using streptozotocin (STZ). After intraperitoneal (i.p.) or intravenous (i.v.) injection, STZ enters the pancreatic β-cell through the glucose transporter 2 transporter (Glut-2) resulting in hyperglycemia [16]. Moreover, STZ is also a source of free radicals that may lead to DNA oxidative damage and subsequent β-cell death. STZ can be administered as a single high dose (e.g., 160 to 240 mg/kg) or as multiple low doses (e.g., 40 mg/kg for 5 days) [18].
Another commonly used diabetic cataract model is AR transgenic mice. The ubiquitous transgenic and lens-specific AR transgenic mice were developed to further prove that polyol accumulation is responsible for diabetic cataract. In both models, sorbitol accumulates in the lens, causing osmotic swelling, and eventually leading to accelerated diabetic cataract formation [10,19].

Selenite-Induced Cataract
Selenite-induced cataract is an effective, rapid, and reproducible model of nuclear cataracts. Selenite cataract is usually produced either by a single dose (19-30 µM/kg body weight) or repeated smaller dosage of sodium selenite (40-50 nmol/g body weight) subcutaneous injection to suckling rat of 10-14 days of age [20]. It has been proposed that selenite treatment leads to altered metabolism in lens epithelium, including loss of small antioxidant molecules such as glutathione (GSH), decreased rate of epithelial cell differentiation, and increased DNA oxidation damage. Such extensive alterations to the epithelium leads to disrupted calcium homeostasis and calcium accumulation in the nucleus of the lens. Increased calcium activates calcium dependent protease m-calpain (calpain II) which results in rapid proteolysis, precipitation of crystallins, and eventually cataract development in rodent lenses [21,22].

UV-Induced Cataract
UV radiation is a major contributor to the pathogenesis of cataract. The strong energy in the UV light can directly cause a DNA lesion in the lens by inducing thymine dimer formation. More importantly, UV can induce cataract formation by the generation of ROS that indirectly induce oxidative Nutrients 2018, 10, 1580 6 of 41 damage to DNA by disturbing cell proliferation in the lens epithelium, altering kinetic properties of enzymes in the energy metabolism, increasing insoluble and decreasing soluble protein, and disturbing the sodium potassium balance, leading to aberrant water balance in the lens [23]. It has been widely accepted that cataract formation is related to oxidative stress induced by continued intraocular penetration of UV light and consequent photochemical generation of ROS such as superoxide and singlet oxygen and their oxidant derivatives such as hydrogen peroxide and hydroxyl radical [24]. Sprague-Dawley rats or mice are exposed to 8 kJ/m 2 UV-B radiation for 15 min to induce cataracts [25].

Steroid-Induced Cataract
As the steroid hormones, glucocorticoids (GCs) have strong anti-inflammatory effects. By binding with the glucocorticoid receptor (GR), GCs have the ability to inhibit all stages of the inflammatory response [26]. Due to its strong anti-inflammatory effects, GCs are widely used in the management of many clinical conditions, including autoimmune disorders, allergies, and asthma, and they also play important roles in chemotherapy and preventing the rejection after solid organ transplantation. However, prolonged use of GCs is associated with the development of posterior subcapsular and nuclear cataracts [26]. The chick embryo has been used to establish an experimental model to study the response of the lens to GCs. When dexamethasone (0.02 µmol/egg) is administered, the lenses of chicken embryos become cataract within 48 h. More recently, the mammalian lens has also been used to establish the steroid-induced cataract models. For example, Brown-Norway rats given a daily 1% prednisolone acetate instillation of a total volume of 1.0 mg/kg or a daily intramuscular injection of 0.8-1.0 mg/kg prednisolone acetate for 10 months successfully induced morphological changes similar to those found in human steroid-induced cataracts [27,28].

Anti-Cataract Phytoconstituents
Based on our literature strategy listed above, the following phytoconstituents are listed in alphabetical order. They have been shown to possess potential anti-cataract efficacy according to the described study models. β-Glucogallin isolated from the aqueous fruit extract of Emblica officinalis Gaertn. (emblic, Indian gooseberry) or Phyllanthus emblica Linn. (Euphorbeaceae) (gooseberry) [29] shows potent activity against human AR in vitro with an IC 50 of 17 µM [30]. Treatment with this compound prevented the sorbitol accumulation by 73% (30 µM) in transgenic human AR expressing lenses ex vivo [30]. This result substantiated the in vitro assay using shared substrate glyceraldehyde at IC 50 of 58 µM. Treatment with β-Glucogallin produced a significant decrease of sorbitol levels in macrophages [31]. Computational molecular docking studies exhibited favorable binding to the active site of between human AR and β-glucogallin. This corroborates the inhibition result of sorbitol production under hyperglycemic conditions in earlier experiments [30] 1,2,3,6-tetra-O-galloyl-β-D-glucose was isolated from the methanolic seeds extract of Cornus officinalis (cornus tree, shan zhu yu) after repeated Sephadex column chromatography. Appeared as an off-white amorphous powder [37], 1,2,3,6-tetra-O-galloyl-β-D-glucose showed the most potent inhibitory activity (IC 50 = 0.70 µM) compared to other secondary metabolites. In addition, AGE formation was also reduced to IC 50 value of 1.99 µM. This compound was further evaluated for its inhibitory effect on ex vivo cataractogenesis activity using rat lenses induced with xylose 20 mM. Treatment with 1,2,3,6-tetra-O-galloyl-β-D-glucose significantly reduced the opacities of the lenses after two days at the concentration of 80 µM [38].

2 ,4 -O-Diacetylquercitrin
Molecular formula: C 25 H 24 O 13 (532.11 g/mol), Melting point: 187 • C. 2 ,4 -O-Diacetylquercitrin was isolated from Melastoma sanguineum (red melastome, fox-tongued melatsome) as a yellow amorphous powder. This compound exhibited the strongest inhibition against RLAR and AGE activities among all the isolated phytoconstituents. IC 50 inhibitory activities for RLAR and AGE were recorded at 0.077 µM and 11.46 µM, respectively. Compared to the positive standards, aminoguanidine (IC 50 = 965.9 µM, AGE) and 3,3-tetramethyleneglutaric acid (IC 50 = 28.8 µM, RLAR), 2 ,4 -O-Diacetylquercitrin inhibited 87 (AGE) and 374 (RLAR) times more efficaciously [40]. Three main constituents of dichloromethane extract from root bark of Garcinia mangostana Linn (mangosteen) were isolated from the hexane/methanol fraction. The result of the study indicated that 3-isomangostin possessed the highest RLAR inhibitory activity with at IC 50 value of 3.28 µM. The presence of cyclization of the prenyl group at the position-two carbon with xanthone derivative enhanced the structure-activity relationship [41]. Gigantol is a bibenzyl-type phenolic compound presents in most herbs of Orchidaceae family [42]. It has been isolated from the stems of various Dendrobium genus such as Dendrobium aurantiacum var. denneanum (die qiao shi hu) [43,44] and Dendrobium chrysotoxum Lindl (fried-egg orchid) [45,46] for anti-cataract activities. As a white solid, gigantol suppresses the damage of rat lenses both in vitro and in vivo in galactose-induced cataractogenesis. The delay in lens turbidity was caused by the inhibition of AR and inducible nitric oxide synthase mRNA expression at an IC 50 of 239.4 µM (65.7 µg/mL) and 32.0 µM (8.8 µg/mL), respectively [43]. Gigantol isolated from Dendrobium chrysotoxum Lindl interpolated into the DNA base pairs in AR gene with a binding constant of 1.85 × 10 3 L/mol, thus, suppressed the gene expression [46].  [48][49][50]. The same constituent was also isolated from Aster koraiensis (Korean starwart) [51], Xanthium strumarium (clotbur, common cocklebur) [33], Artemisia iwayomogi (haninjin) [32] and Artemisia montana [52]. 3,5-di-O-caffeoylquinic acid was reported as the most significant inhibitory activities against AGEs, RLAR and ex vivo xylose-induced lens opacity assays from all isolated constituents. It attenuates AGE formation with IC 50 values ranging from 6 µM to 32 µM, and inhibits RLAR with IC 50 values of 0.2 to 5 µM. These findings are further substantiated by its ability in inhibition of galactitol accumulation at an IC 50 of 153 µM [33] and prevention of xylose-induced opacity of lenses at a concentration of 10 µM [48]. nm (log ε: 3.0) [54]. Isolated from the butanol fraction. 4-O-butylpaeoniflorin was found as an optically active white foam, [α] 25 D -7.8 (c 0.14, MeOH) and later confirmed as an extraction artifact after HPLC analysis. Palbinone inhibits RLAR at an IC 50 value of 11.4 µM. It was suggested that the absence of ring E, side chain of ring D together with double bonds and a conjugated carbonyl group on the ring D played the inhibitory properties. Unlike palbinone, 4-O-butylpaeoniflorin inhibited (IC 50 = 10.8 µM) for AGE activity. The chemical moiety of hydroxy groups in the benzoyl connected to the sugar unit complement the activity [53]. Genistein appears as colorless plates and isolated from the roots of Pueraria lobata (kudzu, Japanese arrowroot) [57,58] and stem bark of Maackia amurensis (Amur maackia) [59]. Both plants are native to Eastern Asia and used as traditional medicine in China, Korea, and Japan. Genistein shows a significant dose-dependent inhibition on RLAR activity (IC 50 = 9.48 µM) compared to the positive control, TMG (3,3-tetramethyleneglutaric acid) (IC 50 = 28.70 µM). Nevertheless, IC 50 was recorded higher at 57.1 µM for the same activity compared to quercetin IC 50 = 10.1 µM) [59]. In an ex vivo lens opacity study genistein suppressed xylose-induced lens opacity at 5 µg/mL (18.5 µM). Further analysis with human lens epithelia cells (LECs; HLE-B3 cells) found that the expression of TGF-β2, αβ-crystallin, and fibronectin mRNAs were reduced, suggesting genistein is protective against lens opacity with antioxidative effects [60]. It is proposed that the chemical moiety with free hydroxyl group at C-7 of genistein attributes to the inhibitory of AR [59].

20(S)-Ginsenoside Rh2
Molecular formula: C 36 H 62 O 8 (622.87 g/mol). 20(S)-Ginsenoside Rh2 is classified under triterpene glycosides and isolated from the root of Panax ginseng C. A. Meyer, (ginseng). It has been used traditionally in East Asia for many years ago with many main active constituents, ginsenosides have been isolated. In RHAR inhibitory activity, 20(S)-Ginsenoside Rh2 showed the most potent inhibitor with an IC 50 of 147.4 µM among all other isolated ginsenosides. It was suggested that the moiety of hydroxyl group at the carbon-20 enhanced the AR activity relationship [61].

Acteoside
Molecular formula: C 29 H 36 O 15 (624.58 g/mol), Melting point: 143-146 • C. Isolated as yellowish amorphous powder from methanolic extract of Abeliophyllum distichum (forsythia) and leaves and stem ethanolic extracts of Brandisia hancei (laijiangteng), acteoside showed the highest RLAR inhibitory activities at IC 50 values ranging 0.83 µM and 1.39 µM compared to four other isolated phenolic glycosides from each plant, respectively. The isolation was conducted by high-speed counter current chromatography using a solvent system of ethyl acetate:n-butanol:water [62]. Isolated acteoside from Brandisia hancei showed potent AGE inhibitory activity with an IC 50 value of 5.11 µM [63].

Canangafruiticoside E
Molecular formula: C 25 H 32 O 12 (524.51 g/mol). The repeated column chromatography of methanolic flower bud extract of Cananga odorata Hook. F. and Thomson generated 25 secondary metabolites and they were tested for RLAR inhibitory activity. The result of the study indicated that among the isolated constituents, canangafruiticoside E possessed the highest activity (IC 50 = 0.8 µM) [71].
Molecular formula: C 45 H 72 O 10 (773.04 g/mol). The isolation of Capsofulvesin A from ethanolic extract of Capsosiphon fulvescens (one of the green algae) showed the strongest RLAR inhibitory activity among all other secondary metabolites, albeit moderate activity at IC 50 value of 52.5 µM. However, the constituent did not show any inhibition against AGE activity [72].

Caryatin-3 methyl ether-7-O-β-D-glucoside
Molecular formula: C 24 H 26 O 12 (506.14 g/mol). The bark of the pecan tree (Carya illinoinensis (Wangenh) K. Koch) has shown good inhibition of AR activity with few compounds have been isolated. Among them, caryatin-3 methyl ether-7-O-β-D-glucoside exhibits the most powerful activity in suppressing the lens AR levels in diabetic cataract rats [73]. The catechol moiety on the B ring of caryatin-3 methyl ether-7-O-β-D-glucoside was suggested to inhibit AR in comparison to the activity of other compounds isolated. In addition, the potent AR activity was also supported by the presence of neighboring O-methyl group in phenolics and an OH group at C-4 [73][74][75]. Caryatin-3 methyl ether-7-O-β-D-glucoside is physically yellow amorphous powder with UV (MeOH) λ max absorption at 350, 330, and 260 nm [73].

C-Phycocyanin (C-PC)
Molecular formula: C 33 H 38 N 4 O 6 (586.67 g/mol). C-Phycocyanin (C-PC), a prominent phytoconstituent found in the stromal surface of thylakoid membranes of Spirulina platensis (a blue-green algae) is a biliprotein that functions to capture light energy to chlorophyll A [76][77][78]. As C-PC is miscible in water but not alcohol and esters, most of the isolations of C-PC use water extraction method [79]. C-PC attenuates selenite-induced cataractogenesis both in vitro and in vivo rat model [78,80]. In vitro study showed C-PC recorded low degree of opacification at 200 µg C-PC with 100 µM sodium selenite [78]. The purified C-PC was active toward the in vivo selenite mediated cataractogenesis showing only slight opacification at 200 mg/kg [78]. Same concentration was observed for naphthalene-and galactose-induced cataract rat models [81]. The protective effect of C-PC in these models were proven from the increment of glutathione, soluble proteins, and water content levels of the lens [79]. Histology study indicated the protection of the lens from oxidative damage. Restoration of lenticular micro-architecture was found with C-PC treated group [77]. C-PC maintains the lens transparency by transcriptional regulation of crystallin, redox genes, and apoptotic cascade mRNA expression [80]. Furthermore, C-PC was suggested to possess protective effects on human LEC by abrogating D-galactose-induced apoptosis through the mitochondrial pathway (p53 and Bcl-2 family protein expression) and unfolded protein response pathway (GRP78 and CHOP expression) [82].

Desmethylanhydroicaritin
Molecular formula: C 20 H 18 O 6 (356.36 g/mol), Melting point: 220-222 • C. The isolation of repeated chromatography of the CH 2 Cl 2 fraction over a silica-gel column and Sephadex LH20 from root methanolic extract of Sophora flavescens (kushen) afforded desmethylanhydroicaritin. Desmethylanhydroicaritin exerted remarkable inhibitory activity of RLAR with IC 50 value of 0.95 µM. Comparable results were observed in RHAR and AGE inhibitions where IC 50 values were observed at 0.45 µM and 294.6 µM, respectively. The presence of prenyl and lavandulyl groups enhanced the RLAR and RHAR inhibitory activities. The 3-hydroxyl group at prenylated flavonoids was suggested for the structural contribution for inhibition of AGE formation [84].
The bioassay-guided isolation of the rhizome of Coptis chinensis Franch (Chinese goldthread) afforded seven secondary metabolites with epiberberine exhibited the highest inhibitory of RLAR activity. The IC 50 of the reported value was 100 µM. Conversely, epiberberine showed a comparable result against RHAR with IC 50 value of 168.1 µM. The chemical moiety of dioxymethylene (ring D) and its oxidized form (ring A) was suggested to enhance the AR inhibitory activities, albeit in moderate effects [88].
Geraniin isolated from Geranium thunbergii shows slightly higher concentration of IC 50 (8.54 µM) in the same activity, however, using rat lens as the source of enzyme [85]. In AGE assay, the activity of geraniin was 96% of inhibition after incubation time of seven days at the concentration of 20 µg/mL (21 µM). Galactitol accumulation in rat lenses incubated with high galactose was inhibited at 39.9% by geraniin with 507.5 µg/lens wet weight (g). It was concluded that geraniin isolated from both plants is a promising agent in the prevention or treatment of diabetic complications.

Hipolon
Molecular formula: C 12 H 12 O 4 (220.22 g/mol), Melting point: 237.5-238.5 • C. Three inhibitors have been isolated from ethanolic extract of Phellinus merrillii (willow) fruiting body and identified as hispidin, hispolon, and inotilone. Hipolon showed highest inhibition against RLAR activity (IC 50 = 9.47 µM) among the three suggesting that phenolic chemical moiety enhanced the activity [90]. Hirsutrin was isolated together with six nonanthocyanin and five anthocyanin compounds from Zea mays L. (corn) for anti-cataractogenesis activity. Isolation of hirsutrin was conducted through bioassay-guided fractionation of ethanolic extract from the kernel of Zea mays L. using repeated column chromatography from ethyl acetate fraction. Hirsutrin showed the highest inhibitory activity in RLAR with an IC 50 value of 4.78 µM and inhibitory constant (K i ) at 7.21 × 10 −7 M from secondary plots of Lineweaver-Burk plots for RHAR assay. Further inhibition by hirsutrin on galactitol formation in rat lens (33.8% inhibition) and erythrocytes (15.7 µM, 32.5% inhibition) supported the efficacy of hirsutrin as the most effective AR inhibitors compared to all isolated compounds [91].

Hopeafuran
Molecular formula: C 28 H 18 O 7 (466.43 g/mol), Melting point: 131-134 • C. Hopeafuran, classified under oligostilbenoids was isolated from the bark of Shorea roxburghii (white meranti) and exhibits the highest RLAR inhibitory activity compared to other isolated secondary metabolites from the same plant. This phytoconstituent inhibits the AR enzyme at an IC 50 value of 6.9 µg/mL (14.8 µM) [92].  [93] and appeared as yellow powder [94,95]. This plant is native to Turkey and widely used as an herbal tea in folk medicine. Isolated hypolaetin has shown the most potent inhibitory activity of AR with IC 50  Isocampneoside II is an active phenylethanoid glycoside isolated from acetone-H 2 O (7:3, v/v) seeds extract of Paulownia coreana (kiri, paotong) at room temperature for 72 h. Paulownia coreana is long cultivated in Eastern Asia, particularly Korea and has been used traditionally in medicines for certain ailments [66]. A total of nine potential inhibitors have been isolated from this plant, however Isocampneoside II is the most potent inhibitor in anti-cataract activities. This compound significantly and uncompetetively inhibited RHAR activity with an IC 50  Cochlospermum religiosum (silk-cotton tree, buttercup tree) has been reported to possess anticataract activity [96]. Purification of hot 95% ethanolic leaves extract of C. religiosum yielded isorhamnetin-3-glucoside. This bioactive compound was obtained as yellow needles and identified as flavonoids with yellowish orange color in alkali, pink in Mg-HCl and reaction with Fe 3+ gives olive green color. Isorhamnetin-3-glucoside at the concentration of 25 µg/mL (52 µM) inhibited further formation of vacuoles and opacity on sodium selenite-induced lens opacity of rat pups. The antioxidant property of isorhamnetin-3-glucoside was suggested to complement its anticataract activity [97]. to as traditional medicine in the form of expectorant. AR activity-guided isolation using column chromatography on a silica gel and gel filtration column afforded kakkalide. Kakkalide significant inhibited AR from Sprague-Dawley rat lenses at an IC 50 of 0.34 µg/mL (0.56 µM), more potent than that of the positive control, tetramethylene glutaric acid (IC 50 = 0.48 µg/mL) [101].
Molecular formula: C 30 H 48 O 4 (472.69 g/mol). Lucidumol A is a new triterpenoid isolated from the ethanolic extract of the fruiting body of Ganoderma lucidum (lingzhi mushroom, reishi mushroom) from a thorough fractionation process [102]. Obtained as a white amorphous powder [103], lucidumol A suppressed the strongest AR activity with an IC 50 [107] and methanolic leaf extract of Vernonia cinereal (purple fleabane) [108] with anti-cataractogenesis activities. Repeated silica gel chromatography after fractionation from both plants yielded lupeol as white needles. It inhibits human recombinant AR activity at IC 50 of 1.53 µg/mL (3.6 µM) [52]. Similar inhibition was observed for AGE with inhibition in the range of 79-82% [107]. The potent activity of lupeol was substantiated with in vivo study using selenite-induced cataract formation in Sprague-Dawley rat pups. Lupeol attenuated the formation of vacuoles and opacity of rat pups lenses at the concentration of 25 µg/g in a dose-dependent manner in selenite-induced cataractogenesis [108]. The potential anti-cataract effect of luteolin is well known [109,110]. As yellow crystalline, luteolin has been isolated from various plants including Platycodon grandiflorum (balloon flower, Chinese bellflower) [111], Vitex negundo (Chinese chastetree, horseshoe vitex) [112], Artemisia montana [52], Perilla frutescens (L.) (perilla, Korean perilla) [69,113,114] and Sinocrassula indica (Chinese crassula) [115]. The selenite-induced oxidative stress treated group with luteolin (isolated from Vitex negundo) demonstrated 80% transparency of the lenses with minor cortical vacuolization and opacity suggesting that the anticataractogenic effect was supported by the antioxidant property based on significant decrease in various antioxidant activities tested [112]. In comparison to the isolated luteolin from different botanicals, luteolin from Platycodon grandiflorum was identified as the highest inhibition with an IC 50 of 0.087 µM (RLAR) [111]. The IC 50 value increases slightly higher to 0.45 µM (Sinocrassula indica) in the same activity [116]. However, isolated luteolin from the same species, Perilla frutescens (L.) of different parts showed different values in RLAR (seeds, IC 50 0.6 µM [113]; IC 50 1.89 µM) [114] and RHAR (leaves, IC 50  Magnoflorine was isolated from Tinospora cordifolia (heart-leaved moonseed, guduchi, giloy) [118] and Coptidis rhizome (coptis root, huang lian) [88] for inhibitory activities against AR. Identification of magnoflorine was conducted with spectroscopic analysis and compared with the literature for both plants. Appeared as yellow powder, this compound exhibited lowest concentration of maximum RLAR activity showing IC 50 value at 3.6 µM from isolation of Tinospora cordifolia. Further analysis showed that magnoflorine inhibited 72.3% of galactose-induced polyol accumulation [118]. Nevertheless, the isolated magnoflorine from Coptidis rhizome possessed marginal inhibition against RLAR with 18% inhibition at a concentration of 146 µM. At this point, it is not clear if the very significant differences in efficacies and potencies were due to technical differences in isolation and/or biological assay. In an attempt to obtain inhibitors of RLAR from Prunus mume (Japanese apricot), mumeic acid-A was found to be the most potent inhibitor from all isolated secondary metabolites. The IC 50 concentration of mumeic acid-A (IC 50 = 0.4 µM) was almost twice that of chlorogenic acid (IC 50 = 0.7 µM) as the positive control [119,120] This was substantiated by the suppression of lens opacity to 72.9% (25 µM) after three days of xylose treatment [127]. A lower IC 50 concentration of RLAR activity was observed for scopoletin isolated from Angelica gigas (dang gui, Korean angelica) with an IC 50 value of 2.6 µM [128] showing the most potent activity among all isolated secondary metabolites. However, scopoletin from methanolic young leaves of Artemisia montana showed higher IC 50 value at 64.5 µM for the same activity [52,129] Semilicoisoflavone B is mostly found in roots and rhizomes of licorice species (Glycyrrhiza sp.) [130]. In searching for potential AR inhibitors, 10 secondary metabolites have been isolated from bioactivity-guided isolation of Glycyrrhiza uralensis with semilicoisoflavone B showed the most potent inhibition of RLAR and RHAR activities. Both inhibition rates were recorded at IC 50 values of 1.8 and 10.6 µM, respectively. Unlike γ,γ-dimethylallyl type prenylated isoflavonoids, semilicoisoflavone B containing γ,γ-dimethylchromene ring on the aromatic ring inhibited AR more strongly. In kinetic analysis of AR inhibition, semilicoisoflavone B did not bind to any substrate and NADPH binding regions of RHAR. Ex vivo analysis showed that this compound highly inhibited sorbitol accumulation in rat lenses incubated with high glucose by 47.0% [131]. The AR and AGE guided isolation of ethanolic bark extract of Rhus verniciflua (lacquer tree) produced nine secondary metabolites with sulferetin and butein as the most potent phytoconstituents for AGE and RHAR, respectively. Sulferetin was isolated as white to off-white crystalline powder [132] and inhibited against AGE activity at IC 50 value of 124 µM, 11 times lower than aminoguanidine (IC 50 = 1450 µM). The RHAR inhibitory activity of butein was reported at IC 50 = 0.7 µM [133]. The efficacies of both phytoconstituents have been suggested on the structure activity relationships of catechol moiety of the B ring and 4 -hydroxyl at the A ring for butein [134] and hydroxyl groups of flavones at the 3 -, 4 -, 5 -, and 7-positions for sulferetin [135].

Syringic Acid
Molecular formula: C 9 H 10 O 5 (198.17 g/mol), Melting point: 206-208 • C. Syringic acid is a phenolic compound and a naturally occurring O-methylated trihydroxybenzoic acid monomer extracted from Herba dendrobii (shi hu). Herba dendrobii, found in the stem of many orchid species of the Dendrobium genus, has been used to improve vision centuries ago [136]. Syringic acid at medium dose (79.97%) isolated from Herba dendrobii improves survival of high-concentration D-galactose-injured human LEC with inhibition ratio of 20.3%. Rat lens turned clear to Grade 0 after 90 days of treatment. Syringic acid inhibits AR activity in a dose-dependent manner with an IC 50 value of 213.17 µg/mL (1075.7 µM). Data suggest that syringic acid downregulates the expression of mRNA of AR [136]. However, the AR inhibition by syringic acid isolated from Magnolia officinalis was weaker with less than 10% of inhibition [137].

Swertisin
Molecular formula: C 22 H 22 O 10 (446.40 g/mol), Melting point: 243 • C. Swertisin appears as pale yellow powdery crystals and isolated from Enicostemma hyssopifolium (najajihva, chota chirayita) methanol extract after repeated column chromatography over silica gel. This compound reacts with ferric chloride and turned greenish brown color as a confirmation test for flavonoids. RLAR activity was significantly inhibited by swertisin at an IC 50 value of 0.71 µg/mL (1.6 µM; 82.3% inhibition at 10 µg/mL) indicating a higher inhibition compared to the other compound isolated, swertiamarin (IC 50 = 7.59 µg/mL). This compound was also found to suppress polyol accumulation (41.7%) in lenses cultured in a galactitol medium [138].

Valoneic Acid Dilactone
Molecular formula: C 21 H 10 O 13 (470.29 g/mol), Melting point: 177-183 • C. The repeated column chromatography and preparative HPLC of seed methanolic extract of Syzygium cumini (L.) Skeels lead to the isolation of six phytoconstituents with valoneic acid dilactone showed the highest activity against RLAR inhibitory activity at IC 50 value of 0.075 µM [87]. Valoneic acid dilactone were the first constituents from this plant reported to possess RLAR inhibitory activity.

Discussion and Outlook
Despite the success in surgical replacement of the cataractous lens with an artificial intraocular lens, discovery of pharmacological prevention and treatment of this blinding disorder has been an earnest, continuous effort in ophthalmology research. In this review manuscript, we summarize findings of phytoconstitutents and their pharmacological effects as potential anti-cataract agents. The large number of interesting compounds is exciting. It raises hope that clinically useful medication may have a good chance to be derived from this sizable collection of chemicals with diverse structural scaffolds.
Many of the compounds have potent and efficacious in vitro pharmacological activities that are consistent with potential anti-cataract effects. For example, 1,2,3,6-tetra-O-galloyl-β-D-glucose inhibits AGE formation with an IC 50 of 2 M. Both 1,3,5,8-tetrahydroxyxanthone and 2 ,4 -O-diacetylquercitrin inhibit AR with IC 50 values below 0.1 M. However, a major limitation of the listed compounds is that although they have been shown to have the appropriate biological actions in a variety of in vitro or ex vivo assays, many of them were not tested in animal cataract models. Additionally, a few have been evaluated in only one animal model. Without relevant in vivo data, it is obviously very difficult to develop the compounds into meaningful treatments for cataract patients.
In addition to the lack of in vivo data, there are other challenges facing development of anti-cataract pharmaceuticals. For example, cataract medication has to compete with the very successfully and generally affordable (as least in developed countries) surgical procedure. Moreover, pharmacological prevention of cataract formation is expected to require a long-term, likely multi-year, administration of medicine, which, to some, is undesirable. Overcoming these challenges necessitates careful considerations of drug safety, convenience of administration, and cost. These concerns may have previously prohibited the development of certain agents. Nonetheless, we feel that phytoconstituents are advantageous compared to conventional synthetic drugs. Many societies have been using plant products from where some of the ingredients are derived for centuries, indicating long-term safety and acceptance. The development path and clinical use will be similar to vitamins and phytochemicals such as lutein and zeaxanthine. If proven safe, cost-effective, and most importantly, efficacious in preventing or reversing cataract formation, phytoconstituents can be a revolutionary approach in the treatment of cataract.

Conclusions
Despite the success in surgical replacement of the cataractous lens with artificial intraocular lens, pharmacological prevention and treatment of this blinding disorder have been an earnest, continuous effort in ophthalmology research. In this review manuscript, we summarize findings of 56 entries of phytoconstitutents and their pharmacological effects as potential anti-cataract agents. The large number of interesting compounds is exciting. It raises hope that clinically useful medication may have a good chance to be derived from this sizable collection of chemicals with diverse structural scaffolds.
Many of the compounds have potent and efficacious in vitro pharmacological activities that are consistent with potential anti-cataract effects. For example, 1,2,3,6-tetra-O-galloyl-β-D-glucose inhibits AGE formation with an IC 50 of 2 µM. Additionally, both 1,3,5,8-tetrahydroxyxanthone and 2 ,4 -O-diacetylquercitrin inhibit AR with IC 50 values below 0.1 µM. However, a major limitation of the listed compounds is that although they have been shown to have the appropriate biological actions in a variety of in vitro or ex vivo assays, many of them were not tested in animal cataract models. And a few have been evaluated in only one animal model. Without relevant in vivo data, it is obviously very difficult to develop the compounds into meaningful treatments for cataract patients. We feel that, by listing the comprehensive collection of phytocontituents in one place, this manuscript serves as an overview and perhaps an inspiration to prompt additional studies in this important research area. Collaborative efforts between phytochemists and cataract researchers are promisingly fruitful.
In addition to the lack of in vivo data, there are other challenges facing development of anti-cataract pharmaceuticals. For example, cataract medication has to compete with the very successfully and generally affordable (as least in developed countries) surgical procedure. Moreover, pharmacological prevention of cataract formation is expected to require a long-term, likely multi-year, administration of medicine, which, to some, is undesirable. Overcoming these challenges necessitates careful considerations of drug safety, convenience of administration, and cost. These concerns may have previously prohibited the development of certain agents. Nonetheless, we feel that phytoconstituents are advantageous compared to conventional synthetic drugs. Many societies have been using plant products where some of the ingredients are derived from for centuries, indicating long-term safety and acceptance. The development path and clinical use will be similar to vitamins and phytochemicals such as lutein and zeaxanthine. If proven safe, cost-effective, and most importantly, efficacious in preventing or reversing cataract formation, phytoconstituents can be a revolutionary approach in the treatment of cataract.