Ethanol Concertration Effects on Cell Membrane for Spectroscopy

Question: Discuss about theEthanol Concertration Effects on Cell Membrane for Spectroscopy. Answer: Objective Of The Experiment The objective of this experiment is to study the effect of different ethanol concentrations on cell membrane integrity by using visible spectroscopy Methods Requirements The requirements are, three test tubes, test-tube rack, cuvettes, beetroot (Beta vulgaris), spectrophotometer, detergent SDS 10%, small dowel rods, room temperature water, 3 replica, corkborer with 4mm inside diameter,6 replicas, 70% ethanol, prepared 50% ethanol +10 ml water, prepared 15ml of 70%ethanol+20ml water, distilled water. Procedure Cut B vulgaris into four equal pieces with the same diameter and length using a cork borer in a size as to fit in a cuvette. Rinse each piece of B. vulgaris with tap water to remove the excess red dye (pigment betalain) that resulted from cutting, number the test tubes from 1-4, mix the contents in the test tubes after placing the contents in each cuvettes and name them for ten minutes, warm the spectrophotometer for at least 20 minutes, place the cuvette with room temperature water(used as the control experiment) in to the spectrophotometer with triangle on the cuvette facing directly 10ml in front of the instrument and press the 0 ABS 100% and then remove it from the instrument, repeat this for the other cuvette A,B and C and record the values obtained, the values obtained are at an interval of 15 minutes, and values are recorded thrice for precision to be compared to the replica. Results The results of absorbance are obtained from the spectrophotometer with the time interval of 15 minutes as shown in the table below; Cuvette Concentration (%) Time in minutes 0.00 15 minutes 30 minutes A 30 0.00 0.90 1.8 B 50 0.01 1.5 0.3 C 70 0.08 2.1 4.2 Getting the concentrations 15ml of 70%ethanol +20ml water 70%of ethanol =70ml ethanol dissolved in 100ml of water 100ml of water contains 70ml of ethanol 15ml of water will contain? (1570) 100=10.5ml Hence 10.5ml of ethanol can be dissolved in 15 ml of water Concentration of the whole solution 15ml of 70% of ethanol+20ml of water (10.5100) 35=30% Molar concentration 10.535=0.3M Concentration of 50% ethanol+10ml of water (50100) 100= 50% 70% ethanol=70% Calculation of absorbance of a substance in each cuvette is based on knowing the transmittance which will build on the absorbance (Ultee, A. 2002). To calculate transmittance, we use the following formula: T=I/IO T represent transmittance I represent incident. IO represents incident intensity. Calculating the different absorbances, we use the following formula. A=-log (T) Whereby T is the transmittance. Calculating absorbance Log-1.41=0.15 Log -7.94=0.9 Log -125.89=2.1 (Lambert, R. 2001). Absorbance given correct to one decimal place This means as the transmittance increases the absorbance decreases hence the transmittance is inversely proportional to the absorbance since the increase in transmittance leads to a reduction of absorbance and a decrease in transmittance leads to an increase in absorbance. To determine the relationship between absorbance and concentration we need to look at Slope of the curve (m) =Y2-Y1/X2-X1 X represents concentration Y represents absorbance. This consequently leads us to the Y=MX+B Which derives another formula: A=?LC A represents the absorptivity From this we find that the absorption is inversely proportional to the transmittance and concentration is directly proportional to the absorbance (Garcia, C. 1994). Discussion The objective of the experiment is to study the effect of different ethanol concentration on the cell membrane integrity by using visible spectroscopy. An increase in the concentration of ethanol results to an increase in the permeability of the membrane hence high absorption results to an increase in the color intensity of the solution, from the results obtained this proved. The absorption by a substance in cuvette is determined by the concentration of the solution. This means that high concentrated ethanol has high interaction between molecules leading to increased absorbance by the solution hence the detection registered a higher absorption than the less concentrated solution. The highly concentrated ethanol (Ethanol 70%) registered high absorbance because there was increased interaction between molecules leading to large permeability of the membrane hence absorption improved due to an increase in the concentration of ethanol. Increase in the color intensity is directly proportion al to the membrane permeability. The cuvette with ethanol at a concentration of 30% has a slightly lower absorbance to the one with the 70% concentration since its molecules have less interaction compared to that of 70% concentration hence B. vulgaris is stronger in the 70% concentration. The ethanol with the lower concentration (ethanol 5%) registers a low absorbance since there is the least interaction between molecules leading to the lowest absorbance by the lowest concentrated solution. The B .vulgaris is not strong compared to the ethanol with 30% concentration. Absorbance is directly proportional concentration. Absorption of solution in cuvette is used to measure the damage on the cell membrane that directly affects the permeability of the cell. Other points to note can be the shape of cuvette can affect light transmittance, the rate at which the beet will be damaged is directly proportional to the absorbance and concentration and the darker the color of the solution due to the damage of the beet the more the absorbance and the more the concentration. In all this absorbance is directly proportional to the concentration of ethanol. The higher the concentration the higher the absorbance and the lower the concentration the lower the absorbance. Limitations When carrying out the experiment the following limitations were encountered, ethanol was flammable and could ignite, steam and hot water may cause burns, equally timing the time allocated by both cuvettes. Remedies This was solved by putting the ethanol away from the sources of ignition, wearing suitable heat proof gloves and opening the water baths carefully. References Ultee, A., Bennik, M.H.J. and Moezelaar, R., 2002. 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