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I. Esterification Reaction 1. Ingredients: Material name, Molecular weight, Moles, Mixing ratio, Amount to be added – Cholesterol, Petroleum ether, Acetic anhydride, Anhydrous sodium carbonate, DMAP (AD), Anhydrous sodium sulfate, 4-Dimethylaminopyridine, 0.5 W/W, 2.5 kg; Hot water (50–60°C). 2. Procedure: Dry the esterification reactor and prepare it for use, and have all the materials required for the reaction ready. 450 liters of petroleum ether are placed into the esterification reactor. Stirring is initiated, and cholesterol, anhydrous sodium carbonate, and AD are added. When the internal temperature rises to 40–45°C, acetic anhydride is gradually added over a period of at least 30 minutes. After adding the liquid, raise the temperature to 64–68°C and allow the reaction to proceed under reflux for 5 hours. Take samples for testing; once they meet the requirements, transfer the solution to the hydrolysis tank and cool it until the internal temperature reaches 50°C ± a certain range. Then add 200 L of hot water (at 50–60°C), being careful not to cause the material to be displaced. The process takes about 20 minutes; after that, stir for 10 minutes and let it stand for 15 minutes. Layer it, then wash the oil layer twice with 200l×2 of hot water; the water layer should be neutral. Separate the aqueous layer, reheat and stir until reflux is achieved; then stop stirring. After half an hour, drain the water from the jacket, allow the mixture to stand for 1 hour, separate the aqueous layer again, add 25 kg of anhydrous sodium sulfate, stir for 20 minutes, and let it stand for 1 hour. Take a sample to determine the moisture content – a value of less than 0.09% indicates that the sample meets the requirements. Filter, and draw the filtrate into the upper bromination pot to allow bromination. 3. Precautions: (1) If the plate test is unsatisfactory, 5 L of acetic anhydride (3.5 kg of anhydrous Na2CO3 and 0.01 kg of DMAP) should be added, the reaction time should be extended by 1–2 hours, and the plate test should be conducted again. ⑵After three washes, if the water layer remains acidic (pH < 6.5), it should be washed again with 200 L of hot water until it reaches a neutral pH. ⑶Each time an esterification is carried out, the residual water in the U-tube below the condenser must be drained completely. ⑷After adding the material to the hydrolysis pot, no more petroleum ether is added. II. Bromination Reaction 1. List of raw materials: Name, molecular weight, number of moles, mixing ratio, and amount to be used – Choline ethyl ester (in petroleum ether solution), additional petroleum ether, NBS, petroleum ether for washing the reactor. 2. Procedure: Dry the reactor used for bromination and set it aside; prepare all the necessary raw materials. Monitor the liquid level in the bromine reactor, add an appropriate amount of petroleum ether (50–100 L), start stirring, add NBS and NaHCO3, and circulate hot water through the jacket to raise the temperature; the temperature should reach 64–68°C after about an hour, after which the reaction is carried out under reflux for 90 minutes. After the reaction is complete, the filtrate is drawn off while still hot to a debromination pot for solvent recovery, and the pot is washed with 50 L of petroleum ether. 3. Precautions: (1) Water must not be introduced during the bromination reaction under any circumstances. NBS, NaHCO3: Be careful to avoid moisture absorption and caking, especially during rainy weather. ⑵When concentrating petroleum ether, the temperature of the jacket water must not exceed 65°C. ⑶During suction filtration and concentration, to prevent the material from being pushed by excessive vacuum, the vacuum can be reduced. The vent outlet should be equipped with a drying device. ⑷The debromination pot must be completely dried and free of moisture in advance. III. Debromination reaction, crystallization. Material name, Molecular weight, Number of moles, Mixing ratio, Amount added: Bromocholesterol ester, Xylene (water content < 0.09% = PD), Pyridine, Tap water, Saturated salt water, Propylene used for washing, Propylene used for pot washing (pre-cooled), Propylene used for rinsing (pre-cooled). 2. Operation: Under stirring, add xylene to the debromination tank and stir for 5 minutes; then add PD and raise the temperature. Timing starts when the internal temperature reaches 125°C, and the reaction is carried out at 125°C–130°C for 100 minutes. After the reaction is complete, the temperature is reduced to 100°C, 250 L of tap water is added, stirred for 5 minutes, and then left to stand for 20 minutes. The lower water layer is extracted once with 25 L of xylene, while the wastewater is retained for PD recovery. The oil layers were combined, and washed twice with 75L×2 of saturated saline solution, with stirring for 5 minutes each time and allowing the mixture to stand for 15 minutes. The water layer is separated; the pH should be around 7. The oil layer is fed into the xylene concentration pot, where xylene is concentrated under reduced pressure. The final vacuum level reaches above -0.09 MPa, and the temperature reaches 95–100°C. When very few distillate droplets are observed, it can be considered that the oil has been completely dried. After circulating hot water through the jacket for 5 minutes, add 150 L of acetone and stir thoroughly for 10 minutes. The mixture is then pumped into the crystallization vessel; subsequently, the concentration vessel is washed with 50 L of acetone, and the wash liquid is also transferred to the crystallization vessel. Start stirring in the crystallization tank, turn on the freezer; once the internal temperature drops to 0°C, continue stirring for another hour, then allow the crystals to settle for more than 8 hours. While stirring, quickly discharge the material while it’s cold to avoid losses. The material is centrifuged for more than 30 minutes; then the centrifuge is turned off. After the rotation speed slows down slightly, it is rinsed with cold acetone pre-cooled to -15°C ±. The same pre-cooled acetone used in the crystallization tank is employed, in a volume of 150 L. Then start the centrifuge and spin it thoroughly for over 30 minutes. The product is discharged and dried in a vacuum oven for 10 hours to yield dehydrocholic acid ester, which is then prepared for saponification. 3. Precautions: (1) To the PD wastewater, add 9.5 kg of solid alkali per 100 L, stir thoroughly to separate the crude PD above the water layer, remove the water layer, and collect the crude PD. To 100 L of crude PD, 25 kg of solid alkali is added for drying and water removal. The dried crude PD is then subjected to vacuum distillation (distillation under reduced pressure is preferred); at a vacuum level of < -0.075, the fraction with a temperature above 110°C is collected, while the earlier fractions are discarded. (The moisture content of crude PD must be <0.6% before distillation can proceed). The distilled fraction, after having its content and water content measured, can be reused. ⑵Debromination is stopped by adding water; when layering occurs, if emulsification takes place, the settling time can be extended by 10–20 minutes. IV. Saponification Reaction 1. Names of raw materials, molecular weights, number of moles, and mixing ratios: Dehydrocholic acid ethyl ester, petroleum ether, methanol, potassium hydrochloride (reagent): 65; Methanol for washing (pre-cooling). 2. Procedure: Prepare a solution of potassium hydrochloride in methanol and set it aside for use. Petroleum ether is added to the saponification tank, and after stirring, ethyl hydrotartrate is poured in. The temperature is raised to 35–45°C, and stirring continues for 15 minutes; once it is confirmed that all the materials have dissolved, the prepared potassium hydrochloride methanol solution is added. Stirring continues as the temperature is raised to 55°C ±, and the reaction proceeds for 20 minutes. Hot water is then removed from the jacket, replaced with cooling water, and the temperature is lowered to 0°C. The mixture is allowed to stand and crystallize for more than 3 hours. The material is placed in a centrifuge and spun to remove excess moisture. It is then rinsed with 100 L of pre-cooled methanol in two stages, spun again to remove moisture, and dried under vacuum to yield the final product, dehydrocholesterol. 3. Precautions: (1) After determining the components of the recovered mixed solvent, it can be used so that the final solution has a ratio of petroleum ether to methanol of 50:75 (V/V).