嗜高渗透压酵母霉菌

1. SCOPE适用范围
   1. This method is applicable to all materials, which contain a high sugar (> 40 ^o Brix ) content, where an estimate of the total Osmophilic yeast and mold count is desired.

1.1这种方法适用于所有高糖度的原料(> 40 ^o Brix )，可以评估嗜高渗压酵母菌和霉菌总数。

2. APPARATUS设备仪器
   1. Sterile sampling devices                                       无菌取样装置
   2. Autoclavable 150 ml screw cap bottle                  150 ml耐高温的带有螺帽的瓶子
   3. Autoclavable Erlenmeyer flasks                            耐高温的锥形瓶
   4. Sterile Petri dishes (150mm)                                 灭菌培养皿 (150mm)
   5. Sterile 10 ml pipettes                                             灭菌10 ml移液管
   6. Bunsen burner                                                       煤气灯
   7. Incubator set at 25 ^o C                                           25 ^o C培养箱
   8. Constant temperature water bath at 45-47 ^o C       45-47 ^o C 的恒温水浴锅
   9. Autoclave                                                               高压灭菌器
   10. Balance: to 1000 grams capacity                          可称量1000克的天平
   11. Stirrer/ Hot Plate, stir bar                                       搅拌器/轻便电炉，搅拌子
3. MEDIA
   1. 40% Sucrose Malt Extract Agar                            40%的蔗糖麦芽浸出物琼脂
   2. Prepare according to the following formula
      1. Malt Extract       麦芽膏30 grams
      2. Mycological Peptone  蛋白胨  5 grams
      3. Agar                    琼脂粉15 grams
      4. Sucrose       蔗糖400 grams
      5. Distilled Water       蒸馏水       1 liter
      6. pH5.4
   3. Alternately, dissolve 400 grams of sucrose and 50 grams of Oxoid Malt Extract Agar in 1 liter of distilled water.

3.3    也可以选择溶解400克的蔗糖和50克的Oxoid公司生产的麦芽精琼脂1升蒸馏水中。

1. Dispense into autoclaving autoclavable Erlenmeyer flask and sterilize by autoclaving at 121 ^o C for 10 minutes. Cool in water bath prior to using.
2. 3.4分装到锥形瓶高压灭菌121 ^o C 10分钟在水浴中冷却待用。

1. Buffered Peptone Water Supplemented with Glucose用葡萄糖补充到缓冲蛋白胨水中
2. Prepare according to the following formula:
   1. Peptone蛋白胨10 grams
   2. Sodium Chlori     氯化钠  5 grams
   3. Disodium Phosphate    磷酸氢二钠  3.5 grams
   4. Potassium Dihydrogen Phosphate磷酸二氢钾1.5 grams
   5. Glucose葡萄糖400 grams
   6. Distilled Water蒸馏水    1 liter
3. Alternatively, dissolve 400 grams of glucose and 20 grams of Oxoid Buffered Peptone Water (CM509) in 1 liter of distilled water.

4.3       也可以选择溶解400克的葡萄糖和20克的Oxoid公司生产的缓冲蛋白胨水(CM509)在1升蒸馏水中。

1. Dispense 94 mls into 150-ml screw cap bottles.

4.4       分装94ml到150-ml的带有螺帽的瓶子中。

1. Cap and sterilize by autoclaving for 15 minutes at 121 ^o C.

4.5      121 ^o C灭菌15分钟。

1. Cool to ambient temperature prior to use. After cooling, this yields 90 ml of Diluent in each bottle.

4.6在室温中冷却。冷却后每个瓶子里有90 ml稀释液。

5. PREPARATION OF STANDARDS
   1. None.
6. PREPARATION OF SAMPLES
   1. Use aseptic technique. Sample at least 20 grams of test material into a sterile container.

6.1      灭菌容器中最少有20克的样品。

INSTRUMENTAL CONDITIONS

1. None

1. None

1. Aseptically pipette 10 grams if test material into 90 mls of Diluent
1.      用灭菌移液管将样品10克加到稀释液中。
2. Shake vigorously to mix.

8.2       剧烈振摇使其混合。

1. Pipette 10 mls into each of the two 150-mm petri dishes.

8.3       吸取10 ml到150-mm的平皿中，并做平行样。

1. Pour 35-40 mls of molten 40% Sucrose Malt Extract Agar into each petri dish and swirl to mix.

8.4    倾倒35-40 ml的40%的蔗糖麦芽精培养基到平皿中，旋转摇匀。

1. After the agar has solidified, invert the petri dish and place in a 25 ^o C incubator for 5 days

8.5       等到琼脂凝固后，翻转平皿，放入25 ^o C培养箱中培养5天。

1. Count the colonies on both dishes and calculate the average to yield the number of CFU's per gram of test material.

8.6       计数两个平皿上的菌落并且计算每克样品中所含有的菌落数。

9. CALCULATIONS
   1. None
10. SAMPLE CALCULATIONS
    1. None
11. REPORTING RESULTS
    1. Record results and compare to specification.