In Vitro Anticancer studies on Colon rectum cell line

CONTENT

Introduction

Literature Review

Procedure

Result and Discussion

Conclusion

References

I. INTRODUCTION

Quercetin

Quercetin is scientifically known as 3,3',4',5,7-pentahydroxyflvanone which was given by IUPAC. Quercetin is derived from a latin word known as "Quercetum" that means oak forest. This comes under the class known as Flavanols that is not produced by human body. The quercetin is yellow in color that is poorly soluble in hot water, partially soluble in alcohol and is insoluble in cold water. It is most widely used Flavonoid worldwide to treat many disorders. It shows many biological activities like antiallergenic, antiviral, vasodilating and anti-inflammatory disorders, anti-platelet, anti-tumor, anti-oxidant and treatment of neuro degenerative disorders. There are many sources of quercetin as it is mostly found in fruits, vegetables and certain beverages. It is used in preventing injuries as they activate the anti-oxidant enzyme, reduction of alpha tocopheryl radicals, inhibits the oxidases, metal chelating activity, mitigation of oxidative stress caused by NO, increases uric acid level, increases the antioxidant properties of low molecular antioxidants and they also oxidize other compounds and act as pro oxidants.

Antioxidant properties:

Quercetin is a plant derived aglycone form of flavonoid glycosides. Quercetin is also supplied as a nutritional supplement as it is beneficial for many variety of diseases. Quercetin has many beneficial effects on cardiovascular protection, anticancer, antitumor, antiulcer, antiallergy, antiviral, anti-inflammatory activity, anti-diabetic, gastroprotective effects, antihypertensive, immunomodulatory, anti-infective, etc. It can also protect from the environmental causes of free radicals like smoking. Example of free radical is cigarette tar which damages the erythrocyte membranes.

Pharmacological importance:

Inflammation is a biological response of body that it shows during the harmful exposure of human body against harmful pathogen. Inflammation helps us in the healing process. Quercetin also have a property to modulate inflammation. It exhibits COX the inflammatory enzyme cyclooxygenase and lipoxygenase. The pharmacological importance of quercetin is shown up as anti-inflammatory, cardiovascular disease prevention, neurodegenerative disorders, antibacterial and anti-viral activity, allergies, asthma, hay fever and hives, and performs pharmacokinetics.

Cancer and apoptosis:

Many surveys collectively showed result that due to the consumption of more fruits and vegetables can help to reduce the risk of cancer. Quercetin have many properties such as growth factor suppression, antiproliferative that in anticancer properties. Flavonoid rich foods and vegetables helps us in cognition functions and also protect neurons by enhancing existing neuronal function. Also, flavonoids have stimulating neuronal regeneration factor.Quercetin is known for apoptosis inductor and has anticarcinogenic properties. Quercetin decreases the growth of tumor in liver, colon, brain and other tissues. It also inhibits the spread of malignant cells. Quercetin can reduce the number and size of ideal with minimal adverse effect. Both invitro and in vivo cancer studies proves that quercetin inhibits chemicals that are responsible and have a beneficial effect against prostate cancer.

Pharmacokinetics:

If quercetin is consumed in toxic dose, quercetin causes hypertension, reduction in serum potassium, emesis and nephrotoxicity. The distribution of quercetin is 0.7-708mm and elimination half life of intravenous quercetin is 3.8-86 min. Its volume of distribution is 3.7l/m square.

Chemical effect of quercetin:

Quercetin is proved to exert beneficial effects on health, including protection against lung cancer, cardiovascular and osteoporosis diseases. The third leading cause of death in the USA is by chronic obstructive pulmonary disease (COPD) which is reduced by the intake of flavonoids. This is the disease caused by disorder of the lung parenchyma and airways. Therapies for COPD, unfortunately said to be partially effect and may be beneficial in its treatment. In pre-clinical studies it has been demonstrated a 4-fold increase in the plasma quercetin level that helped to decrease lung inflammation and prevented disease progression

CUCURBITA:

Cucurbita are commonly known as pumpkin and belongs to the family known as Cucurbitaceae. This comes under cucurbit sp. Wild Cucurbita are too bitter to humans and livestock for feeding larger mammals and resistant to the bitter plants than the smaller ones. Pumpkin is mostly found everywhere but is specially belongs to North America. It is an annual creeping or climbing plant. It has a hollow center and is spherical in shape. It Has shallow root system that is well grown from taproot. Cucurbita has many remarkable properties as it is rich in nutrients, antibacterial, antidiabetic, antifungal, anticancer, antioxidant, anti-inflammatory, anti-ulcer. Nutrient content of pumpkin contains protein, magnesium, manganese, amino acid, fatty acid and zinc. Cucurbita is important to health to its abundance of vitamins and mineral content that our body needs and hence it is to be added in a good quantity in our daily diet. Cucurbita has unisexual flower and hence comes under monoecious plant with female and male on the same plant. Pumpkin flower produces nectar. Seeds of pumpkin is in network structure. Also, seeds are agro- industrial waste and is usually of no use. Seeds are off-white, flat, oval and flat disc in shape. Cucurbita can be consumed in any form either it be roasted, snack or cooked due to its taste of crispiness. Cucurbita has some phytochemical constituents such as flavonoids, alkaloids, oleic acid, palmitic and linoleic acid. Pumpkin provides herbal remedies as they are less toxic with limited side effects. Pumpkin can also be used as to derive oil, seed butter, chocolate, tortilla, candy, cereal bar, vegetable salad, cookies, chips, soups and muffins. It has beneficial effect on blood glucose, immunity, liver, bladder, prostate gland, cholesterol and hence is consumed on a large scale in many counties including India.

II. LITERATURE REVIEW

1) Neel Ratnam et al (2007) said Cucurbita pepo is widely used as food and herbal remedies around the world. It contains several phyto-constituents belonging to the categories of alkaloids, flavonoids, and palmitic, oleic and linoleic acids. Many pharmacological studies have demonstrated hepatoprotection, inhibit benign prostatic hyperplasia, hypoglycemic agent, antioxidant, anticancer, antimicrobial, anti-inflammatory, anti-diabetic, and antiulcer activities supporting its traditional uses. This aims a comprehensive study of the chemical constituents, pharmacological and clinical uses.
2) Karanja J.K et al (2013) explained the nutritional potential of thirteen varieties of Cucurbita fruits collected from selected regions of Kenya were evaluated for their proximate values, mineral and β-carotene content. The results indicated that the moisture content was high and it varied significantly (p<0.05) among the groups. Ash values were significantly different at (p<0.05) and ranged from 0.63 to 1.67%, crude protein was low in all the pumpkin groups and varied from 0.93 to 2.09%. The pumpkins also varied significantly (p<0.05) in crude fibre content (0.47- 1.95) among the groups. Potassium and sodium were the most prevalent minerals potentially useful amounts of potassium (199-172.3 mg/100g), sodium (56.39-118.28 mg/100g) and other essential minerals. The lowest micro-elements were copper and zinc. The results indicated that there were no significant differences among the groups in the β-carotene content (p<0.05). The nutrient information presented in this report should stimulate the nutritionists and agricultural officers in Kenya to consider the question of recommending the pumpkin to be consumed by adults and children alike in Kenya, including pregnant women and others with higher than normal nutritional requirements Keywords Nutritional, Cucurbita, Proximate, Mineral composition
3) Dr.P.Mythili Md et al (2017) suggested pumpkin ( cucurbita maxima ) belongs to cucurbitaceae which include pumpkin, gourds, melons and squashes. They are characterized by their seed oil. Pumpkin plant has been grown since the earliest history of mankind1. ( Brucher – 1989 ). Pumpkin are grown around the world for variety of agricultural purposes, the seeds show anti helmintic, taenicide diuretic, cardio tonic, anti-inflammatory, anti - diabetic, antihyperlipidemic, anti – giardial activity CNS stimulant, melanogenesis inhibitory activity and immune modulatory action. Oil from seeds is nervine tonic. Cucurbita maxima seeds are good source of vitamins, minerals, calories and proteins. Multiple forms of phytase found in germinating cotyledons of cucurbita maxima seeds. Seeds are rich in tocopherols (δ tocopherols), triterpene and unsaturated fatty acids.
4) Xiang D et al (2004) proposed Cucurbita maxima, belongs to family Cucurbitaceae, is commonly known as pumpkin. Several literature reports suggest it to be antidiabetic, antihypertensive, anticancer, immunomodulators, antibacterial and antihyperlipidaemic. One important approach for the treatment of type 2 diabetes mellitus is by decreasing the postprandial hyperglycemia in effect. This is possible by inhibiting certain carbohydrate hydrolyzing enzyme like α-amylase and α-glucosidase. The objective of present study was to evaluate in-vitro antioxidant and α-amylase inhibitory activity of methanolic extract of C. maxima leaves. Methanolic extract of leaves of C. maxima showed strong antioxidant (DPPH scavenging) and α-amylase inhibition (IC50 125 μg/ml and IC50 2.1 mg/ml), respectively. These results suggest the possible use of pumpkin leaves in the management of diabetes mellitus.
5) Jacks TJ et al (1972) investigated quercetin and allopurinol significantly inhibited the TXNIP overexpression, activation of NLRP3 inflammasome, down‐regulation of PPARα and up‐regulation of sterol regulatory element binding protein‐1c (SREBP‐1c), SREBP‐2, fatty acid synthase and liver X receptor α, as well as elevation of ROS and IL‐1β in diabetic rat liver. These effects were confirmed in hepatocytes in vitro and it was further shown that TXNIP down‐regulation contributed to the suppression of NLRP3 inflammasome activation, inflammation and changes in PPARα and SREBPs.
6) Ang-Lee MK et al (2001) studied dietary plants and herbal preparations have been traditionally used as medicine in developing countries and obtained a resurgence of use in the United States and Europe. Research carried out in last few decades has validated several such claims of use of traditional medicine plants. Popularity of pumpkin in various systems of traditional medicine for several ailments (antidiabetic, antihypertensive, antitumor, immunomodulation, antibacterial, antihypercholesterolemia, intestinal antiparasitia, antiinflammation, antalgic) focused the investigators’ attention on this plant. Considerable evidence from several epidemiological studies concerning bioactivities leads have stimulated a number of animal model, cell culture studies and clinical trials designed to test this pharmacological actions. In addition, it was found that technologies such as germination and fermentation could reduce antinutritional materials and affect the pharmacological activities of pumpkin. This review will focus on the main medicinal properties and technologies of pumpkin, and point out areas for future research to further elucidate mechanisms whereby this compound may reduce disease risk.
7) Decker-Walters DS et al (2000) says a review of the activities of members of the Cucurbitaceae was carried out. Many of them are confirmed nutritious and therapeutical. Their global spread, diverse genera and phytochemical profile further confirm them as an attraction for the growth and survival of humanity. The need for alternative control measures to address resistance has heightened the passion for Cucurbitaceae in bioprospecting.
8) M.Manasa et al (2001) explained herbal medicines have gained global importance with bothmedicinal and economic implications these days. The scientific assessment has become a prerequisite for acceptance of health claims. Onion (Allium cepa ) a member of genus Allium, is the second most important horticulture crop all over the world. It is used as important source of phytoconstituents and food flavor. Allium cepa is one of the richest source of flavonoids and organosulphur compounds.Wide spectrum of biological activities makes A.cepa a potential therapeutic agent.
9) Cai X et al (2013) suggested quercetin (QC) is a typical plant flavonoid, possesses diverse pharmacologic effects including antiinflammatory, antioxidant, anti-cancer, anti-anaphylaxis effects and against aging. However, the application of QC in pharmaceutical field is limited due to its poor solubility, low bioavailability, poor permeability and instability. To improve the bioavailability of QC, numerous approaches have been undertaken, involving the use of promising drug delivery systems such as inclusion complexes, liposomes, nanoparticles or micelles, which appear to provide higher solubility and bioavailability. Enhanced bioavailability of QC in the near future is likely to bring this product to the forefront of therapeutic agents for treatment of human disease.
10) Ravi Kant Upadhyay et al (2016) investigated cucurbita contain phenolics and flavonoids that have potential anti-inflammatory, anti-cholesterol, anticancer, and antioxidant properties. Onions contain 89% water, 1.5% protein, and vitamins B1, B2, and C, along with potassium and selenium. It also contains polysaccharides such as fructosans, saccharose, peptides, flavonoids (mostly quercetin), and essential oil. Onion contains numerous sulfur compounds including thiosulfinates and thiosulfonates; cepaenes; S-oxides; S, S-dioxides; mono, di, and tri-sulfides; and sulfoxides. Onion is highly nutritional and its dietary use improves digestion and mental health and lower down toxigenicity of oils. Onion has potential in treating cardiovascular disease, hyperglycemia, and stomach cancer. Onion contains an important antioxidative, i.e., quercetin that is derived from Allium cepa on aldehyde oxidase low-density lipoprotein which reduces hepatocytes apoptosis in streptozotocin-induced diabetic rat. Onion has great ethnomedicinal importance as native remedies used against diabetes, and related complications are from onion. A. cepa red and white varieties showed antimicrobial and antioxidant activities. These are used in traditional Indian spices and are of great health significance. These are curative for implications from and for food cultures for cardiovascular disease and provide longevity
11) Manasa Machavarapu et al (2013) noted allium cepa is the most common and richest natural source of flavonoids. In this study the extraction of flavonoids quercetin and rutin and the total phenolic compounds were carried out. Optimization of physico-chemical parameters namely effects of different solvents, different solvent percentages, soaking time, different amounts of sample (Skin of Allium cepa) and different pH were studied. For the extraction of quercetin, the optimum results were 80% (v/v), 1d, 1% (w/v) and pH 5.5 respectively. The highest quercetin and rutin concentration for optimized conditions were 185.309 µg/L and 124.69 µg/L respectively and it was found to be 19.412 mg/L for total phenolic content
12) Bhimanagouda S. Patil et al (1995) studied the aglycone, or free quercetin, and total quercetin content of 75 cultivars and selections was analyzed by reverse-phase high-performance liquid chromatography. Quercetin glycosides were hydrolyzed into aglycones. Total quercetin content in yellow, pink, and red onions varied from 54 to 286 mg·kg ⁻¹ fresh weight in different onion entries grown during 1992. White onions contained trace amounts of total quercetin. Free quercetin content in all the onions was low (< 0.4 mg·kg ⁻¹ ) except in `20272-G' (12.5 mg·kg ⁻¹ fresh weight). Bulbs stored at 5, 24, and 30C and controlled atmosphere (CA) for 0,1,2,3,4, and 5 months showed a most marked change in total quercetin content at 24C compared to other treatments, with a rise in mid-storage followed by a drop. Storage at 5 and 30C also demonstrated a similar change. However, total quercetin content did not vary significantly in bulbs stored at CA for 5 months. We conclude that genetic and storage factors affect quercetin content on onions
13) Bansode Ashwini Sopan et al (2014) proposed as Antioxidant potential of many plants is mainly due to phenolic components such as phenolic acid, phenolic diterpense therefore total phenolic content and antioxidant potential of cucurbita maxima. (Pumpkin) powder was determined respectively by using Folin-Ciocalteu reagent and diphenyl-picryl-hydrazyl (DPPH) assay. In this method the antioxidants present in the plant extracts reacted with DPPH, which is a stable free radical and converted it to 1,1-diphenyl-1,2-picryl, hydrazine. Total phenolic content of pumpkin was determined using Folin-Ciocalteu reagent and gallic acid as standard, amount of phenolic compound was found to be 6.5 mg/gm powder of the drug. The high fiber content of pumpkin helps to aid proper digestion and in traditional medicine it is used as an anthelmintic, taenicide, diuretics and in the treatment of hemorrhoids so we compared the anti oxidant potential of pumpkin powder with marketed preparation (Pilex tablet). The half maximal inhibitory concentration (IC50) is a measure of the effectiveness of a compound in inhibiting biological or biochemical function. IC50 value of pilex tablet = 8.53 µg/ml & IC50 value of pumpkin powder =9.12 µg/ml.
14) Md. Irshad et al (2014) noted pumpkin, as a dietary plant, has been used in traditional medicine around the world. In addition, during the last decade, antidiabetic, antihypertensive, antitumor, intestinal antiparasitic, antibacterial, anti hypercholesterolemia, anti-inflammatory, immunomodulatory and analgic effects of pumpkin has been reported. The aim of the present study was to determine the effects of different extracts of Cucurbita pepo L. on the immune responses. Methanolic, chloroform and ethylacetate extracts of C. pepo fruits was obtained using percolation method. Mice were used to study the effects of C. pepo extracts on the acquired immunity. Sheep red blood cell (SRBC) was injected (S.C., 1×108 cells/ml, 20 μl) and 5 days later, methanolic, chloroform and ethylacetate extracts of C. pepo at diiferent doses (10, 100 and 500 mg/kg), betamethasone and levamisol at equal doses (4 mg/kg) as positive controls and normal saline as a negative control were given i.p. After 1 h SRBC was injected to the footpad (S.C., 1×108 cells/ml, 20 ml) and the footpad swelling was measured up to 72 h. To investigate the effects of C. pepo on the innate immunity the same procedure was used, but animals received only one injection of SRBC 1 h after i.p. injection of test compounds. Our findings showed that SRBC induced an increase in the paw swelling with maximum response at 6-8 h. Betamethasone inhibited the paw swelling in both models. In both innate and acquired immunity models, methanolic, chloroform and ethylacetate extracts of C. pepo fruits significantly reduced the paw swelling dose dependently. The data suggest that the pumpkin extracts may have immunomodulatory effects.

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