Sterilization steam bath

Agar Culture Medium Add 6.0 g of agar to 500 mL of Liquid Culture Medium, and heat with stirring on a steam bath until the agar dissolves. Add approximately 10-mL portions of the hot solution to test tubes, cover to prevent contamination, sterilize by heating in an autoclave at 121 ° for 15 min, and cool tubes in an upright position to keep color formation to a minimum. Store at 10° in the dark. 

A solution of 10 grams of d-glucoheptonic acid lactone in 50 ml of distilled water is warmed on a steam bath for about 2 hours to hydrolyze the lactone to the acid. The mixture is cooled and 100 ml of 95% ethanol are added. To the solution of glucoheptonic acid are added about 37 grams of erythromycin and the volume of the reaction mixture is brought to 200 ml by the addition of 95% ethanol. The reaction mixture is stirred for about 2 hours and is filtered through a porcelain filter candle of porosity 02. To provide a sterile product, aseptic technique is used throughout the remainder of the procedure. 

The components listed above are heated in a flask for 20 minutes in a steam bath, filtered, cooled down and then made up to 1000 ml with demineralized water. 10 ml quantities of the broth are poured into test tubes and then sterilized in stages (3 times 20 minutes) on a steam bath. 

The stated quantities of the components are dissolved in the water by heating in a steam bath. The pH is then set to 7.2. The culture medium is either sterilized in stages (heating three times for 20 minutes in a steam bath with intervals of 24 hours between each heating process) or in an autoclave for 20 minutes at 121 C. The culture medium is then poured out into Petri dishes. 

The specialist trade also supplies dry culture media in powder form, which contain all the components which are necessary for the cultivation of microorganisms on the particular culture medium. In accordance with the recipes which are given, a definite quantity of the powder must be weighed out. A specified quantity of demineralized water, tap water or the water which is to be analyzed is then poured over this powder. This is then generally dissolved with heating and the pH is set to the desired value. After the solution has been put into test tubes or flasks, it is sterilized either in an autoclave or by sterilization in stages in a steam bath. It is always to... 

The steam bath is usually used for sterilizing sensitive culture media... 

Glucose and nutrient salts are dissolved separately in 500 ml redistilled water, sterilized for 30 minutes in steam bath and the separate solutions combined after cooling. 

Accuracy of thermocouples should be 0.5°C. Temperature accuracy is especially important in steam sterilization validation because an error of just 0.1 °C in temperature measured by a faulty thermocouple will produce a 2.3% error in the calculated F0 value. Thermocouple accuracy is determined using National Bureau of Standards (NBS) traceable constant temperature calibration instruments such as those shown in Figure 6. Thermocouples should be calibrated before and after a validation experiment at two temperatures 0°C and 125°C. The newer temperature-recording devices are capable of automatically correcting temperature or slight errors in the thermocouple calibration. Any thermocouple that senses a temperature of more than 0.5°C away from the calibration temperature bath should be discarded. Stricter limits (i.e., <0.5°C) may be imposed according to the user s experience and expectations. Temperature recorders should be capable of printing temperature data in 0.1 °C increments. 

Equipment where necessary should be properly sterilized before use, preferably by pressurized dry steam. Other methods can be accepted if steam sterilization cannot be used because of the nature of the equipment It is important not to overlook such individual items as bench centrifuges and water baths. 

Reuss and Smith (1965) added NaN02 to a Bath soil (pH 4.6) and a Walla Walla soil (pH 5.6) and observed that after 24 h about 25% of the nitrogen had been oxidized to nitrate in the more acid soil and 14% in the Walla Walla soil. Experiments in steam-sterilized and non-sterilized soils gave identical results. 

Sterilization of the surfaces of vessels, pipes and valves may be achieved by heat, radiation or chemicals [81]. The use of steam has already been mentioned apart from cost it tends to be a slow procedure for sterilizing brewery vessels when steam pressures are low. Furthermore, it may carry undesirable particles and odours while the condensate is often drained unsatisfactorily. Radiation sterilization is rare although ultraviolet light irradiation is used for the treatment of water on a continuous basis. One of the simplest chemicals used for sterilization is ozone but this has proved corrosive [82]. More widely employed is hydrogen peroxide which, with peracetic acid, is added to soak baths for flexible pipes and items of fermentation equipment. 

Purified water is water that has been purified from the hardness minerals by one of the methods mentioned by Ph. Eur. Ph. Eur. has two types of Purified Water Purified Water in bulk and Purified Water in containers. It is used in the pharmacy in several ways as a raw material for the manufacture of non-sterile medicines, sterile medicines that are not necessarily free of endotoxins, in water baths, as cooling water for steam sterilisers, and sometimes for rinsing glassware or packaging material. Eor Highly purified water Ph. Eur. the microbiological and endotoxin requirements are the same as for Water for injections. 

Dispense 120 ml. volumes into 250 mL conical flasks and add 2 g. of agar to each. Plug the flarits and autoclave them at 120°C. for 15 minutes. The sterile medium is stored at 4°C. and has a storage life of 4 to 6 weeks. When required for use, the flasks are steamed for 30 to 40 minutes to melt the agar, and then 10 ml. of glucose solution is added to each. The glucose is sterilized separately as a 13% w/v solution for 15 minutes at 120°C. The molten medium is allowed to stand in a water bath at a temperature of 46-48°C. 

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