Silymarin: a review on paving the way towards promising pharmacological agent

ABSTRACT Advancements in the field of research and development have paved toward the use of herbal agents to cure and control health maladies. Milk thistle (Silybum marianum L.) is one of the herbs that possess pharmacological and therapeutic attributes. Various bioactive components in milk thistle have a key role in pathological conditions like blood-related disorders, hepatic ailments, arthritis, ulcerative colitis, and carcinomas. Among various bioactive components, silymarin is an active agent which isa standardized mixture of flavonolignans (70%–80%) along with silybin, silydianin, and silychristin, with silybin being the main active chemical component. Silymarin has been used medicinally to treat liver disorders, including acute and chronic viral hepatitis, toxin/drug-induced hepatitis, cirrhosis, and alcoholic liver diseases. Traditionally, the plant was employed to increase human reproduction ability, possess reactive oxygen species (ROS) scavenging, antioxidant, anti-inflammatory and immunomodulatory properties, has UV-induced skin damage protection, maintains the menstrual cycle, lessen viral infection, decrease fibrosis, and blood glucose as well as improve functions of the liver, pancreas, and reproductive cells. This review summarizes the basic as well as advanced therapeutic effects of silymarin which will ultimately lead researchers in developinphytopharmaceutical-based complementary medicine.


Introduction
Herbal drugs have been considered the major agents to treat and prevent pathologies from the time of ancient Greeks (Theophrastus, 4th century B.C.) and Romans (Pliny the Elder, 1st century A.D.), among which milk thistle seeds (Silybum marianum), commonly termed as lady's thistle and Saint Mary's thistle), have been employed as hepatoprotective.The advancement in plant-based extracted

Chemical nature of silymarin
Flavonolignans are the common name of silymarin compounds.Even this basic name indicates that the molecular structure of this compound can be separated into two components, one is hybrid or the other is non-conventional lignans which are more suitable for the silymarin compounds group.Flavonolignans are naturally found in the form of stereoisomers because the basic structure contains several symmetrical centers.The chemical structure of milk thistle was described in Figure 1. [19]The main silymarin compounds reported in the scientific literature are taxifolin, silybin A and B, silychristin, isosilychristin, silydianin, and silychristin.Naturally occurring iso-and trans-diastereoisomers of silibinin (silybin A and silybin B) and isosilibin (issilybin A and issilybin B). [20]

Effect of silymarin on oxidative stress
Various studies revealed that Silymarin could exert antioxidant properties in several mechanisms, which includes direct hindrance in free radical production, progression and enhancement of antioxidant enzyme activity by the transcription factor, and activation of an array of vitagenes which are ultimately responsible for protective molecule synthesis. [21]It is considered that the existence of hydroxyl groups in the molecular configuration of silymarin components is responsible for antioxidant and radical scavenging activity.Therefore, Silymarin can counteract the negative effects of the inflammatory process and oxidative stress by scavenging free radicals and controlling inflammatory cytokines.Silymarin used in vitro antioxidant activity by tackling the free radicals 1,1-diphenyl-2-picryl-hydrazyl (DPPH) and 2,2'-azino-bis (3-ethylbenzene-thiazoline-6-sulfonic acid diammonium salt) (ABTS). [22]The antioxidant properties of Silymarin may operate through a variety of methods.These comprise hindering the enzyme activity that produces reactive oxygen species, stopping free radicals production, intestinal ion chelation, encouraging the manufacture of protective molecules, and activating antioxidant enzymes. [21]Silymarin's antioxidant effects have been shown to increase poly-(ADP-ribose)-polymerase function by maintaining NAD+ homeostasis, sirtuin 1 activity, and the AMP-activated protein kinase pathway (all significant regulatory processes associated with oxidative stress). [23]Furthermore, the radical scavenging properties of Silymarin enhance hepatic lipid homeostasis by reducing denovo lipogenesis through the down-regulation of acetyl-CoA carboxylase, fatty acid synthase, and peroxisome proliferator-activated receptor. [24]

Effect of silymarin in reproductive system
The community's psychological and economic load caused infertility in humans and requires the use of fresh, practical, and economical treatments.Investigations revealed that oxidative stress plays a significant role in human infertility.Furthermore, smoking, Alcohol abuse, chronic stress, obesity, reproductive system inflammation, urogenital trauma, heavy metals, chemicals, heat, pesticides, and electromagnetic radiation are also considered triggering factors that directly impact spermatogenesis and hence induce infertility. [25]everal plant extracts are available which help to scavenge free radicals and defend cells.As a result, administering antioxidants like Silymarin can aid in boosting conception rates.
In addition, numerous investigations revealed that Silymarin has antioxidant and preventive capabilities against the destruction that chemotherapy drugs and environmental pollutants can cause to sperm, and eggs.Studies on male and female animals have shown that Silymarin has a positive impact on fertility.To decide the dosage and length of treatment, more research, especially in humans, should be conducted while taking into account some phyto-estrogenic properties of Silymarin.Numerous studies have demonstrated that antioxidants may enhance oocyte and sperm motility that increased healthy pregnancies. [26]A specific level of reactive oxygen species (ROS) is essential for optimal cell function; an imbalance between the generation of ROS and antioxidants can cause cell cycle abnormalities and the advancement of apoptosis.The production of ROS is heavily influenced by both endogenous factors like the mitochondrial respiratory chain and external factors like alcohol, smoking, and exposure to environmental contaminants. [27]Oxidative stress caused reproductive systems cell damage that is responsible for infertility in both males and females. [28]

Male infertility
The majority of studies have focused on men.Because oxidative stress caused the harmful effects on sperm than oocytes.Different studies conclude that oxidative stress caused male infertility in 2 different ways: (1) sperm membrane dysfunction, decreased sperm movement, and subsequent disruption in sperm union with the egg (2) DNA damage and embryo abnormalities. [29,30]onsumption of antioxidants with diet modification is an effective solution to decrease DNA and RNA abnormalities and enhance fertility. [31]Sperm may be affected by free radicals production and infection response, which is linked to DNA break or change in the motility, fusion, and viability of reproductive cells.All of the above problems could lead to fetus abnormalities, abortion, and infertility. [32]hen human lymphocytes and sperm cells are open to the elements of food mutagens like 3-amino-1-methyl-5 H-pyrido (4,3-b) indole (Trp-P-2), 2-amino-3-methylimidazole-(4,5-f) quinoline) and flavonoids, as a result, DNA damage was reduced by the antigen-toxic activity of flavonoids (Silymarin, myricetin, quercetin, kaempferol, rutin, and kaempferol-3-rutinoside). [31]The effects of Silymarin, myricetin, and quercetin on sperm were antigenotoxic.Phenolic hydroxyl groups have been linked to antioxidant benefits, and the addition of hydroxyl groups to A and B rings revealed an increased effect of these effects.Antigenotoxic effects in lymphocytes and sperms at the same dose demonstrated that this has a protective effect. [33]ne more study showed that sodium arsenite stimulated the generation of free radical species and induced oxidative stress in sperm cells.Therefore, Silymarin has antioxidant activity, so it can increase sperm survival, motility, and mitochondrial membrane potential by scavenging free radicals and boosting antioxidant enzyme activity. [34]Another study by Eskandari and Momeni explained that ram epididymal spermatozoa that were exposed to arsenite had considerably higher plasma membrane and acrosome integrity as compared to the control group. [35]nother most important harmful sperm disruption is due to lipid peroxidation (LPO) during sperm storage.Silymarin polyphenols can reverse this obliteration process as well as improve the quality of ram sperm when it is used as a supplement with caproic acid. [36]Another study held on bull sperm that showed Silymarin provides a protection medium and optimistic effect on bull sperm in chilled and frozen storage conditions. [37]he mammalian spermatozoa cell membrane contains polyunsaturated fatty acids, which create a susceptible situation that encourages LPO to produce Radicals to damage the cell.Significant increases in testosterone levels, spermatid diameter, and testicular-associated factors were observed when impact of silymarin was observed on testicular tissues. [38]Abedi and his coworkers studied the effects of Silymarin on spermatogenesis, testicular tissues, and hypothalamic-pituitary-gonadal axis hormones (testosterone, luteinizing hormone, follicle-stimulating hormone).Experimental rat's blood levels confirmed an increase in spermatids, spermatozoa cells, FSH, LH, gonadotrophin hormone and testosterone levels. [39]El-hanbuli and his research associates gave silymarin to Rats and studied testosterone levels the results cleared that silymarin drug has an important function in the reduction of histopathological and testosterone biochemical adverse reactions. [40]When milk thistle seed and rosemary leaves were fed to rabbits, the results concludes that sperm concentration, live sperm amount, sperm production quantity, motile sperm, fertility rate, and testosterone level were improved. [41]s a result, Silymarin demonstrated a remarkable contribution to the enhancement of spermrelated parameters and fertility.Men who have varicocele may experience infertility due to hypoxia (caused by venous stasis), Leydig cell death, decreased testosterone levels, elevated testicular temperature, and accumulation of phospholipid and androgen receptor dysfunction.Studies revealed that Silymarin can lessen the negative effects of hypoxia.Additionally, Silymarin improved glucose in germinal cells, spermatozoa nuclei maturity, and spermatogenesis. [42]Furthermore, Silymarin enhances the molecules of thiol in experimental mice.So, sperm movement increased and DNA disruption was reduced. [43,44]The results for the combined effect of celecoxib and Silymarin on varicocele cells revealed that these two drugs play a vital role in lowering the varicocele-induced injuries by inhibiting enzymes (cyclooxygenase) and enhancing the antioxidant activity to safeguard the germinal cell RNA. [9]More studies have revealed radioprotective properties of Silymarin in the testis tissues of mice and rats. [45,46]Silymarin's antioxidant potential improved the reproductive function and decreased the progesterone, estrogen, and testosterone level of thymoquinone in male rats exposed to a chemical pollutant (benzo pyrene). [47]ethotrexate (cytotoxic) [48] doxorubicin (antineoplastic) [49] and Nickel chloride [50] damage the testicles, shrink seminiferous tubules and the main spermatocyte, resulting in a decrease in sperm motility and count, and the production of immature and aberrant sperm.Silibinin has the ability to increase the amount and quality of sperm. [51]The agonistic or antagonistic effects of Silymarin on the estrogen receptors might depend on the body's estradiol level, amount, and duration of, therapy and this drug cause a dramatic drop in diabetes-induced reduction in sperm count. [9]

Female infertility
Female pregnancy issues such as abortion, miscarriage, preeclampsia, endometriosis, PCOS and infertility may be caused by an imbalance among pro-oxidants and antioxidants. [52]Additionally, several investigations have demonstrated that oxidative stress causes cell death in preovulatory follicles.Consuming synthetic antioxidants like vitamin C, E, A, and B complex, coenzyme Q10, carnitine, zinc, selenium, copper, and taurine as well as nonenzymatic antioxidants such supplements might help to maintain stability. [9]The sound effects of Silymarin on granulosa cell apoptosis and folliculogenesis in healthy females that have undergone in-vitro fertilization (IVF) were assessed in a randomized, double-blind experiment.However, they were unable to identify any associations between silymarin benefits and other effects such as endometrial thickness, oocyte retrieval, or follicular maturation.But results revealed that using Silymarin and gonadotropin simultaneously from the first day of ovulation can cause a decline in granulosa cell apoptosis. [53]xidative stress (induced by sodium nitroprusside in this study) is one of the variables that hinder the IVF of bovine embryos.Silymarin may decrease the adverse effects of LPO and improve the viability and morphology of bovine oviduct epithelial cells.Moreover, an increase in the growth rate of bovine IVF embryos has been observed. [54]n the ovarian tissue of juvenile female fish, the extracts of Cucurbita pepo, Silybum marianum, Linumusitatissmum, and Vitex agnus-castus plants were evaluated and concluded that due to 17 estradiol Silybum marianum has the most effect on improving the fertility among these extracts. [55]he estrogenic effects of Silymarin were demonstrated by useful modification in experimental rats treated with the silymarin drug In vitro research cleared that Silymarin activates some receptors such as silybin and taxifolin have partial to complete estrogen receptors.Other flavonolignans and silybin A lacked estrogenic characteristics (Figure 2).It is suggested that extracts are purified using flavonolignans without estrogenic properties because estrogenic effects should be taken into consideration as a clinical negative. [56]yclophosphamide stimulates free radical production in female follicles.Drastically, the development and number of antral follicles were enhanced in the silymarin-treated group, therefore, Silymarin might be used as an ovarian follicle protector. [57]

Effect of silymarin on hepatic cells
In the past years, there was a considerable rise in the disorder of chronic liver disease with increasing incidence of nonalcoholic fatty liver malady and chronic liver disease, a key cause of global death. [58]tudies suggest that hospitalized patients due to chronic liver disease have doubled over the previous years.The rise in hepatic diseases has raised serious concerns about the people at risk for cirrhosis and liver transplants. [59,60]As a result, reasonable and efficient behavior options to lower the morbidity and mortality linked to chronic liver disease.Several plant-based medicines, Silybum marianum, also identified as milk thistle, were used to cure and prevent liver disorders.Preclinical tests employing a variety of medicines, including Legalon, which comprises the Eurosil drug, and Silymarin have specifically demonstrated promising protective properties. [24]Silymarin's antioxidant, regenerative, antiproliferative, anti-inflammatory, and immunomodulatory characteristics help to improve liver function.For clinical purposes, Silymarin has a wide range of chronic liver disease curative agents.Liver diseases such as viral hepatitis (HCV), primary liver malignancies, cholestasis (pregnancy and nonpregnancy related), nonalcoholic fatty liver disease, and drug and toxin-induced hepatic disorders are included. [61]coholic liver disease Fatty liver disease (FLD) is caused by the accumulation of excess fat in the liver, which can lead to serious liver disease for many people.Approximately liver cirrhosis is caused by alcohol, which is a major risk factor for liver problems.Significant alcohol intake causes fatty liver disease at first, which can turn into cirrhosis. [59]The drug Silymarin has been investigated in patients with liver cirrhosis and/or alcohol-related liver disease [62] Salmi et al [63] studies 97 patients of liver patients, in these patients 78% of patients take alcohol on a daily basis.Salmi administered 420 mg of silymarin for dose per day up to 4 weeks and they reported that ALT, AST, and liver function improved.Feher et al [64] study the 36 patients with liver diseases.They concludes that the intake of silymarin for 6 months decreased the ALT, AST, and bilirubin levels and synthesized the procollagen. [65]

NonalcoholicFatty liver disease
The use of Silymarin for nonalcoholic fatty liver disease has shown positive results in several randomized control study (often referred to as RCTs) that demonstrates the therapeutic potential of Silymarin. [66]Hepatic fibrosis and inflammation are two clinical symptoms of nonalcoholic fatty liver disease.This is because Silymarin has metabolic effects on insulin resistance and hyperlipidemia, along with its anti-inflammatory, antioxidant, antifibrotic, and pro-regenerative activities, which are linked to nonalcoholic fatty liver disease. [67]The use of Silymarin for the prevention of nonalcoholic fatty liver disease is favorable but still needs more research with bigger standardized RCTs.As a recent meta-analysis encompassing 500 individuals suggests an improvement in liver function when they consumed silymarin drugs. [66]ilymarin drug (140 mg, 3 times for 28 days) has more bioavailability and no side effects in individuals with acute hepatitis.After receiving silymarin therapy, the participants in this clinical research followed an additional 4 weeks of screening. [68]In one more study, nonalcoholic fatty liver disease patients when consumed a dose of silybin (94 mg) and phosphatidylcholine (194 mg) with vitamin E (89.28 mg), twice a day for a year.Then this drug had no significant adverse effects.However, some side symptoms like diarrhea, itchiness, and dysgeusia are developed. [69]

Drug-induced liver injury
When humans are exposed to various chemicals, significant increases in oxidative stress can lead to drug-induced liver damage. [70]Free radical production and the body's capacity to activate antioxidant enzymes that acts as counterbalances as a complementary defense mechanism typically coexists in harmony.The antioxidant potential of Silymarin has significant preventive measures for chronic liver diseases such as jaundice, cirrhosis, and hepatitis. [71]Silymarin administration prevents hepatic dysfunction and restored normal liver functionality in studies on CCl 4 -hepatotoxicity in rats. [72]ilymarin functions as an antioxidant by reducing oxidative stress and preventing glutathione loss. [21]According to the Hep573 study, Silymarin and antioxidants in a complex naturopathic cocktail may help HCV patients achieve normal ALT levels and a higher quality of life overall by treating diseases that aren't often appropriately treated by the antiviral "treatment."These results were primarily seen in HCV genotype 1 patients, these people lack endogenous antioxidants such as glutathione. [73]These studies justified that Silymarin has God-gifted therapeutic potential for liver diseases.Figure 3 shows the pharmacological agents of silymarin for the treatment of various diseases.

Effect of silymarin on fibrotic cell
Hepatitis B and C virus (HBV, HCV) infection can lead to the development of chronic liver disease that can lead to liver fibrosis and eventually to liver cirrhosis.Silymarin's antifibrotic effect is mainly carried by its ability to prevent hepatic stellate cells from changing into myofibroblasts by blocking fibrogenic pathways, likewise, these are involved in cytoskeletal development, profibrogenic collagen, and electron transfer chains.In particular, Silymarin inhibits NF-kB, down-regulates TGF-ß1 mRNA, and suppresses activation of liver stellate cells.Studies in animal models that showed Silymarin can decrease the development of early fibrosis. [74]hen the recommended dosage of silymarin is given in the early phases of fibrotic that is capable of blocking the fibrogenesis mechanism and the advancement of liver fibrosis.50 mg and 200 mg of silymarin dose can be used to reduce fibrogenetic mechanism and progression of liver fibrosis.Silymarin significantly decreased the elevation of aspartate aminotransferase (AST), alanine aminotransferase, and alkaline phosphatase in serum, and also reversed the altered expressions of α-smooth muscle actin in fibrotic tissue. [75]kda et al. [76] currently reported that silymarin with ginger has significantly decreased the severity and incidence of liver fibrosis.The activation of hepatic stellate cells via TGF-1 expression the and stability of mast cells were linked to silymarin's anti-fibrotic and anti-inflammatory actions.These results indicate that silymarin prevents hepatic fibrosis by reducing inflammation and hypoxia throughout the fibrogenesis process. [77] Promising pharmacological agents of silymarin for the treatment of various diseases.

Effect of silymarin on immune function
Inflammasomes and NF-B activation are blocked, which are essential for controlling the human defense response in inflammatory conditions, Silymarin's immunomodulatory activity has an antiinflammatory effect. [78](Silymarin can also activate the farnesyl X receptor, which in response can lessen liver inflammation, and repair the insulin receptor substrate-1/PI3K/Akt pathway, which can alleviate NAFLD-induced insulin resistance and steatosis. [79]Therefore we can say that Silymarin's anti-inflammatory and antioxidant properties decreased virus diseases such as chronic HCV infection. [13]Numerous studies have demonstrated that silymarin reduces inflammation by inhibiting the nuclear factor-kappaB (NF-B) signaling pathway and TNF activation.Additionally, it has several immunomodulatory actions that are dose and time-dependent.Cyclooxygenase-2 (COX-2), prostaglandin E2 (PGE2), and inflammatory cytokines are NF-B regulated gene products that are suppressed by silymarin, which has a strong anti-inflammatory effect.Silymarin functions as an immunomodulator by inhibiting T-lymphocyte function at low dosages while inducing inflammation at high doses.According to studies, silymarin can inhibit oxidative stress and nitrosative immunotoxicity as well as immune-mediated liver disorders, allergies, preeclampsia, cancer, and autoimmune illnesses.Moreover, silymarin has been shown to have dual effects on the growth and cell death of several cell types.Neurodegenerative illnesses like Parkinson's disease, Alzheimer's disease, and multiple sclerosis arise in the central nervous system (CNS) as a result of inflammatory reactions mediated by activated microglia (the resident macrophages in the CNS).Notably, some studies investigated the neuroprotective effects of silymarin in mesencephalic mixed neuron-glia cultures against lipopolysaccharide (LPS)-induced neurotoxicity.Researchers demonstrated that silymarin inhibits microglia activation, and protects dopaminergic neurons from lipopolysaccharide (LPS)-induced neurotoxicity.According to these findings, silymarin decreases the synthesis of inflammatory mediators including TNF-and nitric oxide (NO), which minimizes the destruction of LPS [80]

Protective effect against viral infection
The main mechanisms by which Silymarin prevents additional damage in situations of toxin-related hepatic injury are the modulation of membrane permeability and the aggressive hang-up of toxins at binding sites.This stops these hazardous compounds from being absorbed, especially in the hepatic phalloidin-transporting system. [81]he management of viral hepatitis has undergone a significant change since the introduction of antiviral medication.Despite this, there may be a demand for other therapy approaches like Silymarin due to high treatment costs and accessibility concerns.Unfortunately, [82] designed a double-blinded experiment comparing the effectiveness of Silymarin to placebo but it did not reduce serum alanine aminotransferase (ALT) levels.Additionally, silymarin treatment for HCV patients resulted in somewhat lower ALT and aspartate aminotransferase (AST) levels, but these benefits were found to be too unpredictable to have any evident therapeutic value. [69]atients with hepatitis C virus (HCV) infection and non-cirrhotic hepatitis were not successful in one trial using the interferon drug.When these patients received Silymarin for 7 days orally in doses of 140, 280, 560, and 700 mg twice a day.Nausea and headaches were reported who consumed 280 mg of Silymarin.The remaining doses have no drug-related side effects were recorded. [83]The HCV-infected participants received PEGylated-interferon alfa 2b, ribavirin, and one supplement (94 mg of silybin, vitamin E, and phospholipids) every day for a year.Although some symptoms like reduction in weight, muscle cramps, irritability, hypertension, joint pain, and high cholesterol were noticed during this experiment, therefore no severe adverse effects were recorded. [84]In the second experiment, 20 mg/kg of silymarin injection was given over the course of 20 days to 27 male and female patients who had been treated with ribavirin and interferon but still had hepatitis C virus.Blood was drawn on specified days of 1, 8, 15, and 22 as well as four weeks later.However, some patients reported heat sensations and some patients reported gastrointestinal symptoms while patients receiving silymarin medication, showed that Silymarin did not have significant toxicity.But each participant in this trial had a high level of bilirubin (0.98 to 2.12 mg/dL). [85]630 mg/d for 6 months dose of Silymarin has high bioavailability for HCV patients. [86]Silymarin (1,400 mg/day over 110-350 minutes) was given intravenously to hepatitis C patients for two days as part of PEGylated-interferon alpha-2a treatment, and it was well tolerated.However, some modest adverse effects were also noted, including nausea, vomiting, diarrhea, and a feeling of heat.These issues did not last more than two days. [87]Another study found that oral doses of 420 and 700 mg of Silymarin twice a day for 6 months were acceptable.Therefore, it can be said that Silymarin is used as a medicine for hepatitis C patients. [82]

Antituberculosis
One Silymarin (140 mg) tablet was administered to tuberculosis patients after every 8 hours they had taken antituberculosis medication.No major side effects of Silymarin were noticed after 2 & 4 Weeks.In various studies, this dosage of Silymarin was used for 8 weeks without any negative side effects. [88]oreover, taking 140 mg of Silymarin orally twice a day for 2 months was acceptable, and no negative effects were reported. [89]In another clinical trial counting tuberculosis patients, oral administration of 140 mg of Silymarin thrice a day for 15 days was examined, and concluded that this dose is also acceptable and had no significant adverse effects.But there were some intestinal issues produced, such as nausea, vomiting, and anorexia observed. [90]

Immune deficiency virus (HIV)
Silymarin (150 mg/d dose) was given for 15 days to HIV-infected patients to investigate its effects on the pharmacokinetics of darunavir-ritonavir.In this study, the Darunavir-ritonavir drug with silymarin drug was delivered to 15 experimental males.In combination with ritonavir -darunavir, Silymarin was more bioavailable and safe. [91]

Effect of silymarin on tumor cell
In cancer patients, silymarin drug with 140 mg silymarin concentration taken 2 times per day before cisplatin drug intake for 1 week has bioavailability and no reaction. [92]In a different study, cancer patients received the medication silymarin at a dosage of 420 mg per day for 20 days.There have been no known side effects. [93]In another study, 2-20 mg of silybin-phytosome (siliphos) when taken orally, three times a day, to males with prostate cancer for one month.There have been some reported adverse reactions, including gastrointestinal issues and liver malfunction, as well as additional undesirable effects, including a higher calcium, creatinine, and halitosis levels. [94]ne more research was conducted to evaluate the effects of Silymarin on the severity and incidence of mucositis caused by radiation.After receiving radiotherapy, people having cancer of the neck and head, were given silymarin pills (420 mg/day) for 15 days while the behaviour was monitored, the study reported that silymarin had no negative effects on the body. [95]n another study, a group of patients with prostate cancer was given Silymarin with selenium in different combinations.The Silymarin with a selenium dose of 570 mg/d, for 6 months caused no side effects and was effective in reducing prostate cancer growth. [95]Colorectal cancer patients were given silybin, phosphatidylcholine, and vitamin E daily in addition to regorafenib.The latest research concluded that this experiment has no harmful effects on silybin. [96]ilymarin shows anti-cancerous properties considered to be linked to oxidative stress inhibition, apoptosis induction, growth cycle arrest, and mitochondrial pathway inhibition.Animals with hepatocellular carcinoma (HCC) treated with Silymarin have revealed the anticancer effects at various stages of hepatocarcinogenesis cells.Silymarin is ideally suited as a potential therapy for people with chronic liver disease due to its capacity to promote hepatic regeneration.Particularly, there is a relationship to ribosomal RNA synthesis, probably through polymerase 1 activation. [97]

Effect of silymarin on arthritis and ulcerative Colitis
Arthiritis and ulcerative colitis is a chronic inflammatory diseases.This can be treated with antioxidant and anti-inflammatory compounds.Silymarin was supplied at a dose of 300 mg/day for 1 month to patients with knee osteoarthritis to examine its anti-inflammatory impact.Silymarin was studied as compared to non-steroidal anti-inflammatory drugs.As a result, there was a significant decrease in the levels of interleukins, and the patients tolerated Silymarin well with no harm. [13]Patients with rheumatoid arthritis received treatment for three months with methotrexate in a dose of 12 mg/ week along with silibinin in the range of 120 mg twice a day.There were no negative effects or reports of silibinin toxicity in this trial. [98]lcerative colitis is a chronic inflammatory bowel disease.This can be treated with antioxidant and anti-inflammatory compounds.50 mg and 100 mg of silymarin dose per kg have been effective on acetic acid induces colitis rats.100 mg/kg administration dose of silymarin has reduce the inflammation, crypt damage, and macroscopic damage. [99]hen randomized clinical study was conducted on individuals with ulcerative colitis to evaluate the favorable effect of Silymarin.As part of their routine treatment, they received silymarin, and after six months, the dosage was decreased to 140 mg daily when taken orally.Silymarin side effects include nausea, vomiting, headaches, and diarrhea at high doses.Although these side effects were temporary, however, it did not show any harmful or toxic effects. [100]Moreover, Hoseini et al. [101] worked on the nanosilymarin drug for colitis patients.They experimented on ulcerate colitis rats.They administered 50, 100, and 200 mg/kg nano silymarin dose in ulcerates colitis rats orally for 14 days.Results indicate that nanosilymarin reduced the rats colon inflammation by inhibit the TLR4/NFκB molecular pathway. [101]

Effect of silymarin on pancreas
In an experimental study, 140 mg of silymarin was given three times daily to diabetic individuals for three months together with renin-angiotensin inhibitor enzyme.But one patient had a previous cardiac illness, therefore, the patient died as a result of myocardial infarction during this study.Common side effects of Silymarin include headaches, diarrhea, and vomiting. [102]Another study with the patients having hypercholesterolemia and diabeties, was given tablets of Berberol® (each tablet containing Berberis aristata extract 588 mg, and silymarin 105 mg) 2 times a day for 1 year.Berberol® had no toxic effect and was safe for consumption, but it eventually caused asthenia and headache.Additionally, one patient in the Berberol® plus statins group experienced headaches and constipation for up to several days, and two patients in the group experienced muscle cramps. [103]In another study, patients with type 2 diabetes were given a total daily dose of 1,000 mg of berberine and 210 mg of Silymarin for 4 months.However, 15% of the participants from this research observed mild abdominal discomfort that was temporary with no other side effects. [103]Another study showed that the diabetic patients that take 140 mg of dried Silymarin extract orally after every 8 hours for 2 months, there were no side effects or symptoms observed. [104]

Effect of silymarin on blood diseases
Children did not experience any damage from milk thistle supplements that were diagnosed with acute lymphoblastic leukemia and hepatic toxicity.One standard silibin capsule taken at the recommended dosage and frequency for a month had no negative side effects. [65]Another study reveals that the patients with beta-thalassemia were given deferiprone in the range of 20-40 mg/ kg/day and 140 mg of Silymarin three times a day for 180 days without experiencing any negative side effects.When the medicine was removed, ferritin and iron levels declined, but there were no discernible changes in aspartate aminotransferase, serum creatinine, aminotransferase and alanine levels.Silymarin also had an iron-chelating effect. [105]A study showed that the patients were given Silymarin three times a day orally in the dose of 140 mg for about six months long with subcutaneous desferrioxamine.Only three of the trial's participants experienced bloating and dyspepsia.However, after 4 weeks, these problems subsided, and Silymarin was more widely accepted with no significant side effects. [106]In this trial, Silymarin was given orally at a dose of 420 mg daily for 6 months to patients with thalassemia to assess its efficacy on the serum levels of interleukins. [107]Patients with thalassemia were administrated with desferrioxamine in combination with Silymarin for about 8-9 months to lower serum ferritin levels and did not show any side effects. [108]To test the medication's ability to chelate iron, patients with hereditary hemochromatosis were given 140 mg of silybin (Legalon® Forte) every day for about three months, the study showed silybin's ability to chelate iron, which can be used to cure hereditary hemochromatosis without having any negative side effects. [109]

Conclusion
We have compiled both human and animal studies in one study.This study also concludes Silymarin's mutagenicity, genotoxicity, and cytotoxicity.Although some studies utilized silibinin, Silymarin has typically been standardized based on silybin.This analysis has several restrictions.In the majority of in vivo studies, Silymarin was combined with another medication to enhance the therapeutic result or lessen the adverse effects of the primary medication.Consequently, side effects of the primary medication may hide silymarin benefits.Few papers in animal research specifically addressed Silymarin's toxicity but the majority of studies demonstrated that Silymarin along with other chemicals or medications can be used.As a result, silymarin consumption during pregnancy in humans should be monitored.Silymarin has low bioavalibity due to low water solubility, therefore formulations for Silymarin are another important topic.This can be improved by appropriate modification.The formulation and absorption of silymarin were often not discussed in the research.The US Food and Drug Administration has not yet given its consent to the essential problem of Silymarin standardization.Several flavonolignans, including silybin, silibinin, silydianin, and silychristin, are present in milk thistle extract.It is best to evaluate the toxicity and safety of these chemicals separately, especially silybin, which is the main component of Silymarin and accounts for the majority of its medical effects.As the numerous animal studies in this essay come to an end, it can be said that milk thistle is safe for use by humans, has therapeutic potential, and rarely causes side effects like minor gastrointestinal discomfort.More pregnancy-related studies are required.Although it has important pharmacological agents that improve human physiology.

Figure 1 .
Figure 1.Chemical structure of the basic components of Silymarin.

Figure 2 .
Figure 2. Pharmacological effect of silymarin on the male and female reproductive system.