1.

Title

: Absorption Analysis of EPMS and Acid-Base Indicators by Using UV-Visible Spectrophotometer

2. Objective : Determining the absorption of EPMS Determining the absorption of Teleng flower (Clitoria ternatea L.) Determining the absorption of phenolphthalein

3. Basic Theory: Ultraviolet-Visible Spectrometry Ultraviolet-visible spectrometry is an analytical method based on molecular absorption using ultraviolet radiation and visible light, with wavelengths between 160-780 nm. This method is widely used in quantitative measurement of various inorganic and organic compounds. Ultraviolet light has a wavelength between 160-400 nm and visible light that humans have typically seen the wavelength range between 400-800 nm.

Picture 1. Visible Spectrum Description:

Violet : 400 - 420 nm Indigo : 420 - 440 nm Blue : 440 - 490 nm

Green : 490 - 570 nm Yellow : 570 - 585 nm Orange : 585 - 620 nm Red : 680 780 nm

Ultraviolet-visible radiant energy with wavelengths between 160-780 nm corresponds to the transition of electrons involved in bonding in a molecule. In organic molecules, radiant energy absorbance in the ultraviolet-visible causes transition of electrons involved in bonding phi () primarily engaged in the phi electron conjugation system. Organic molecules have phi electrons involved in phi conjugated bond. While on an inorganic molecule, absorbance of radiation in the region appear visible to the transition of electrons in d orbital, and this is more common in complex compounds, because it is generally colored complex compounds. Sunscreen Lotion The wavelength range emitted by the sun's ultraviolet radiation covers that cannot be viewed eye. Ultraviolet is very dangerous for some types of skin and therefore skin should be protected with sunscreen lotion to avoid direct irradiation of the skin from the tropical sun. The sun emits electromagnetic radiation into the earth's surface with a range of 290 nm to 800 nm. Range of ultraviolet radiation is divided into two parts roughly UV A: 320-400 nm and UV-B: 280-320 nm. UV-B is responsible for skin damage (sunburn and tanning). When skin is exposed to ultraviolet radiation, skin cells will produce brown pigment called melanin and will become more brown skin and thinning. More and more exposed to ultraviolet radiation in the range 290-320 nm, the more melanin is formed and the skin will get darker. Furthermore, ultraviolet radiation causes skin damage to DNA and proteins, thus cause very harmful effects of skin cancer or melanoma. In an effort to minimize the effects of direct irradiation of ultraviolet radiation, various lotions (sunscreen) (sunscreen lotion) have been developed. The purpose of the lotion or cream is to filter out radiation that can potentially damage the skin, the wavelength of 290-320 nm. Sunscreen products will be effective if it contains compounds that can absorb radiation at these wavelengths. Usually more than one compound should be used to convince a range of radiation is absorbed. EPMS is extracted from greater is one of the active compound to make sunscreen.

Ethyl p-Methoxycinnamate (EPMS) Greater (Kaempferia galanga L.) is one type of drug plants belonging to the Zingiberaceae. Rhizome or rhizome of this plant contains essential oils and alkaloids are used as a stimulant. A compound most widely in the rhizome essential oil of greater is ethyl p-methoxy cinnamate (EPMS). Rhizome isolation of greater obtained EPMS by 2.4% of dry weight, because it can easily be isolated from the tuber using a solvent of petroleum ether or ethanol. Ethyl pmethoxycinnamate crystal has a melting point of about 48o-49oC.
O

OC 2 H 5 H 3 CO

Picture 2. Greater and Ethyl p-Methoxycinnamate (EPMS) Ethyl p-methoxy cinnamate (EPMS) is one compound which is the basic ingredient of sunscreen which protective skin from the sun. EPMS included in the class of ester compounds containing benzene rings and methoxy group which is nonpolar and also the carbonyl groups that binding of ethyl are less polar so that the extraction can use solvents that have a polarity variation such as: ethanol, ethyl acetate, methanol, water, and hexane. In isolation experiments of EPMS can be used soxlet extraction method. Separation principle is based on the distribution ratio of solute in two solvents that do not dissolve each other. Greater-paste, put in soxlet tool that has been wrapped with filter paper. Extraction sokhlet discontinued when the suspected substance in the circulation that will be extracted are exhausted, this is indicated by no color change of solvent after passing through the sample. The organic solvent used in isolation EPMS using soxlet extraction method is ether. Crystals of ethyl pmethoxy cinnamate obtained usually is mixed with impurities. Purification can be

done by recrystallization using ethanol and proceed with the test crystal melting point EPMS.

Acid-Base Indicators Acid-base indicator is a substance whose color can change as interact or react with acidic compounds and basic compounds. Indicators of acid-base commonly used are artificial and natural indicator. a. Artificial Indicators The nature of acid/base of a substance can be determined by taste. However, not all substances to be sampled prior safe chemicals in the laboratory. Accordingly, for purposes of experimentation, scientists create litmus. Litmus is a similar substance obtained from the types of lichens (Roccella tinctoria). In addition to litmus, are still a lot of artificial acid-base indicators such as phenolphthalein, methyl red and blue bromthymol. Various types of artificial acid-base indicator and changes color in a solution of acid or base and its pH range can be seen in the following table: Color Indicators In acid Blue thymol Blue bromophenol Orange methyl Red methyl Blue chlorophenol Blue bromotimol Red cresol Phenolphthalein Red Yellow Orange Red Yelow Yellow Yelow Colorless Yellow Blue purple Yellow Yellow Red Blue Red Reddish pink In base 1.2 2.8 3.0 4.6 3.1 4.4 4.2 6.3 4.8 6.4 6.0 7.6 7.2 8.8 8.3 10.0 pH range*

* PH range is defined as the range in which the indicator color changes from acid to alkaline color Phenolphthalein is one of artificial indicator. Phenolphthalein as an acidbase indicator, and will know that it is colorless in acidic conditions and magenta

(bright pink) in an alkaline solution. This color change related to changes in the molecule explains below: The structures of the two differently colored forms are:

Both of these absorb light in the ultra-violet, but the one on the right also absorbs in the visible with a peak at 553 nm. The molecule in acid solution is colorless because human eyes can't detect the fact that some light is being absorbed in the ultra-violet. However, human eyes do detect the absorption at 553 nm produced by the form in alkaline solution. This absorption at 553 nm is in the green region of the spectrum (the complementary color of green is magenta - and that's the color human see). Human have is a shift to absorption at a higher wavelength in alkaline solution. As human have already seen, a shift to higher wavelength is associated with a greater degree of delocalization. Here is a modified diagram of the structure of the form in acidic solution - the colorless form. The extent of the delocalization is shown in red.

Notice that there is delocalization over each of the three rings - extending out over the carbon-oxygen double bond, and to the various oxygen atoms because of their lone pairs. But the delocalization doesn't extend over the whole

molecule. The carbon atom in the centre with its four single bonds prevents the three delocalized regions interacting with each other. This is comparison with the magenta form:

The rearrangement now lets the delocalization extend over the entire ion. This greater delocalization lowers the energy gap between the highest occupied molecular orbital and the lowest unoccupied phi anti-bonding orbital. It needs less energy to make the jump and so a longer wavelength of light is absorbed. b. Natural Indicators Besides the artificial indicators such as phenolphthalein, methyl red and blue bromthymol, compound acid/base can be identified by using natural indicators, such as hibiscus, turmeric, red cabbage and various other plant species. The procedures for making a natural indicator of flower are as follows: The characteristics of a good flower used as a pH indicator that is still fresh flowers dark colored (light), used only crown flower, while stamen and pistil not in use. In the preparation of liquid indicators, flowers are washed with clean running water for flower color pigments are intended to not participate soluble in water. Flower that has been washed and cut into small pieces to expand the surface so that the process of dissolution rates of flower more effectively. The more surface area of flower, the more color pigment of flower is soluble in the leaching process. In the process of cutting, flower does not chopped into small pieces. After cut flower, dried flower later in the oven to reduce the water content contained, conducted at a temperature of 50C for 15 minutes. At these

temperatures, flower pigments have not changed so that when dissolved will produce a readily observable color. When drying performed at a temperature greater than 50C then the flower color will change because the characteristics of early flower color is lost. Dried flowers are already included in the jar and alcohol 70% is added to approximately 0.5 cm submerged flowers. Flowers left in place overnight for flower color pigment soluble in alcohol. Alcohol is actually 70% ethanol, which was chosen as a solvent in addition to views of polar characteristics also be seen from the economic aspect. Ethanol is more readily available and cheaper than other types of alcohol. The use of solvents to dissolve the flower used to taste because if excessive then the resulting solution will be diluted, causing the resulting product is less good. After last night, the solution is filtered to obtain filtrate flower extract. Flower extract is a liquid indicator. Then the indicator liquid is poured in another jar and stored in a refrigerator until used. How to use liquid indicator that these indicators shed on the solution to be tested for its pH. Solution will provide the color change and then change the color matched the color of the route pH indicator. Each color on the route has different pH. The color of the same solution with the color on the route showed that the pH of the solution pH equal to pH at pH route indicator. Flower is used as a natural indicator by observer is teleng flower. These plants spread and usually found in the yard or forest edge. This plant family of the tribe of legumes (papilionaceae) is derived from tropical Asia, but now has spread throughout the tropical regions. Teleng flower (Clitoria ternatea L.), known as the blue flower, telang flower. Teleng flower has a variety of chemical constituents, including: saponins, flavonoids, alkoloid, Ca-oxalate and sulfur, especially its leaves: kaempferol 3-glucoside and triterpenoids. The flowers also contained delphinidin 3.3'.5' and triglucoside, phenol. Roots are poisonous.

Picture 3. Teleng flower Since the first plant in the garden is planted as an ornamental plant because the flowers are bright blue. The flowers in various parts of Southeast Asia are used as food coloring or cake. In Malaysia, crown flower extract is used to color the sticky rice. In Thailand, the flowers are used as blue-colored beverages. Other variety of this flower is a flower with a white crown.

4. Chemical and Chemical Apparatus : In this experiment, the chemical and chemical apparatus used are: Apparatus Digital Scales volumetric flash 25 mL volumetric flash 50 mL volumetric flash 100 mL Mortar and pastle Measuring pipette 5 mL beaker 100 mL Spatula dropped Watch glass stirring rod Amount 1 1 piece 1 piece 1 piece 1 set 1 1 piece 1 piece 3 pieces 1 piece 1 piece Chemicals solution of EPMS Clitoria ternatea L. Ethanol distillate water solution of PP solution of HCl 0,1 M solution of NaOH 0,1 M Description 10 mL 15 tangkai 500 mL 500 mL 10 mL 5 mL 5 mL

Petri dish UV-Vis spectrophotometer tube Tube rack

1 piece 1 set

12 piece 1 piece

5. Procedure and Observation Result Part I : Measurement EPMS No 1 Work procedures EPMS is still in the form of slightly viscous liquid was dissolved by using 95% ethanol. Observations EPMS slightly brown colored fluid. When reconstituted with 95% ethanol solution formed a transparent brown.

Cuvette is filled with EPMS solution, EPMS spectra measurement, there are two peaks then measured spectrum by using 95% ethanol as a reference sample. where the maximum absorption at a wavelength of 307.0 nm with the absorbance is 1.834.

Part II : Measurement of Teleng Flowers No 1 Work procedures Flowers teleng smoothed by using a mortar and pestle. Observations Teleng Flowers (Clitoria ternatea L.) is a blue flower. When mashed produced little dark purple liquid.

Flowers that have been mashed dissolved using 95% ethanol.

When reconstituted with using ethanol solution remained purple.

Cuvette filled with extracts of Teleng flower then measured spectrum by using 95% ethanol as a reference sample.

In the spectrum measurement teleng flower extract contained five peaks where maximum absorption at a wavelength of 211.0 nm with the absorbance is 1.042.

Extracts that had been measured earlier added a few drops of HCl 0.1 M. Furthermore, its spectrum was measured again.

In the spectrum measurement extracts of teleng flower were added with HCl there are four peaks where maximum absorption at a wavelength of 268.0 nm with the absorbance is 1.022. When the first extract of teleng flower purple plus HCl solution turns red.

Teleng flower extract on cuvette replaced with a new pupil flower extract and then added a few drops of NaOH. Next measured spectrum.

In the spectrum measurement teleng flower extract is added with NaOH there are five peaks where maximum absorption at a wavelength of 277.0 nm with the absorbance is 0.826. When the first extract of teleng flower purple plus NaOH solution changes color to green.

Part III : Measurement of PP No 1 Work procedures PP solution made by dissolving 1 gram of solids into a 100 mL phenolphthalein aquades. Observations PP is a white solid. When diluted with water does not all dissolve PP. Colorless solution was formed.

Cuvette filled with a solution of PP

In the spectrum measurement of PP solution there are

and its spectrum was measured using three peaks in which the maximum absorption at a 95% ethanol as a reference sample. wavelength of 229.0 nm with the absorbance is 0.396.

Cuvette filled PP solution is then added with HCl and measured spectrum.

In the spectrum measurement of PP were added to a solution of HCl, there are three peaks in which the maximum absorption at a wavelength of 228.5 nm with the absorbance is 0.329. PP solution is a colorless solution when added to HCl solution remains colorless.

Cuvette PP filled with a new solution is then added with NaOH then measured spectrum.

In the spectrum measurement of PP were added to a solution of NaOH, there are two peaks where the maximum absorption at a wavelength of 553.0 nm with the absorbance is 0.407. PP solution is a colorless solution when added to the NaOH solution became pink.

6. Discussion The discussion of this experiment are : Wavelength Absorption Determination of Compound Lotion Active Ingredients in Sunscreen EPMS In this experiment the wavelength of absorption measurements were taken the active ingredients in products sold sunscreen free. One of the active compound in question is ethyl the methoxycinnamate. Before the measurement wavelength absorption in these compounds, the methoxy ethyl which is in the form of viscous liquid was dissolved in ethanol. Ethanol solvent was chosen because a compound will be dissolved (EPMS) are compounds that dissolve in organic solvents besides ethanol also showed no absorption at 200-1000 nm spectral region. This ethanol solution was put into cuvette. Subsequently a solution of ethyl p-

methoxycinnamate which has been diluted and incorporated into spectra cuvette measured. EPMS is one of the active ingredients found in sunscreen lotions. EPMS Solution absorbance was measured with UV-Vis spectrometer instrument, the way to put the solution on cuvette and operated tool for measuring absorbance. The EPMS absorption curve generated from UV-Vis instrument as follows.

Figure 4. EPMS absorption curve From the absorption curve above can be seen that the absorption EPMS gives two peaks and one valley, where the highest peak shows a maximum absorption of Ethyl p-methoxycinnamate which occurs at a wavelength of 307 nm with 1.834 absorbance. Determining The Absorption of Teleng Flower (Clitoria Ternatea L.) Teleng flowers or Clitoria ternatea L. is blue flowers that including Papilionaceae family of plant. The variety chemical content, already been known that is: Saponins, flavonoids, alkoloids, Ca-oksalat and sulphur. In this experiment the flower was crushed and dissolved in ethanol produces a dark purple solution. After that the Solution was diluted with ethanol until the color is transparent. This diluted solution that has been tested by UV-Vis and obtained the following data:

Figure 5. Absorbance curve of neutral solution From the curve is known that compound on solution of Clitoria ternatea L. solution can absorb light with a wavelength of 211.0 nm with absorbance 0168, wavelength 268.0 nm by absorbance 0179, wavelength 308.0 nm by absorbance 0870, at a wavelength of 577.0 nm with 1.012 absorbance , at a wavelength of 623.0 nm with the absorbance of 1.042. This shows that the compound on Clitoria ternatea L. solution can absorb visible light (wavelength 577.0 nm and 623.0 nm) and ultraviolet light (wavelength 211.0 nm, 268.0 nm, and 308.0 nm). But there is maximum absorption at a wavelength of 211.0 nm that is equal to 1.042 so that the maximum absorb ultraviolet radiation and can be used as a sun screen lotion. Not only absorption test in neutral circumstances, but also absorbance at acidic and alkaline conditions of Clitoria ternatea L. solution was determined. To pickle Clitoria ternatea L. solution, some of 0.1 M HCl was added and the color

of solution turns red. This solution was tested by UV-Vis absorbance. From the measurement results obtained the following data:

Figure 6.Absorbance curve of Clitoria ternatea L. solution in acidic conditions From the curve is known that Clitoria ternatea L. solution in compounds in acidic conditions can absorb light with a wavelength of 225.0 nm with absorbance 0845, wavelength 268.0 nm by absorbance 1.022, wavelength 299.0 nm with the absorbance of 0.884, and at a wavelength of 549.0 nm with the absorbance 0174. This shows that the compound on Clitoria ternatea L. solution can absorb visible light (wavelength 549.0 nm) and ultraviolet light (wavelength 299.0 nm, 268.0 nm, and 225.0 nm). But there is maximum absorption at a wavelength of 268.0 nm that is equal to 1.022 so that it can absorb ultraviolet radiation and can be used as a sun screen lotion.

Furthermore, to Clitoria ternatea L. solution in a state of alkaline condition can be made by adding 0.1 M NaOH solution, the solution changes color to green. From the measurement results with UV-Vis data obtained following data:

Figure 7. Absorbance curve of Clitoria ternatea L. solution in an alkaline condition From the curve is known that Clitoria ternatea L. solution in alkaline conditions, which can absorb green light with a wavelength of 229.0 nm with absorbance 0.738, wavelength 277.0 nm by absorbance 0.826, wavelength 312.0 nm by absorbance 0.558, at wavelength of 396.0 nm with absorbance 0.617, and at a wavelength of 593.0 nm with the absorbance of 0.092. This shows that the compound on Clitoria ternatea L. solution can absorb visible light (wavelength 593.0 nm) and ultraviolet light (wavelength 229.0 nm, 277.0 nm, and 312.0 nm, and 396 nm). But there is maximum absorption at a wavelength of 277.0 nm that

is equal to 0.826 so that the maximum absorb ultraviolet radiation and can be used as a sun screen lotion. From these observations may also note that the flower extract Clitoria ternatea L. can be used as an indicator of acid-base due to show obvious color changes. It changes because of anthocyanin that contented on the flowers. Anthocyanin is one kind of flavonoid that has conjugated double bond. This bounding make the flowers can absorb visible light. This following figure is the structure of anthocyanin

Figure 8. Structure of anthocyanin

Wavelength Absorption Determination of PP In The State of Acids and Bases The wavelength of absorption measurements was taken the active ingredients in the PP (phenolphthalein). The structure of PP when added with the acid and base are as follows.

Figure 9. Fenolfttalein reaction with acids and bases

PP In Neutral Condition Before the measurement wavelength absorption in the compound, the PP is in the form of a white crystalline solid that is dissolved in distillate water. After reconstitution formed a colorless solution. First of all water put into cuvette one as a reference. Furthermore, PP solution incorporated into the two and spectra

measured. PP neutral solution absorbance measured by UV-Vis spectrometer instrument, the way to put the solution on cuvette and operated tool for measuring absorbance. The PP neutral absorption curve generated from UV-Vis instrument as follows.

Figure 10. PP neutral absorption curve From the absorption curve above can be seen that the absorption of PP gives three peaks, namely at a wavelength of 342.5 nm with 0.130 absorbance, at wavelength 276 nm with 0.163 absorbance, and at a wavelength of 229 nm with 0.396 absorbance. Where the highest peak is shows a maximum absorption of the PP that occurs at a wavelength of 229 nm with 0.396 absorbance.

PP In Acid Solution Then the second experiment, a solution of 0,1 M HCl added PP Once added, a solution of PP is not being changed. The following reaction is the reaction between the PP with acid.

Addition of acid (hydrogen ions) excess will shift the position of equilibrium to the left, and turn the indicator colorless. As already seen, the shift to higher wavelength associated with a greater degree of delocalization. Here is a diagram of a modification of the structure forms in acid solution. Delocalization level shown in red.

Figure 11. The level of acid delocalization Notice that there is delocalization over each of the three rings - extending out over the carbon-oxygen double bond, and to the various oxygen atoms because of their lone pairs. But the delocalization doesn't extend over the whole molecule. The carbon atom in the centre with its four single bonds prevents the three delocalized regions interacting with each other. So the acid molecule in solution is colorless because our eyes cannot detect the fact that light is absorbed in the ultra-violet. Further absorption wavelength measurement of PP in a acidic atmosphere. PP solution in acidic conditions was measured absorbance with UV-Vis spectrometer instrument, the way to put the solution on cuvette and operated tool for measuring absorbance. The absorption curves of PP in a state of acids produced from UV-Vis instrument as follows.

Figure 12. Absorption curves of PP in an atmosphere of acid From the absorption curve above can be seen that the uptake of PP in a acidic conditions gave three peaks, namely at a wavelength of 342 nm with 0.079 absorbance, at wavelength 276 nm with 0.108 absorbance, and at a wavelength of

228.5 nm with 0.329 absorbance. Where is the highest peak of maximum absorption of the PP in an atmosphere of acid occurs at a wavelength of 228.5 nm with 0.329 absorbance.

PP In Base Solution The third experiment, a solution of 0,1 M NaOH added PP once added, the solution changes color from colorless PP turn pink. The following reactions that occur between the PP with the bases.

This is caused by the addition of hydroxide ions which remove hydrogen ions from the equilibrium that leads to the right to replace the indicator change to pink. In this case occurred the rearrangement now lets the delocalization extend over the entire ion. This greater delocalization lowers the energy gap between the highest occupied molecular orbital and the lowest unoccupied pi anti-bonding orbital. It needs less energy to make the jump and so a longer wavelength of light is absorbed. The following structures.

In this case, our eyes do detect absorption at 553 nm generated by the shape of the base solution. Where 553 nm is in the green region of the spectrum. When seen again the color wheel, will find that the complementary color is magenta and green color that is being viewed. Furthermore, uptake measurement wavelength of PP under alkaline conditions. Alkaline solution of PP in a state of measured absorbance on UV-Vis spectrometer, by putting the solution on kuvet and operated tool for measuring absorbance. The absorption curves of PP in alkaline conditions resulting from UV-Vis instrument as follows.

Figure 13. Absorption curves of PP under alkaline From the absorption curve above can be seen that the uptake of PP in alkaline conditions gave two peaks, i.e. at a wavelength of 553 nm with 0.407 absorbance and the wavelength of 370 nm with 0.082 absorbance. Where is the highest peak of maximum absorption of the PP under alkaline conditions occurs at a wavelength of 370 nm with 0.082 absorbance.

7. Conclusion According to the discussion above, it can concluded that: a. The absorption of EPMS is 1.834 at a wavelength of 307.0 nm b. The absorption of Teleng flower (Clitoria ternatea L.) is 1.042 at a wavelength of 211.0 nm c. The absorption of phenolphthalein is 0.407 at a wavelength of 553.0 nm

REFERENCES Anonim. 2009. Analysis UV-Visible. Accessed on April 1st from

http://www.chemguide.co.uk/analysis/uvvisible/theory.html#top Clark, Jim. 2007. Phenolphthalein. Accessed on April 1st 2011 from http://www.digipac.ca/chemical/mtom/contents/chapter3/phenolphthalein. htm Muderawan, I Wayan. 2010. Analisis Instrumen. Singaraja: Jurusan Pendidikan Kimia, UNDIKSHA.

Wikipedia.

2009.

Phenolphthalein.

Accessed

on

April

1st

from

http://en.wikipedia.org/wiki/Phenolphthalein