Extraction Of Natural Dyes And Its Applications

Artificial dyes are widely used in everyday items, with side-effects not known to the people. They are toxic, harmful and carcinogenic. There is a worldwide increase in demand of natural dyes due to its therapeutic properties and no known side-effects. The current research aimed at extracting dyes of different colors from natural plant sources, carrying out its characterization and its applications. The dyes were tested for their antimicrobial ability and were found to be inhibitory to common organisms like S. typhi, C. diphtheriae, S. aureus, etc. Phytochemical tests were performed and they confirmed presence of several important metabolites like Phenols, Tannins, Terpenoids, and many more. The dyes were tested for anti-oxidant activity by FRAP assay and antioxidant levels ranging from 0. 62 to 2. 00 were obtained. Preparation of cosmetics like lip balms, was done and dyes were also tested as natural food coloring agents. The results indicated that the dyes caused inhibition of wide range of bacteria and proved to be good sources of antioxidants. They departed a natural color to food items and hence can replace artificial food colorings. Thus the use of natural dyes should be promoted as they are cheap and biodegradable.

Introduction

Dye is a natural or synthetic substance used to add a color to or change the color of something. It is a substance used to impart color to textiles, paper, leather, and other materials such that the coloring is not readily altered by washing, heat, light, or other factors to which the material is likely to be exposed. Artificial dyes have been used widely since its inception. They are used in everyday items like clothes, food products, cosmetics, plastics, etc. The harmful effects of these dyes, however, go unnoticed by the people. In the recent years, due to the scientific development, the adverse effects of these dyes are being discovered. They have been found to cause or contribute to various allergic reactions, toxicity, mental disorders like ADHD, tumor formations, etc. Recent studies show that seven of the artificial dyes contributed to cancer in lab animals, including brain and testicular tumors, colon cancer, and mutations. These dyes not only pose a risk to the biotic components, but are also responsible for degradation of abiotic components like soil and water. The contamination of soils and waters by dye-containing effluents is of environmental concern. Water pollution can also result from manufacturing synthetic dyes when untreated dye effluent is dumped directly on bodies of water.

Dyes derived from natural materials such as plant leaves, roots, bark, insect secretions, and minerals were the only dyes available to mankind for the coloring of textiles and other products until the discovery of the first synthetic dye in 1856. Rapid research strides in synthetic chemistry, supported by the industrialization led to the development of synthetic alternatives to popular natural dyes and gradually pushed the natural dyes into oblivion. However, environmental and health issues have once again revived interest in production and use of natural dyes. Natural dyes are considered eco-friendly as these are renewable and bio-degradable; are skin friendly and may also provide health benefits to the wearer. Natural dyes are harmonizing and soothing to the human eye. Some of its constituents are anti-allergens, hence prove safe for skin contact and are mostly non-hazardous to human health.

Beetroot (Beta vulgaris) is high in immune-boosting vitamins and minerals, it is also good for the bones, liver, kidneys, it reduces risk of heart disease and improves endurance and blood pressure. The petals of Rose (Rosa) are beneficial to the skin and cure dry and patchy skin; they are also used in health tonics. Carrot (Daucus carota) is a rich source of vitamin A, C, K and B8 as well as pantothenic acid, folate, potassium, iron, copper, and manganese. It is also a good source of antioxidants. Marigold (Tagetes) has been used to make healing herbal ointments; it is a rich source of flavonoids and anti-oxidants. Pomegranate peels (Punica granatum) are effective detoxifying agents, combat tool against heart diseases, in prevention of skin cancer, for coughs and sore throats, etc.

Henna (Lawsonia inermis) has the ability to relieve headaches, detoxification, skin protection, boost hair health, etc. Spinach (Spinacia oleracea) is high in protein, zinc, fibre, vitamins, etc; its health benefits include skin care, eyesight, regulated blood pressure etc. It is a good source of water soluble vitamins like thiamine, riboflavin, niacin and ascorbic acid. It contains enough amount of vitamin C to prevent and cure scurvy. Peppermint has significant antimicrobial and antiviral activities, strong antioxidant and antitumor actions, and exhibits some antiallergenic potential.

Bioactive compounds in plants are known to be responsible for various bioactivities of the plant. Phytochemical assays are often considered to be the first step for characterization of a plant sample depending on the presence of the various metabolites. A positive response of the plant sample for the presence of Terpenoids, Flavonoids and Glycosides indicates it potential use for defence against micro-organisms, since most of these metabolites impart the ability to inhibit the growth of organisms. Phenolic compounds possess properties such as antiaging, anticancer, anti-inflammatory, antiatherosclerosis, cardiovascular protection. Coumarins have bacteriostatic and anti-tumor activity and help in inflammatory responses and act as anti-coagulants. (16) Quinones provide protection against reactive oxygen species (ROS) and have a diverse role in medicine, including anti-cancer agents and anti-aging and arteriosclerosis.

Free radicals are highly reactive and capable of damaging biomolecules. They are associated with certain inflammatory diseases like rheumatoid arthritis and respiratory disorders. Various studies have demonstrated the adverse effects of antioxidant supplements such as high risk of cancer, increased mortality, etc. Several nutrients in food and/or plants like vitamin C and E, beta-carotene, phenols, flavonoids, etc. are commonly studied dietary antioxidants.

Antimicrobial agents are essentially important in reducing the global burden of infectious diseases. Alternative antimicrobial strategies are urgently needed, and thus this situation has led to a re-evaluation of the therapeutic use of ancient remedies, such as plants and plant-based products. The use of plant extracts and phytochemicals, both with known antimicrobial properties, can be of great significance in therapeutic treatments.

With regards to the increased scope of natural dyes, its study, production and usage; the present research is designed for the extraction of dyes from natural plant sources, its characterization and studying its applications.

Materials and Methods

Selection and collection of plant materials-8 different plant sources were chosen for extraction of dyes of 4 different colors. Beetroot and Rose for Red, Pomegranate peel and Henna for Brown, Spinach and Peppermint for Green, Carrot and Marigold for Orange. Extraction of natural dyes-Dyes were extracted using both water and organic solvents. Water-based dyes were prepared by blanching, grinding and filtering the plant material using distilled water. These were further lyophilized for concentration. Organic solvents were used for extraction of the color-imparting phytochemical. Methanol was used for Beetroot, Rose, Marigold, Henna. Ethanol was used for Pom peel, while Petroleum ether was used for Carrot. Spinach and Peppermint were extracted using acetone. Qualitative phytochemical analysis-The water-based dyes were used for phytochemical screening. Phytochemicals like Proetins, Glycosides, Terpenoids, Quinones, Coumarin and Saponins were analysed. Phenols. Tannins, Carbohydrates, Flavonoids were also analysed.

Antimicrobial ability – the ability of the dyes to inhibit bacteria was tested using the Agar dilution method. 1g of concentrated dye mixed with 2ml 20% DMSO was incorporated into molten nutrient agar butts which were poured onto plates. 6 common pathogenic organisms, namely, E. coli, S. typhi, P. aeroginosa, S. aureus, S. faecalis and C. diphtheriae. were spot inoculated on the plates and incubated to check for results.

Anti-oxidant activity – the ability of the dyes to scavenge free oxygen radicals was determined using the FRAP (Ferric Reducing Ability of Plasma) assay.

Applications of the dyes

1. Use as Food coloring agents – Noodles were colored by boiling them in water containing the dye for 5-10 minutes, while, doughs of white flour were prepared using dye mixed with water.
2. Cosmetic production (Lip-balm) – Natural ingredients like aloe gel, almond oil, beeswax, rice powder and cornstarch were mixed well. 5-6 drops of the dye was added and mixed until a good consistency was obtained. The prepared lip-balms were then stored at 4°C and RT.

Results

The extracted dyes showed a good color imparting ability and were of the desirable concentration. Phytochemical analysis revealed the presence of important metabolites like Proteins, Carbohydrates, Phenols, Tannins, Terpenoids, Glycosides. These are major metabolites present which are responsible for several additive properties of the plants. In the table given below, ‘+’ and ‘-’ indicates presence and absence of that phytochemical respectively. The dyes were checked for their antimicrobial ability by the Agar Dilution method. Organisms like S. aureus, P. aeroginosa, were inhibited by all the dyes. While, S. typhi, E. coli, C. diphtheriae and S. faecalis were inhibited by most of the dyes. The following is a tabular representation of the antimicrobial ability of the dyes as observed, upon comparing with the controls.

The anti-oxidant activity of the dyes was tested using FRAP assay and ascorbic acid as standard. Highest anti-oxidant activity was shown by Beetroot dye, followed by Henna, Rose, Carrot dye. Other dyes also showed a good anti-oxidant activity.

The noodles and whilte flour dough that were prepared using the dyes retained a good color and no considerable change in taste was imparted to the food products. The prepared lip-balms were found to be stable upon storage at RT and 4°C for upto 4 weeks. They had a good consistency and gave a vibrant color, Also, since the dyes were prepared using all natural ingredients, they gave a refreshing and soothing feel to the skin.

Discussion

The dyes extracted from natural sources are rich sources of important bioactive compounds. Plants like Henna, Beetroot, Carrot, Spinach, etc are known to have therapeutic properties. The extracted dyes showed prominent and lucid colors. Upon testing for presence of various phytochemicals, the dyes showed presence of important metabolites like Proteins, Tannins, Carbohydrates, Phenols, Glycosides and many more. These phytochemicals are known to have therapeutic properties like protecting the cellular systems from oxidative damage, modulating immune system, preventing cancer and lowering the risk of chronic diseases like cardiovascular diseases, etc. The antimicrobial ability of the dyes was tested using the agar dilution method and the dyes showed inhibition to common pathogenic organisms like Staphylococcus aureus, which is a common skin infection causing bacterium; Pseudomonas aeroginosa, which is a frequent cause of various nosocomial infections. Organisms like Streptococcus faecalis, Salmonella typhi, Pseudomonas aeroginosa, and Corynebacterium diphtheria, that are the causatives of various afflictions were also inhibited by most of the dyes. Being extracted from natural plant sources, the dyes were found to be rich in anti-oxidants using the reduction based analysis – FRAP (Ferric Reducing Ability of the Plasma) assay. The highest anti-oxidant level found spectrophotometrically was that in Beetroot Red dye (FRAP value-2. 00). The other dyes obtained were also found to have a good anti-oxidant activity. The food products colored using the dyes showed no change in taste and retained a vibrant color. Also, the cosmetic (lip-balms) prepared using the dyes were found to be stable upon storage and gave a refreshing and soothing feel to the skin.

Natural dyes are on the rise ever since the harmful effects of artificial dyes have begun to uncover. Presence of important metabolites and high anti-oxidant activity are an indication of the beneficial purposes these dyes will serve in food industries. The ability to inhibit commonly pathogenic organisms is proof of the medicinal and therapeutic properties of these natural dyes. Natural dyes can, thus, replace artificial dyes in the near future.