Application of the Dye from Caesalpinia Sappan Essay Example
Application of the Dye from Caesalpinia Sappan Essay Example

Application of the Dye from Caesalpinia Sappan Essay Example

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The development of dyes to stain microorganisms was a significant advance in microbiology. Stains serve purposes such as it differentiate microorganisms from their surrounding environment and it allows detailed observation of microbial structures at high magnification. (http://insist. Back. Wise. Deed/, 0 2006-2013 Microbiology Laboratories) Sexagenarians (2005) defined staining as the method of artificially producing color in microorganisms to allow for the visualization under the microscope.

He further stated that stains are used not only to make the organisms kibbles but also to understand their structure and chemical nature. Gram staining is a technique commonly used in the laboratory. It is used to differentiate two large groups of bacteria based on their different cell wall constituents. It distinguishes between Gram positive and Gram negative groups by coloring these cells red or violet.

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When bacteria are stained by crystal violet, Gram positive bacteria stain violet due to the presence of a thick layer of pedagogical in their cell walls which retains the crystal violet while Gram negative bacteria stain red which is attributed to a winner pedagogical wall, which does not retain the crystal violet during the discoloring process (http://sere. Carleton. Deed/18370, 2012). The Escalating span is characterized as small shrubby tree which can grow up to 4-mm tall. The roots are fibrous and wiry, lacking nodules and dark colored.

Under natural conditions, Escalating span grows mostly in hilly areas with clayey soil and calcareous rocks at low and medium altitudes. (http://www. Glommed. Com/, 2010-2011) The wood of Escalating span is the wood-dye that is known to have been used at large, international scale throughout the world history. It remained a major source of red dye

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up to the end of the 19th century. It is still used for dyeing textiles, but only on a smaller scale by craftsmen and artists. The main dye component in the heartwood of Escalating span is brazilin.

It is weakly colored product which easily oxidized to give the deep red pigment brazilin Manson, 2005). By manipulating the pH of the dye bath by the addition of wood ash water or vinegar, the dyer's produce everything from deep egg plant to lavender and maroon to ox blood red color (Bedlam, 2004). Thus, this study will determine the effectively of the dye from Escalating span for retain bacterial species. Statement of the Problem The study is entitled "Application of the dye from Escalating span (Sapping) for bacterial staining".

The main objective of the study is to assess the dye from Escalating span as an affordable and effective staining agent for microscopic observation. Specifically, this study seeks answers to the following questions: What is the color reaction of Escalating span as a stain to Gram-positive and Gram-negative Etcetera? 2 want Is ten detectably AT ten aye sailplane span (Sapping) as a bacterial stain in comparison with Gram stain in terms of: . 1. Color, 3. 2. Contrast; and, 3. 3. Sharpness? 3. Is there a significant difference in the effect of the dye Escalating span as a bacterial stain in terms of: 4. . Color, 4. 5. Contrast; and, 4. 6. Sharpness? 4. Is Escalating span (Sapping) effective in staining the bacterial cells under the microscope? Hypothesis There is no significant difference in the effectively of the dye Escalating span as bacterial stain in terms of color, contrast,

and sharpness. Significance of the study The outcome of the study will benefit the following. To the School Laboratories. The dyes used in laboratories nowadays are expensive. In a school setting, the laboratory facilities lack the capacity to provide stains to do comprehensive experiments.

By assessing the dye Escalating span, this study will be deemed significant because it will provide school laboratories or institutions with a cheap, conventional stain. To the Medical Laboratory Students. This study will be deemed significant as it will help the medical laboratory students in having additional information pertaining to natural dyes that can be used as an alternative stain. To the Environment. This study ill be deemed significant because the dye that was studied will help in reducing water contamination because it is naturally made thus it is also non-toxic to the environment.

The dyes being used in the laboratory brings great hazard to environment due to improper waste disposal of used and expired chemically made stain. To the Future Researchers. This study will be deemed significant to the future researchers as it will guide them as a reference pertaining to researches about indigenous ways to stain microorganism. Scope and Limitations of the Study This study is only limited on the investigation of the dye from Escalating span s a bacterial stain and the said dye will be only tested on certain species of bacteria. Gram-positive bacteria are those that are stained dark blue or violet by Gram staining.

This is in contrast to Gram-negative bacteria, which cannot retain the crystal violet stain, instead taking up the countersink (seafaring or fuchsias) and appearing red or pink. Gram-positive organisms are able

to retain the crystal violet stain because of the high amount of pedagogical in the cell wall. The respondents will only be limited to the College of Medical Technology faculty members and students f Our Lady of Fatima University-Antipodal City; Five (5) microbiology faculty members and 30 students currently taking the bacteriology course will be asked to answer the corresponding questionnaire given by the researchers.

This research will start November 27, 2012 and will end at October 7, 2013. Definition of Terms The following important terms are defined below for a clear understanding of the study: Brazilin. It is a natural dye which is extracted from the heartwood of Escalating span. Sailplane span. It Is a small sundry tree wanly can grow up to 4-mm roots are fibrous and wiry, lacking nodules and dark colored. Dye. It is described as a colored substance that has an affinity to the substrate to which it is being applied. Escherichia coli.

It is one of several types of bacteria that normally inhabit the intestine of humans and animals. Gram negative. This is characteristic of bacteria that have a cell wall composed of a thin layer of a particular substance called pedagogical. Gram positive. This is characteristic of bacteria that have a cell wall composed of a thick layer of a particular substance called epidemiological. Gram stain. A microbiological procedure that categorizes bacteria based on the physical ND chemical structure of their outer surface. Span wood.

A dyed wood yielded by Escalating span. Stain. It is used to add color to tissues and microbes to make them optically distinct (Organisms and Beggarly, 1993). Staphylococcus erasure. A species of Staphylococcus

that produces a golden pigment with some color variations and is commonly found on the skin or nose of healthy people. Synthetic. Produced by synthesis, especially not of natural origin. The review of related literature and studies will provide useful insights on the conduct of the present study.

The concepts and ideas generated from the review will be used as guide to direct the course of the present study particularly on the ability of Escalating span to stain bacterial species. These readings will guide the researchers in the formulation of their questionnaire and utilization of appropriate methodology and procedure. According to Wiley in 2008, even though microorganism can be directly examined with the light microscope, in order for them to increase visibility and accentuate its specific morphological features and preserve them for future study they must be fixed and stained first.

Black (2005) defined stain, or dye, as a molecule that can bind to a cellular structure and give it color. She stated that staining techniques make the microorganisms stand out against their backgrounds. They are also used to help investigators group major categories of microorganisms, examine the structural and chemical differences in cellular structures, and look at the parts of the cell. Hans Christian Gram developed the Gram staining; a century old method that still remains the most universal diagnostic staining technique for bacteria.

It divides bacteria into two classes based upon the color reaction of the cells. Gram-positive bacteria stains purple while gram-negative bacteria stain red (Cowan, 2006). The staining procedure differentiates organisms of the domain Bacteria according to cell wall structure. Gram-positive cells have a thick pedagogical layer and stain blue to

purple. Gram-negative cells have a thin pedagogical layer and stain red to pink (http://www. Microbiological. Org). Various theories have been proposed to explain why some bacteria retain the dye and some don't.

The thickness of Gram positive cell wall and presence of more lipids in gram negative cell walls have been more acceptable reasons for Gram stains reactions http://www. Micro. Com). Gram-positive microorganism has a thicker pedagogical layer that can protect and surround the entire cell, which interact with negative changer Doctoral components Ana stalls ten Etcetera cells purple (http//:www. Subsistence. Com). While gram-negative microorganism contains thin pedagogical layer which will be decolonize by alcohol and will be produce a red color due to the countersink seafaring (Toronto, 2011).

Escalating span Escalating span (Sapping) is described as a small tree that reaches a height of 8 to 10 m high and a diameter of 30 to 40 CM at maturity. Its trunk and branches have spines that distinguish it from other fuel wood species. The leaves are compound with a pair of spines underneath the midrib. It has yellow flowers. The fruits are oblong, hard, and its shiny pods contain 3 to 4 seeds (Florid, 2002). The generic name is after A. Cleansing, 1519-1603, Italian physician and botanist (Raw et al. 2009). The common name of Escalating span in the Philippines is Span in regions of 'locos Sure, La Union, Bataan and Racial.

It is called Kabuki's in the provinces of Samaras Island, Negroes, Indoor, Guenon Province, Bucolic Region, Sambaing and Basilar (Florid, 2002). Japans is found locally abundant throughout the Philippines at low and medium altitudes in dry thickets. It is extensively cultivated in

Lillo, Capri, Klan, Antique, Negroes Occidental, and Samaras Island. It is also abundant in Bucolic Region (Florid, 2002). It can be seen in low elevation and stony areas. It propagates thru seeds (Baton, Sir, 2009). The plant has been used as a traditional medicine.

It possesses medicinal abilities as an anti-bacterial and for its anti-coagulant properties. A detection of wood is said to be very useful in curing dysentery and diarrhea (http://wonderers. Rug). In the study conducted by Bedlam in 2004, the detection of the wood is used as a powerful mongoose in India, Brazil, China and several other countries. It is used as an anti-anemic and anti-malarial in Philippines and Malaysia, respectively. Brazilin is the main dye component of the heartwood of Escalating span. It is also present in Brazil wood (Escalating Chianti Lam).

It is a weakly colored product which is easily oxidized to give Brazilin, a deep red pigment Manson, 2005). Staphylococcus erasure. The genus Staphylococci usually occur in grapevine clusters. Staphylococcus erasure (abbreviated as S. Urges) is the most important staphylococcal species. It is a gram positive microorganism which is characterized by spherical cell and has yellow pigmented colonies thus the name erasure which is Latin for gold (Tractor, 2011). It is a facultative anaerobe and opportunistic pathogen that is a major cause of skin, soft tissues, respiratory, bone, joint, and endeavourers infections.

Staphylococcus erasure is usually involve in most pimples, boils, carbuncles, and styles. It is also one of the four most common causes of monoclonal infection causing post-surgical wound infections (Engineering, 2007). Escherichia coli The American Heritage Medical Dictionary defines the genus Escherichia as an aerobic, gram-negative, rod-shaped bacteria

found in nature. Escherichia coli (E. Coli) is a member of the normal intestinal flora (Brooks, 2007). According to the Center for Disease Control and Prevention (CDC), some E. Olio is pathogenic which meaner that they can cause illness either diarrhea or an illness outside of the intestinal tract. CDC also stated that the types of E. Coli that can cause diarrhea can be transmitted through contaminated water or food or through contact with infectious animals or errors. Related Studies According to the study made by Braided et al in 2010, most dyes that are used is chemical synthesized. I nee are expensive Ana can De nauseous to unman Neal their study, they utilized four Nigerian plant extracts to determine their staining potentials.

They used extracts from Hibiscus seafaring, Bias Orwellian, Lawson mannerism, and Pterosaurs sun. They observed that Pterosaurs sun (uh) exhibited the best staining result. They further stated that the result obtained indicate that extracts with good staining ability have the potential use in diagnostic bacteriology. In the study conducted by 'human et al in 2012, they stated that over 2000 dyes are synthesized from various parts of plants, of which only about 1 50 have been commercially exploited. Among such plants is the H. Beautify Lain which is a very important dye-yielding annual/perennial plant species. Apart from the use of the extract in biological staining, the extracts can also be used as a therapeutic, a laxative, an interventions, and a cholesterol lowering medicine. In their study, they investigated the potential of methanol extracts from H. Subsidiary Lain as a staining agent for fungal species. They concluded that the Methanol extracts from H.

Subsidiary could be a good replacement for Electroplate in cotton blue stain because this plant species is available at affordable low market price.

They suggested that there is also a need to investigate the potential use of methanol extracts in the staining of other microorganisms aside from fungal species. Another study was made utilizing H. Subsidiary Lain (Rosella calyx) as fungal stain. Babushka (2012) attempted to use the aqueous and ethanol extracts of the rosella calyx in mycologist staining. Calyx is a very important dye-yielding organ of Rosella H. Subsidiary Lain. He concluded that Red calyx from rosella plants produced an extraordinary red color which had the affinity for mycologist staining.

In the study conducted by Lie et al in 2012, extraction of the methanol or ethanol dye of (broadsword/sapping) Escalating span L. After crystallization has a reddish brown color. If dissolved in hot water, the crystal will exhibit a deep red color. Brazilin has been isolated as the major component of the extract. However, brazilin is easily oxidized by air and light to produce brazilin. The oxidized dyestuff component has n intense red color which has been used as a traditional food coolant in several Asian countries. The traditional practice to extract the dye of the broadsword is simply by boiling the wood.

They further stated that besides the function of the ethanol or methanol dye extract of the broadsword as a natural food and beverage coolant, it has also been investigated that it can be used as an anti-inflammation, anti-oxidant, and anti-microbial. They also added that in ancient centuries, the dye has been known to be used in historical painting and textiles. Justification

of the Study In the study of W. Braided, C. Icebound, R. N. Announcing and L. C. Incredible in 2010 dyestuffs from four (4) Nigerian plants were extracted and their staining ability on some species of bacteria and McCollum were determined.

The study shows results the dyestuffs were extracted using water and organic solvents. Acetic acid treatment of the extracts increased the ability of the extract to stain both bacteria and McCollum. All these researches are related to the present study for they all pertain to the dye, stain, microorganism, indigenous plants and Escalating span. Although previous tidies manifest viability of the application of natural dyes, stain, microorganism, Indigenous plants Ana sailplane span In relation to some variables, teeny lean towards the necessity of the instrument.

They differ, however, from the present study which deals specifically on the application of the dye from Escalating span (Sapping) for bacterial staining.This chapter presents the research method used, the respondents of the study, instruments, data gathering procedures, and statistical treatment of data. Research Method This study will employ the quasi-experimental method research design. This method will include the use of variables: The extracted dye from Escalating span as the independent variable, color, contrast, and sharpness as the dependent variable, and gram stain as the control.

In terms of the clarity in viewing the specimen stained with the dye from Escalating span (Span), it will used the descriptive type of research to compare with the conventional staining method to answer the question tool given. The experimentation and biological test of this research will be conducted at Our Lady of Fatima University (Antipodal Campus), Medical Laboratory Science Laboratory. The experiment

will be conducted from April to May 2013. Instrument/Materials Test Organism The heartwood/bark of Escalating span will be used in the study.

The span plant will be obtained at FEZ Fruit Tree Seedling Nursery. Preparation of raw materials The heartwood/bark of Escalating span will be sun dried and chopped into pieces and further dried in a laboratory oven for 4 days at ICC. The chips will be broken into smaller pieces using laboratory mortar and pestles and will be further reduced to powder by grinding. The powder will be sterilized in a hot air oven at ICC for 1 hour (Braided et al, 2011). Extraction of Pigment The plant materials will be boiled in distilled water followed by soaking the plant materials in organic solvents.

Ten grams of the sapping will be soaked 100 ml of the solvents (80% ethanol and absolute ethanol, methanol and chloroform) and aqueous solutions (hot and cold) in 250 ml Erlenmeyer flask. The mouth of the flask will be stopped with cotton wool wrapped with aluminum foil and allowed to stand in the dark for 48 hours at ambient temperature. The quantities of the plant materials will be boiled in 100 ml distilled water for 5 minutes and allowed to cool before reification (Braided et al, 2011).

Purification of extracts (dyestuff Filtration I en content AT ten Atlas will De Telltale. Nine rescue Trot ten contractor extracts wall be put back into the Erlenmeyer flask and 20 ml of distilled water will be added into it and shaken gently until a strong color becomes visible and the contents will be filtered again (Braided et al, 2011). Evaporation of solvents The

organic solvent in the extract will be evaporated by passing warm air over the mouth of the flask. The extract will be further dried in the hot air oven at ICC for la (Braided et al, 2011).

Reconstitution of dyestuffs The dried extract will be reconstituted by adding 20 ml of distilled water. The reconstituted extracts will be transferred into a screw capped bottle, corked tightly, labeled and preserved in the refrigerator prior to use (Braided et al, 2011). Preparation of staining reagents (dyestuff from plant extracts Two millimeters of the reconstituted dyestuffs will be transferred into a test tube and 0. 2 ml glacial acetic acid will be added and the whole contents will be shaken to mix then it will be corked and labeled appropriately (Braided et al, 2011). Microorganisms

Pure cultures of the bacterial species (Staphylococcus erasure, Escherichia coli) will be cultured at Our Lady of Fatima University (Antipodal Campus), Medical Laboratory Science Laboratory. The pure cultures will be screened to confirm their identity using the standard methods. Gram Stain A heat-fixed bacterial smear on a glass slide will be treated with crystal violet, the basic dye. The smear will be then covered with Gram's Iodine solution. After a rinse with water and decentralization with acetone, the preparation will be thoroughly washed in water and countersigned with seafaring.

The stained preparation will be inside with water and air dried (2004). Staining of tested microorganisms Staining of bacteria A small piece of bacterial culture will be removed and transferred on a cleaned grease free slide using an inoculating needle. A drop of the extracted dye from Escalating span will be placed and heat

fixed. Gram staining reagents (crystal violet, logo's iodine, acetone and seafaring) will be used. Control slides will also be prepared and stained by Gram's Method (Braided et al, 2011). The slide will be covered with cover slip carefully to avoid air bubbles.

The slide will be examined first under owe power lox, then in high power XIX, and then oil immersion XIX of the microscope objectives respectively (Babushka, 2012). Respondents of the Study There will be two (2) groups of respondents for this study: composed of 20 third year students taking up Bacteriology course and five (5) Microbiology faculty instructors to assess the effectiveness of Escalating span as a stain in terms of specimen color, contrast, and sharpness. Instrument uses The researchers will use a researcher-constructed questionnaire.

Inputs from books, Journals, and other pertinent sources will be incorporated to Justly assess the rabbles under study. The instrument will be submitted by the researchers to their adviser and selected panel member for checking and validation. Upon approval, the final copy will be made ready for distribution to the target respondents. Data Gathering procedure Prior to the administration of the researchers' instrument, the researchers will sought the permission of the Dean of the College of Medical Laboratory Science, Dean Jose Surely Novo, ARM, PhD, and the College adviser, Mrs...

Dianne Capistrano- Mendoza, ARM, MPH, to allow them to involve their faculty members and students in the study. If the permission will be granted, they will administer and retrieve the questionnaires from the target respondents. After the retrieval of all the accomplished questionnaires, the researchers will then proceed to encoding, summarizing, and tabulating of the data for statistical

analysis and interpretation. Statistical Treatment of Data The researcher will ask for the assistance of an expert statistician for the computation of data to be utilized in the analysis and interpretations thereof.

The Chi-square statistics will be utilized for the statistical analysis of data to be gathered. Chi-square test that will be used in this study will determine the difference of the result of the experiment.

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