What is the pH of trimyristine

Experiment 1 Extraction of nutmeg powder with a lipophilic solvent: Isolation of trimyristine (a triglyceride)

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1 organic-chemical internship for technical biologists 1 experiment 1 Extraction of nutmeg powder with a lipophilic solvent: Isolation of trimyristine (a triglyceride) organic-chemical internship for technical biologists 2 separation operation. The trimyristine is stored in a labeled preparation glass for experiment 12. Do not discard the ethanolic mother liquor, but use it in experiment 5. H 3 C (CH 2) 12 (CH 2) 12 CH 3 (CH 2) 12 CH 3 ground nutmeg dichloromethane X n R20-40, S23-24 / 25-36 / 37 95 percent ethanol F R11, S7-16 calcium chloride X i R36, S22-24 Approx. 20 g of ground nutmeg are weighed into a 250 ml round bottom flask. Approx. 150 ml of dichloromethane and a few boiling stones are added and the flask is attached to the stand. The reflux condenser is attached and the mixture is heated under gentle reflux for about 90 minutes. The liquid level in the flask should not be below that of the bath. It is then allowed to cool to room temperature and filtered through a folded filter (wipe the oil off the outer wall of the flask beforehand), which has been weighed beforehand, into a round-bottomed flask. The insolubles remaining in the extraction flask are rinsed completely into the folded filter with as little dichloromethane as possible and the residue located there is washed three times with 20 ml of dichloromethane each time. The residue including the filter is placed in a beaker and allowed to air dry (approx. 1 week). The extraction residue can then be weighed out. Calcium chloride is added to the dichloromethane extract to remove traces of water and it is moved in a circular motion for a few seconds. The round bottom flask is closed with a polyethylene stopper and left to stand for more than eight. The CaCl 2 is then filtered off in a previously weighed round bottom flask. The solvent is distilled off on a rotary evaporator. Weigh the arrears! What is its consistency and smell? The residue is dissolved in 95 percent ethanol on the boiling water bath. 150 ml of ethanol are used for 10 g of extract. Over eight is allowed to cool to room temperature, the trimyristine crystallizing out. The precipitated white solid is filtered off with suction and washed with as little ice-cold ethanol as possible. The trimyristine is dried over Sicapent for eight in a vacuum desiccator. If the trimyristine still smells of nutmeg, it is recrystallized again from ethanol (approx. 30 ml per gram). Pure trimyristine is completely odorless and has a melting point of 54-55 / C. Determine the yield (in percent) and melting point of the trimyristine you isolated. Weigh out the insoluble residue as well. Create a flow diagram of the

2 Organic-chemical practical course for technical biologists 3 Experiment 2 Separation of benzoic acid from autoxidized benzaldehyde Organic-chemical practical course for technical biologists 4 Experiment 3 Extraction of caffeine from tea leaves H Autoxidation 2, h. ν Benzaldehyde X n R22, S24 tert-butyl methyl ether FR 11, S atrium carbonate X i R36, S22-26 atrium sulfate concentrated hydrochloric acid C R34-37, S26-36 / 37 / 39-45 crude benzaldehyde (15.0 g, 141 mmol, 15.8 ml ) is diluted with 50 ml of tert-butyl methyl ether in a separating funnel. 100 ml of a 10 percent atrium carbonate solution are prepared in water (dissolves slowly) and the benzaldehyde solution is shaken out in portions (4 x 25 ml) in a separating funnel. The aqueous phase should finally show an alkaline reaction (pH indicator paper). Separate the organic phase. The aqueous phases are combined and washed with 2 × 20 ml of tert-butyl methyl ether. The organic extract is added to the first separated organic phase. The organic phases combined in a 100 ml round bottom flask are dried over atrium sulfate (the flask is closed with a polyethylene stopper). As little air as possible should stand above the drying solution in order to avoid renewed autoxidation of the benzaldehyde. To obtain the benzoic acid, the combined aqueous phases are carefully acidified to about pH 3 with concentrated hydrochloric acid. Benzoic acid should precipitate (if this is not the case, it could also be isolated by shaking with solvent). The benzoic acid is filtered off with suction and dried in a desiccator. Determine the amount and melting point of the isolated benzoic acid. Formulate the auto-oxidation of benzaldehyde in all sub-steps. Before use (experiment 25), the solution of the benzaldehyde is distilled in vacuo over a small Vigreux column. Caffeine tea (cheap quality is sufficient, bring your own, no fruit tea, etc.) Dichloromethane X n R40, S23-24 / 25-36 / 37 In a 600 ml beaker, 10 g of tea with approx. 200 ml of water are brought to a gentle simmer for 15 minutes heated (hot plate; boiling stones). The tea leaves are filtered off and the filtrate is concentrated to approx. 75 ml in a beaker by boiling the water. After the solution has cooled to room temperature, it is transferred to a 250 ml separating funnel, mixed with 30 ml dichloromethane and not shaken too vigorously (risk of emulsion formation). The two phases are separated and the aqueous phase is extracted twice more with 30 ml of dichloromethane each time. The combined dichloromethane extracts are dried over eight with magnesium sulfate. After filtering off the desiccant, the solvent is removed in a 250 ml round bottom flask on a rotary evaporator. Weigh out the raw product and determine the melting point in the sealed tube. The crude product is dissolved in the round bottom flask with as little dichloromethane as possible. This solution is transferred to the sublimation tube with a pipette and the dichloromethane is evaporated with a hair dryer (shake gently to avoid delayed boiling). Using the spatula, scrape the raw caffeine from the wall all the way to the bottom of the sleeve and put the cold finger on. A vacuum is carefully applied with the membrane pump and the sublimation apparatus is heated to approx. 200 ° C. in an oil bath. When enough sublimate has accumulated on the cold finger, it is carefully removed from the apparatus and the caffeine is removed with a spatula. This procedure is repeated several times if necessary. Draw a flow diagram of the extraction of caffeine from tea leaves. Calculate the percentage yield, give the melting point you observed and the melting point of the literature. The sublimed caffeine is stored in a preparation glass for later investigations (experiment 8).

3 Organic-chemical internship for technical biologists 5 Experiment 4 Purification of anthracene by adsorptive filtration over aluminum oxide Anthracene Anthracene (technical quality) X n R20 / 21 / 22-42 / 43, S22-24 aluminum oxide (basic, activity level I) Sea sand toluene F , X n R11-20, Organic-chemical internship for technical biologists 6 Determine the yield of recrystallized substance and the melting point. Observe the fluorescence of the purified anthracene. Anthracene can be oxidized to anthraquinone (experiment 18). This experiment is an example of what is known as adsorptive filtration. A solution of the mixture is continuously applied to the column until it is saturated with all adsorbable substances. In this process only the weakest adsorbed substance, which therefore runs on the front, is kept pure. All the others are mixed with the following fractions from the previous fractions. The process is suitable for cleaning chemical preparations from all more strongly adsorbable impurities. The accompanying substances of the techn. Anthracene is more strongly adsorbed than anthracene itself and is therefore retained. a) Filling the chromatography tube (slurrying in) A chromatography tube (40 x 1.6 cm, if the tube is small, halve all amounts) is fastened vertically to a stand with two clamps. Place an Erlenmeyer flask under the spout. With the tap closed, the mandrel tube fills up to about a quarter with toluene. With the help of a long glass rod, you push a wad of cotton wool into the narrowing of the spout and remove any air bubbles that may have formed by pushing it. A bubble-free suspension is prepared from 50 g of aluminum oxide and approx. 80 ml of toluene in an Erlenmeyer flask by shaking it and it is poured as vigorously as possible into the separation column through a powder funnel. Excess toluene is allowed to drain off by opening the tap. The suspension remaining in the Erlenmeyer flask is rinsed quantitatively into the column with toluene. The settling process is supported by tapping with a piece of vacuum hose. It is important to ensure that there is always a layer of solvent over the filling (the column must not "run dry"). When the aluminum oxide has settled, about 1 cm deep of sea sand is allowed to trickle in. Before adding the substance mixture to be separated, the supernatant liquid is drained off by opening the tap so that the sea sand is only covered with a thin layer of solvent. b) Separation process 0.5 g of anthracene (technical grade) is dissolved in about 50 ml of toluene. The solution is allowed to drop through the storage vessel onto the column - at the same time the stopcock at the lower end of the chromatography column is opened. The eluate should emerge with 1-2 drops per second. It is collected in a 250 ml round bottom flask, the weight of which has been determined. When all of the anthracene solution has been given up, it is eluted by dropping toluene on it. The eluate from anthracene is checked from time to time by placing a few drops on the joint of the piston stopper and blowing the liquid off to evaporate. The elution is stopped when no residue can be seen on the polished section (approx. 100 ml of toluene are required for elution). note the amount required. The eluate is distilled to dryness on a rotary evaporator and the crude yield is determined. The anthracene is recrystallized from 96 percent ethanol in the same flask. To do this, the crude product is dissolved in ml of EtH on a boiling water bath (note the amount) and allowed to crystallize over eight. The crystals are filtered off with suction, washed with a little ice-cold EtH and dried over calcium chloride in a desiccator.

4 Organic-chemical internship for technical biologists 7 Experiment 5 Organic-chemical internship for technical biologists 8 Fig. 1: Gas chromatogram of the fatty acid methyl ester from the saponified nutmeg extract Processing of the trimyristine mother liquor (ingredients of the nutmeg extract: the neutral fraction) Myristicin of the ethanolic mother liquor -Crystallization (from experiment 1) Potassium hydroxide C R35, S26-37 / 39-45 petroleum ether (low boiling) F R12, S ethyl acetate (ethyl acetate) F R11, S dichloromethane (methylene chloride) X n R40, S23-24 / 25-36 / 37 Magnesium sulfate To saponify the remaining triglycerides, 25 ml of a 50 percent potassium hydroxide solution is added to the ethanolic solution in a 250 ml flask. The dark brown reaction mixture is allowed to stand over eight. The solution is then transferred to a 500 ml separating funnel, 200 ml each of water and 100 ml petroleum ether are carefully added (do not shake - risk of emulsion formation!) And left to stand for more than eight (place a suitable size collecting vessel under the separating funnel, if this leaks). Fig. 2: Capillary gas chromatogram of the neutral fraction of nutmeg. The organic phase is separated off and dried with magnesium sulfate for at least 1 hour. After filtering off the drying agent, the solvents are distilled off on a rotary evaporator. The mixture of substances obtained in this way is weighed out and examined by thin layer chromatography (TLC); Pure myristicin (available from the assistant) is spotted as a reference substance. Development is carried out with the solvent mixture petroleum ether / EtAc (10: 1). Detection takes place under UV light and by spraying the film with the vanillin-sulfuric acid reagent (the stain is made visible by briefly heating the film with a hair dryer). From the substances contained in nutmeg, fats and essential oils were extracted from the builders in experiment 1 and a large part of the fat (trimyristine) crystallized out. The triglycerides still contained in the mother liquor are now saponified and remain in the aqueous phase during the extraction of the essential oil (salt formation in the basic medium). Fig. 1 shows a gas chromatogram of the methyl esters of these fatty acids. The complex composition of the essential oil in the petroleum ether extract is documented by the capillary gas chromatogram in Figure 2. The main component of this oil is the myristicin.

5 Organic-chemical internship for technical biologists 9 Experiment 6 Isolation of the essential oil from caraway seeds by steam distillation Carvone caraway spice (ground) n-hexane F, X n R11-48 / 20, S / magnesium sulfate Limonene 50 g caraway spice are crushed and put together With 300 ml of water in a 500 ml round-bottomed flask, add some boiling stones and add the Clais condenser (heating: oil bath with contact thermometer). A 100 ml round bottom flask serves as a template. un heated to vigorous boiling. After the templates are almost too full, mandie steam distillation stops. The distillate is poured into a 250 ml separating funnel, the round bottom flask is rinsed with 20 ml hexane and also poured into the separating funnel, which is shaken briefly. One waits until the floating hexane layer, which contains the essential oil, has separated cleanly and the water phase is allowed to drain off. The hexane phase is drained into a small round-bottomed flask and dried over eight with magnesium sulfate (close the flask and shake occasionally). Analyze the essential oil of the caraway seeds using thin layer chromatography (TLC). Use petroleum ether / EtAc (20: 1) as the eluent (limonene and carvone for reference are available from the wizard). Detection is carried out by UV light or spraying with vanillic sulfuric acid. Then separate the two main components of the caraway seed oil by column chromatography. For this purpose, 40 g of silica gel are suspended in 100 ml of petroleum ether / EtAc (20: 1) in a 250 ml beaker and filled into the chromatography tube. After the silica gel has settled, cover with approx. 0.5 cm of sea sand. 0.6 g of caraway oil are added and the mixture is eluted with petroleum ether / EtAc 20: 1. After a preliminary run of approx. 40 ml, the first fraction (approx. 25 ml) is removed. The following 80 ml of eluate can be discarded before another fraction (approx. 170 ml) is collected. In order to identify the ingredients of the caraway oil, the mixture and the individual fractions are spotted on a TLC plate. After the development, the R f values ​​of the individual substances are noted. The respective fractions are concentrated and weighed. Organic-chemical internship for technical biologists 10 Experiment 7 Boiling analysis of a mixture of hexane and glacial acetic acid n-hexane F, X n R 11-48 / 20, / acetic acid (glacial acetic acid) C R10-35, S Two students at a time carry out the experiment together . Build a distillation apparatus consisting of a 250 ml round-bottomed flask (joint S 14.5), Claisen condenser and a vent. The 10 ml measuring cylinder is used instead of the piston (provide 2 measuring cylinders that are used alternately). 100 ml of a mixture of equal parts by volume of n-hexane and acetic acid are filled into the flask and the magnetic stir bar is added. Heat slowly on the oil bath and watch the thermometer. If the boiling point of the more volatile component is reached, condensate forms in the cooler. Collect 9 fractions of exactly 10 ml as possible. The dripping speed must not exceed 1 drop per second. For each fraction, the vapor temperature is noted at the beginning and at the end of the withdrawal (if the first four fractions were not collected at constant temperature, you have heated up too quickly and your dripping speed is too high. If necessary, stop the distillation and start again). The ratio of n-hexane to acetic acid can be determined by extraction. To do this, after each removal, transfer the fraction into the graduated test tube and make up to 20 ml with water. Cork the test tube and shake. The acetic acid changes into the water phase, while the n-hexane forms a layer over it. Determine the volume of the n-hexane layer of each fraction as precisely as possible. Present in tabular and graphical form the amount of hexane in mole percent and the mean steam temperature of each fraction, both as a function of the fraction number. Does your result correspond to the theory? The hexane fractions of all groups are collected, dried over calcium chloride and redistilled from the participants in one group.

61 percent methanolic solutions. The painkiller is ground to a fine powder in a mortar (1 tablet per group). It is shaken in a test tube for a few minutes with 5 ml of methanol (extraction) and the undissolved components are filtered off (folded filter). The methanolic filtrate is spotted on a TLC foil (40 × 80 mm, 0.25 mm silica gel with fluorescent indicator) using a chromatography tube (single spotting is sufficient). The previously prepared solutions of the reference substances are spotted at a distance of 5-10 mm. The plate is left in the air for a few minutes or evaporation is accelerated by blowing with a cold hair dryer. You can check under the UV lamp whether enough substance has been applied. The chromatogram is developed with the solvent mixture toluene / ether / acetic acid / methanol (120/60/18/1).After marking the solvent front and evaporating the solvent, the TLC is viewed under the UV lamp. Mark the spots, determine the R f values ​​and compare the DC of the tablet extract with the reference substances. Make a note of the analysis result and discuss the sequence of the R f values ​​of the individual components. Also characterize the substances using spray reagents. The spray reagent is prepared by dissolving K 3 Fe (C) 6 and 0.60 g FeCl 3 in 30 ml water. After spraying, the spots of acetylsalicylic acid and phenacetine turn brown and blue, respectively. a) Preparation of the 2,4-dinitrophenylhydrazone In a 25 ml Erlenmeyer flask, 0.40 g (2.02 mmol) of 2,4-dinitrophenyl hydrazine are added and 2.0 ml of concentrated sulfuric acid are carefully added. The dinitrophenylhydrazine is crushed with a glass rod or spatula until a clear solution has formed. 3 ml of water are added dropwise while swirling the Erlenmeyer flask. 10 ml of 95 percent ethanol are carefully added to this warm solution. The unknown carbonyl compound (500 mg) is dissolved in 20 ml of 95 percent ethanol. The freshly prepared sulfuric acid 2,4-dinitrophenylhydrazine solution is added and the mixture is left at room temperature. stand. As a rule, the 2,4-dinitrophenylhydrazone crystallizes in 5 to 10 minutes, then it is cooled in an ice bath for 20 minutes and the dinitrophenylhydrazone is filtered off with suction. For recrystallization, the crude 2,4-dinitrophenylhydrazone is heated in 30 ml of ethanol in a 100 ml round bottom flask with a reflux condenser. If the substance dissolves immediately, water is slowly added with a pipette at the boiling point until a slight cloudiness is visible. If the dinitrophenylhydrazone does not dissolve in the specified amount of ethanol, small amounts of ethyl acetate are added to the boiling solution through the reflux condenser until the dinitrophenylhydrazone is completely dissolved. The hot solution is cooled to room temperature without shaking. For complete crystallization, the closed flask is left in the refrigerator for more than eight. The product is filtered off with suction, washed with small amounts of ice-cold ethanol and dried over P 4 10 in a desiccator. Determine the melting point and compare it to the melting points of 2,4-dinitrophenylhydrazones given in the list. b) Representation of the semicarbazone

7 Organic-chemical internship for technical biologists 13 Dissolve 500 mg (4.48 mmol) of semicarbazide hydrochloride and 1.00 g (7.35 mmol) of atrium acetate trihydrate in 4 ml of water in a test tube. To dissolve, it can be heated in a boiling water bath for about 2 minutes. The reagent solution is poured into the ketone (500 mg), which is located in a second test tube. Heat the mixture in a water bath for 4 minutes. If the precipitate that occurs when adding the ketone does not dissolve within the first 30 seconds of heating, add 1 ml of 95 percent ethanol. If the precipitate has not dissolved after 30 seconds when heated, add 1 ml of ethanol again. It may be necessary to repeat this process up to four times. If it was necessary to add ethanol, heat the solution for a further 6 minutes after the last addition. The test tube is then almost completely filled with ice water and the precipitated semicarbazone is separated by suction. The semicarbazone is recrystallized from 25 percent ethanol (approx. 5 ml) over eight. The next day, suction off the semicarbazone, dry it over P 4 10 in the desiccator and determine the melting point. Compare it with the melting points of the semicarbazones given in the list. What is the structural formula and the name of the ketone found? Write down the reaction equation for the reaction with semicarbazide and with 2,4-dinitrophenylhydrazine. State the yield in milligrams. Calculate the percentage yield based on the ketone used. State the melting points of the recrystallized products and the literature values ​​(see Table 3/1). Table 3/1: Melting points of the 2,4-dinitrophenylhydrazones and semicarbazones of the ketones r. Ketone 2,4-dinitrophenylhydrazone melting point [/ C] semicarbazone melting point [/ C] 1 2-butanone methylcyclohexanone pentanone cyclopentanone cycloheptanone pentanone cyclohexanone acetophenone organic-chemical practical course for technical biologists 14 experiment 10 reaction of acetic acid with isoamyl alcohol in the presence of acid HHH 2 S 4 acetic acid (glacial acetic acid) C R10-35, S isoamyl alcohol (3-methylbutan-1-ol) X n R10-20 sulfuric acid C R35, S dichloromethane X n R20, S24 Carrying out the reaction 10.9 ml (8.80 g. 100 mmol) isoamyl alcohol, 23.0 ml (24.0 g, 400 mmol) acetic acid and 3.00 ml (5.50 g, 540 mmol) concentrated sulfuric acid. Add a magnetic stir bar and attach a reflux condenser. The mixture is heated to a gentle boil in an oil bath for approx. 1-2 hours. The flask is removed from the oil bath and cooled with cold water below the boiling point of dichloromethane. Approx. 50 g of ice are then placed in a separating funnel and the reaction mixture is then poured into the separating funnel. 40 ml of dichloromethane are added (some of the dichloromethane is used to rinse the reaction flask) and shake thoroughly. After the phases have separated, the dichloromethane phase is allowed to flow off into an Erlenmeyer flask. The aqueous over phase is extracted again with 40 ml of dichloromethane. The two dichloromethane phases are then combined and washed with 50 ml of water in a separating funnel. The remaining acid in the dichloromethane phase is then removed by shaking twice with 25 ml of carbonate solution each time (dissolve 3 g of atrium carbonate in 50 ml of water). [Caution, when eutralizing C 2 develops, ventilate the separating funnel so that no overpressure builds up!]. The dichloromethane phase separated off in the separating funnel is dried over eight with magnesium sulfate. The desiccant is then filtered off in a round-bottomed flask and most of the dichloromethane is removed on a rotary evaporator. The liquid residue is transferred to a smaller distillation flask. The liquid is distilled in a vacuum (membrane pump) with a Vigreux column and spider. note the boiling points of the fractions. Complete the reaction equation. Discuss the influence of sulfuric acid and the relative molar amount of the starting materials. Determine the amount (in grams) of each fraction after distillation. Calculate the percentage recovery of isoamyl acetate. At 756 Torr, isoamyl acetate is said to boil at 142 ° C. What is the boiling point in the vacuum of your diaphragm pump? [The constants A and B of the vapor pressure equation are A = and B = The vapor pressure is calculated using the following equation: lg p = (A / T) B. Note: Sting pheromone of the honeybee: Aldrichimica Acta 1983, 16, 41.

8 Organic-chemical practical course for technical biologists 15 Experiment 11 Reaction of salicylic acid with methanol in the presence of acid Organic-chemical practical course for technical biologists 16 Experiment 12 Preparation of myristic acid (alkaline saponification of trimyristine) HHH 3 C (CH 2) 12 (CH 2 ) 12 CH 3 HH 2 S 4 KH, EtH reflux salicylic acid X n R22 methanol T, F R11-23, R25, S sulfuric acid C R35, S hexane X n, FR, S / Carry out the reaction in a 250 ml round bottom flask 16 ml of concentrated sulfuric acid were added to a mixture of 20.7 g (150 mmol) of salicylic acid and 71.5 g (2.23 mol) of methanol with ice cooling and magnetic stirring. The mixture is then refluxed for 2 hours. After cooling to room temperature, the reaction mixture is poured onto 100 g of ice in a separatory funnel. The organic phase is separated off and the aqueous phase is extracted twice with 25 ml of hexane each time. The combined organic phases are first washed with a 5 percent aqueous atrium hydrogen carbonate solution, then with 50 ml of water and dried with atrium sulfate [caution, when neutralizing C 2 develops, vent the separating funnel so that no excess pressure builds up!]. After filtering and concentrating on a rotary evaporator, the residue is distilled from a small round-bottomed flask over a short Vigreux column in a low vacuum. The methyl salicylate boils at 105 / C / 14 torr. Salicylic acid methyl ester is contained in percentages in wintergreen oil, which is obtained by steam distillation of the flowers, leaves and stems of a heather plant mainly found in North America. Wintergreen oil is mainly used in perfumery and for making soaps, as well as in medicine. (CH 2) 12 CH 3 Materials and instructions from GetStoftV Trimyristin from experiment 1 Ethanol F R11, S7-16 Potassium hydroxide C R35, S / 39 Hydrochloric acid C R35-37, S7 / 3.50 g of potassium hydroxide are weighed into an Erlenmeyer flask. Make up to 100 g with ethanol and loosen the KH by swiveling the flask. Transfer all trimyristine from experiment 1 into a round bottom flask with ground joint S 29. Add the ethanolic KH solution (15 ml KH solution are required for 1.00g trimyristine) and reflux in a water bath. Make sure that the piston is not more than half full. After boiling for 1 hour, the mixture is allowed to cool and water is added (for 1.00 g of trimyristine, 30 ml of H 2). The solution is concentrated somewhat on a rotary evaporator until the ethanol has distilled off. The alkaline solution is filtered and carefully (pipette) sufficient hydrochloric acid is added until the ph value is approx. 3 (dots on indicator paper). Myristic acid should separate out. The batch is left to stand at room temperature until the myristic acid solidifies. The crude acid is filtered off with suction and washed with water. The myristic acid is dissolved in as little ethanol as possible at room temperature. Then add enough water with a pipette until a cloudiness remains visible. Let crystallize in the refrigerator for about eight. The purified myristic acid is suctioned off through a small Buchner funnel and dried in a vacuum desiccator over P 2 5 or Sicapent. Write down the equation for the reaction of trimyristine with KH in ethanol. Discuss the work-up of the reaction. Calculate the percentage yield and give the melting point you observed.

9 Organic-chemical practical course for technical biologists 17 Experiment 13 Reaction of salicylic acid with acetic anhydride Organic-chemical practical course for technical biologists 18 Experiment 14 Reaction of p-aminophenol with acetic anhydride H H H H H 3 P 4 HCl aac. 3 H 2 salicylic acid X n R22 acetic anhydride (acetic anhydride) C R10-34, S26 85 percent phosphoric acid (orthophosphoric acid) C R34, S26 Carrying out the reaction 2.00 g (14.5 mmol) salicylic acid and 4.30 g are placed in a 100 ml flask (4.00 ml, 42.0 mmol) acetic anhydride. 5 drops of 85 percent phosphoric acid are added and the components are mixed by swiveling the flask. A reflux condenser is attached and the mixture is heated to 100 ° C. in an oil bath for about 5 minutes. Then the oil bath is removed. After the oil bath has been removed, 2.0 ml of water are carefully added to the still warm reaction mixture via the reflux condenser. The excess acetic anhydride is hydrolyzed [exothermic reaction! The resulting heat of reaction can cause the contents of the flask to boil]. When the vigorous reaction has subsided, 40 ml of water are added and the mixture is allowed to cool to room temperature. Remove the reflux condenser and stir the mixture with a glass rod. If the separated oil does not crystallize immediately, rub it with the glass rod (inside!) ("Scratch" it to crystallize). To increase the yield, it is cooled in an ice bath. After complete crystallization, the product is suctioned off with a small Buchner funnel, washed with a few drops of water and dried over phosphorus pentoxide in a desiccator. Give the percentage yield and the melting point of the crude acetylsalicylic acid. Pure acetylsalicylic acid melts at 135 / C. If the melting point is too low, recrystallize the product from a little water or isoamyl alcohol. Complete the reaction equation. Discuss the mechanism of the reaction. Why is phosphoric acid added? What should you watch out for when handling acid anhydrides and acid halides? How are these disposed of? Formulate the response. The experiment demonstrates acetylation of a phenol on a small scale (derivatization). The industrial process for the synthesis of the active pharmaceutical ingredient acetylsalicylic acid is similar. Acetylsalicylic acid was first produced by Gerhardt in 1853 and marketed as aspirin by Bayer in 1899. In the western countries the amount of industrially produced ASA was tons per year. At that time, every German took an average of 8 g of ASA per year, in the USA it was said to be as much as 50 g. H 2 p-aminophenol X n R, S28 acetic anhydride (acetic anhydride) C R10-34, S26 hydrochloric acid C R34-37, S2-26 atrium acetate trihydrate S22-24 / 25 activated carbon F R11, S16 carrying out the reaction 2.10 g (19.2 mmol ) p-aminophenol and 35 ml of water are placed in a 100 ml round bottom flask. 1.5 ml of concentrated hydrochloric acid are added and the round bottom flask is swirled for a few minutes until the p-aminophenol has completely dissolved. If the compound does not dissolve, add more drops of concentrated hydrochloric acid and continue swirling. A buffer solution is also prepared from 2.50 g (18.4 mmol) of atrium acetate trihydrate in 7.5 ml of water, which is filtered if it is cloudy. For acetylation, the magnetically stirred hydrochloric acid solution of the p-aminophenol is added in one pour with the atrium acetate buffer solution. Immediately add 2.16 g (2.00 ml, 21.2 mmol) acetic anhydride drop by drop (pipette) and stir further (approx. 10 min). The reaction mixture is cooled in an ice bath while stirring is continued; crude p-hydroxyacetanilide crystallizes out. If crystallization does not occur, scratch the (inner) wall of the bulb with a glass rod or obtain seed crystals. The mixture is left in an ice bath for about 1 hour and the crystals are then filtered off with suction. They are washed with ice water. Weigh the crystals and determine the raw yield. Recrystallization takes place from water (approx. 10 ml of distilled water are required for 500 mg of paracetamol). The crude product is placed in a 100 ml round-bottomed flask, the water and two spatula tips of activated charcoal are added and the mixture is heated to the boil (shake occasionally). If the boiling solution does not become discolored, it is allowed to cool down again and more activated carbon is added [be careful when adding activated carbon to hot solutions! There may be violent foaming]. A small funnel is placed on the flask, which is heated by the rising solvent vapors and at the same time serves as a reflux condenser. So much water is added in small portions from a measuring cylinder until the substance is completely dissolved at the boiling point. When adding the solvent, the funnel is removed so that it does not cool down. After each addition of solvent, the mixture is allowed to boil for a few minutes in order to reach saturation. note the amount of solvent required. The hot solution is filtered as quickly as possible through the hot funnel, in which a suitable filter paper has been placed, into a round bottom flask and allowed to cool slowly. If the substance should crystallize in the funnel, the filtrate is finally heated again to vigorous boiling with the funnel attached, until the crystals begin to rise

10 organic-chemical internship for technical biologists 19 have solved solvents. The solution is allowed to crystallize in the closed flask in the refrigerator for eight. The crystals are filtered off with suction and washed with ice water. They are dried over calcium chloride in a desiccator. Complete the above equation. Weigh out the substance and determine the melting point and the percentage yield (m.p./C). The p-hydroxyacetanilide is collected as a reference substance for thin layer chromatography (experiment 8). The working technique given here is suitable, since it is very rapid, for recrystallizing small samples of substances, e.g. B. for analytical purposes, from non-flammable, non-toxic solvents. It cannot be used because of the risk of fire for recrystallization with flammable solvents such as ether, petroleum ether, acetone etc. In these cases, crystallization is carried out in a ground-joint flask with a reflux condenser in an electrically heated oil bath. When decolorizing solutions, contaminated coloring by-products (mostly high molecular weight compounds), which often make the crystallization of the main product of a reaction more difficult, should be removed. If these impurities are physically and chemically significant differences from the main product, they can be selectively removed from the solution in question by adding a suitable adsorbent. The adsorbed impurities are filtered off with the adsorbent and discarded. In order to avoid losses in the main product, the smallest possible amounts of adsorbent must be used. Solutions in polar solvents are decolorized with activated charcoal; in non-polar solvents one works with aluminum oxide. When decolorizing with activated charcoal, it should be noted that sensitive substances are easily oxidized by adsorbed oxygen, especially when exposed to heat. Organic-chemical internship for technical biologists 20 Experiment 15 TLC analysis of 2,4-dinitrophenylhydrazones Finished film for TLC (0.25 mm silica gel with fluorescent indicator) Petroleum ether F R11, S Ethyl acetate (ethyl acetate) F R11, S Remove a trace of the 2, 4-Dinitrophenylhydrazone from Experiment 9 in a few drops of ethyl acetate (small preparation glass).Speckle 3 concentrations (1 drop, 2 drops, 3 drops) of this solution onto three adjacent starting points of a silica gel TLC slide. When the solvent has evaporated, a yellow spot must be visible at the starting point. Develop the TLC in a chamber with the following solvents (see the elutropic series): a) EtAc b) Petroleum ether / EtAc (8: 1) c) Petroleum ether / EtAc (4: 1) d) Petroleum ether / EtAc (2: 1 ) Mark the front of the solvent in each case, blow off the solvent with the hair dryer and determine the R f value of the dinitrophenylhydrazone. Transfer the chromatograms to your protocol. Record the R f values ​​for the different chromatographic systems. Discuss the influence of the solvent on the R f value.

11 Organic-chemical practical course for technical biologists 21 Experiment 16 Synthesis of methyl methyl ester organic-chemical practical course for technical biologists 22 Experiment 17 Reaction of resorcinol with phthalic anhydride in the presence of zinc chloride HH 3 / H 2 S 4 ZnCl 2, HCl methyl benzoate X n R36 / 37 / 38-42 / 43; S / 37/39 methanol F, T R11-23 / 25; S nitric acid (concentrated) C R35; S sulfuric acid (concentrated) C R35; Procedure Note: Halve the entire batch. 13.6 g (100 mmol) of methyl benzoate and 27 ml of ice-cold concentrated sulfuric acid are placed in a 100 ml flask. A mixture of 9 ml of concentrated nitric acid and 9 ml of concentrated sulfuric acid is slowly added dropwise with magnetic stirring and ice-cooling. The mixture is then stirred for a further 15 minutes. Work-up and purification The reaction mixture is poured into a beaker onto 100 g of ice. The crude product is filtered off with suction, washed with water until it reacts neutrally and then washed in an Erlenmeyer flask with 15 ml of ice-cold methanol. It is filtered off with suction and recrystallized from approximately the same amount by weight (based on the product) of methanol. Melting point: 78 / C. Yield: 80%. Experimental Results and Conclusions Determine the yield and the melting point of the methyl methybenzoic acid. What type of reaction is this implementation? Formulate the reaction mechanism! Phthalic anhydride X i R36 / 37/38 resorcinol X n R22-36 / 38, S26 zinc chloride (anhydrous) C R34, S7 / 8-28 hydrochloric acid (concentrated) C R34-37, S26-36 / 37 / 39-45 Carry out the Reaction 3.00 g (20.3 mmol) phthalic anhydride g (40.0 mmol) resorcinol and 2.00 g (14.7 mmol) anhydrous zinc chloride are placed in a 100 ml round bottom flask. The open flask is heated to 200 ° C. in an oil bath. After the reaction has ended (approx. 2 hours, the substance should now be solid), the mixture is allowed to cool to about 90 ° C. A mixture of 10 ml of water and 2 ml of concentrated water is carefully added. Hydrochloric acid is added and the mixture is heated to 100 ° C. for another 30 minutes. Occasionally stir with a spatula to loosen the product from the wall of the flask. The mixture is allowed to cool to room temperature, the dark red solid which crystallizes after about minutes (if necessary scratch again with a spatula) is filtered off with suction and washed with ice water until it no longer shows an acidic reaction. The product is dried in a desiccator. Determine the yield of fluorescein. Flourescein melts at / C, but the substance decomposes. Dissolve a grain of fluorescein in a little ammonia and dilute in a large beaker with 1 liter of water. Observation? H

12 Organic-chemical practical course for technical biologists 23 Experiment 18 Conversion of anthracene with atrium chlorate in the presence of vanadium (V) oxide to form 9,10- anthraquinone Organic-chemical practical course for technical biologists 24 Experiment 19 Oxidation of (-) - borneol to ( -) - Camphor acl 3, V 2 5 H acl glacial acetic acid H Materials and instructions from GefStoffV Anthracene X i R36 / 37/38, S23-24 / 25 atrium chlorate, X n R9-22, S Vanadium (V) oxide T, R20 / / 48/23/51 / 53-63, S36 / glacial acetic acid C R10-35; S sulfuric acid C R35, S ethanol F R11; S7-16 Carrying out the reaction Note: Batch size 1 g of anthracene is sufficient in principle! In a 250 ml three-necked round-bottomed flask with a KPG stirrer (grease the cylindrical ground joint well with silicone oil) and reflux condenser, add 5.35 g (30.0%) to 60 ml glacial acetic acid and 20 ml 0.4 sulfuric acid (solution of 0.4 g conc. Sulfuric acid in 20 ml water) mmol) anthracene, 4.80 g (45.0 mmol) atrium chlorate and 40.0 mg vanadium (V) oxide as a catalyst for the oxidation. The reaction mixture is heated to the boil with stirring and reflux; if too vigorous a reaction sets in, the oil bath is briefly removed. The mixture is then refluxed for a further hour. When the reaction mixture is cooled in an ice bath, the reaction product crystallizes out. It is suctioned off on a Buchner funnel / suction bottle and washed twice with 10 ml of ice water each time. Air-dry suction is carried out at the membrane pump, then the raw anthraquinone is transferred to a tared porcelain dish or a tared beaker and dried thoroughly in the desiccator over potassium hydroxide. For purification, the crude product is recrystallized from high boiling ethanol (100 ml round bottom flask with reflux condenser). When the reaction mixture is cooled in an ice bath, the reaction product crystallizes out. It is suctioned off on a Buchner funnel / suction bottle and washed with 10 ml of ice-cold ethanol. Yield 38-48%, pale yellow crystals, melting point / C test result, conclusion Complete the reaction scheme and formulate the redox equation for the reaction. Determine the percentage yield and melting point of anthraquinone. Materials and instructions of GefStoffV (-) - Borneol F R10, S24 / 25 glacial acetic acid C R10-35, S atrium hypochlorite (5.25% solution) X i Ethanol F R11, S7-16 atrium disulfite Carrying out the reaction In a 100 ml round bottom flask a Mixture made from 1.00 g (-) - borneol and 5 ml glacial acetic acid. A 5.25 percent solution of atrium hypochlorite (common household bleach) is added drop by drop (no faster than 5 ml in 5 minutes) and with vigorous stirring. After the addition of 15 ml of the bleach, the conversion is checked by means of thin layer chromatography with hexane: ethyl acetate = 9: 1 as eluent (mobile phase). The anisaldehyde reagent is used to stain the chromatography plate. If starting material can still be seen, a further 10 ml of bleach are added as described above. As soon as the starting material is no longer visible in thin layer chromatography, the addition of the bleach is stopped and the reaction mixture is checked with iodine starch paper (or KI starch solution on filter paper). If the latter turns blue, an aqueous atrium disulfite (a 2 S 2 5, structure?) Is added until the test with the iodine starch paper is negative. The reaction mixture is cooled in an ice bath and the precipitate is isolated by vacuum filtration through a Buchner funnel. After recrystallization from ethanol / water, the product is dissolved in a minimum amount of diethyl ether and small portions of atrium sulfate are added with swirling until the solution becomes clear. The supernatant solution is then decanted into a tared flask or filtered and the ether removed on the rotary evaporator (do not leave too long on the rotary evaporator, the ether is quickly removed, but the product is also highly volatile). Measure the melting point and create a thin layer chromatogram of your product (literature melting point of optically rhyming (-) - camphor / C, of ​​racemic camphor / C). Experimental result, conclusion Complete the reaction scheme and formulate the redox equation for the reaction. Determine the percentage of camphor recovery. Save your camphor for experiment 27 (close the flask tightly, the camphor sublimates slightly ...).

13 Organic-chemical practical course for technical biologists 25 Experiment 20 Oxidation of hydroquinone with cerium (IV) ammonium nitrate (CA) Organic-chemical practical course for technical biologists 26 Experiment 21 Representation of 1,2: 5,6-di-isopropylidene-aD -glucofuranose (and 1,2-isopropylidene-aD-g1ucofuranose) (H 4) 2 Ce (3) 6 H 3 CC, H 2 HHHHHHH 2 S 4 Materials and instructions of the GefStofffV Hydroquinone X n R20 / 22, S24 / 25 -39 Cer (IV) ammonium nitrate R8, S17 acetonitrile F, T R11-23 / 24/25, S Dichtormethane X n R20-40, S24 Carrying out the reaction In a small Erlenmeyer flask, mix 6 ml acetonitrile with 2 ml water and dissolves therein 220 mg (2.00 mmol) 1,4-dihydroxybenzene. 2.50 g (4.56 mmol) cerium (IV) ammonium nitrate are added and the flask is moved in circles until the cerium salt has dissolved. What color effect are you observing? After about 2 min, at room temperature, the mixture is poured into a small separating funnel onto 10 ml of water and extracted twice with 15 ml of dichloromethane each time. The dichloromethane phases are combined and dried over eight with magnesium sulfate. After the dichloromethane extract has been dried, the magnesium sulphate is filtered off through a small folded filter and concentrated on a rotary evaporator. p-benzoquinone remains as a yellow solid. Do not leave the flask on the rotary evaporator for too long as the substance sublimates easily. Determine the yield and melting point of the crude p-benzoquinone. Formulate the stoichiometric equation of the redox reaction. The p-benzoquinone obtained according to this procedure is sufficiently pure for conversion to quinhydrone (experiment 22). The water phase, which contains acetonitrile and cerium (III) salts, is not poured away, but disposed of together. Acetone F R1, S sulfuric acid (concentrated) C R33, S potassium hydroxide C R35, S / 39 dichloromethane (methylene chloride) X n R20-40, S24 cyclohexane F R11, S n-hexane F, X n R, S / D-glucose -Monohydrate (2.00 g mmol) is suspended in 20 ml of acetone in a 100 ml round bottom flask with a magnetic stir bar. It is cooled in an ice bath and, with ice cooling and stirring, 0.6 ml of concentrated sulfuric acid is added with the pipette. After that, mandase ice bath removed; After the mixture has reached room temperature, add another 1.3 ml of concentrated H 2 S 4. The mixture is allowed to stir for 2 hours at room temperature, whereupon the glucose almost dissolves and a yellow coloration occurs. While the batch is being stirred, 100 ml of aqueous potassium hydroxide solution (c = 1 mol / liter) are prepared, which is cooled with ice. After the reaction of the glucose has ended, the reaction mixture is poured into the potassium hydroxide solution with stirring. The yellow color disappears. The mixture is extracted three times with 20 ml of dichloromethane each time; the combined dichloromethane phases are washed three times with 20 ml of water each time. The washed dichloromethane extracts are dried over eight with atrium sulfate. After filtering off the desiccant, the solvent is removed on a rotary evaporator. The oily residue is taken up in approx. 10 ml of warm cyclohexane and stored in the refrigerator for over eight to crystallize. The precipitated crystals are filtered off with suction, washed with n-hexane and air-dried. The yield of 1,2: 5,6-di-isopropylidene-α-D-glucofuranose is 4 g (31%); Melting point 110 / C. Discuss the reaction, paying particular attention to the following questions: a) Crystalline glucose is in the pyranose form. However, the products contain hydrogenated furan rings. Explanation? b) What is the function of sulfuric acid? c) Why is the mixture poured into potassium hydroxide solution and not into pure water? d) What is the purpose of shaking out the dichloromethane extracts with water?

14 Organic-chemical practical course for technical biologists 27 Experiment 22 Presentation of quinhydrone Organic-chemical practical course for technical biologists 28 Experiment 23 Oxidation of 3-methylpyridine (ß-picoline) with potassium permanganate H KMn 4 p-benzoquinone (from experiment 20) T R23 / 25-36 / 37/38, S Hydroquinone X n R20 / 22, S24 / 25-39 H Make separate solutions of equal amounts of quinone (use all p-benzoquinone try 20) and hydroquinone in warm distilled water (for 100mg quinone and 100 mg of hydroquinone are required in each case 10 ml of H 2). Pour the two solutions together. Almost immediately, the green aromas of quinhydrone crystallize out. The product is filtered off with suction and washed with water. Dry the needles between a filter paper in the non-evacuated desiccator. Determine the quinhydrone yield. H 2, 90 C 3-methylpyridine (ß-picoline) X n R1-20 / 21/22, S26-36 potassium permanganate, X n R8-22, S2 hydrochloric acid (concentrated) C R34 / 37, S2-26 in a 100 In a ml flask, mix 10 ml of water and 1.00 g (10.7 mmol) of 3-methylpyridine. A magnetic stir bar is added and 4.20 g (26.6 mmol) of potassium permanganate are added in portions. The flask is then heated in an oil bath on a magnetic stirrer to approx. 90 / C until the violet coloration has disappeared from the permanganate (approx. 90 minutes). Manganese dioxide (brownstone) is deposited in the process. The still hot reaction mixture is suctioned off using a suction filter and the manganese dioxide precipitate is washed four times with 3 ml of hot water each time. The combined filtrates are concentrated to approx. 16 ml on a rotary evaporator and carefully adjusted to pH 3 (just add individual drops with the pipette) with concentrated hydrochloric acid (adjust the ph value precisely; dab on indicator paper!). The resulting voluminous, white precipitate (if nothing fails, evaporate and absorb in a little water, check and adjust the ph value again) is brought into solution by heating on an oil bath, adding more water if necessary using a pipette. The hot solution is allowed to cool slowly and is placed in the refrigerator for more than eight hours to complete the crystallization. The precipitated crystals of icotinic acid are filtered off with suction and washed with a few drops of ice water / ethanol mixture. The product is dried in the desiccator over calcium chloride (over eight). Yield: approx. 580 mg (44%) of colorless crystal powder; M.p./c. The mother liquor still contains approx. 420 mg of icotinic acid, which could be obtained by a second crystallization. Determine the percentage yield of pyridine-3-carboxylic acid and formulate the redox equation for the reaction. Manganese dioxide and icotinic acid are collected. Icotinic acid (pyridine-3-carboxylic acid) is a vital compound for humans and mammals, a vitamin. In the 1930's it was found that the disease pellagra is due to a vitamin deficiency and can be cured by the administration of icotinic acid. Hence the name vitamin PP (pellagra preventive factor). This disease with its typical "3-D symptoms" dementia, dermatitis and diarrhea is no longer a problem in developed countries thanks to improved nutrition (Pellagra occurs in cases of malnutrition, e.g. one-sided corn diet) Human icotinic acid is about mg / kg. Since the animal organism needs a lot of icotinic acid, it is now an important industrially produced feed additive. Of course, the very low toxic effect of icotinic acid is not surprising. The LD 50 at