Frundzhiam V G, Brovko L Iu, Babunova V S, Kartashova V M, Ugarova N N
Lomonosov Moscow State University, Russia.
Prikl Biokhim Mikrobiol. 1999 May-Jun;35(3):358-65.
A rapid (30-35 min) bioluminescence assay of total bacterial contamination (TBC) of raw milk was optimized. This method includes incubation of milk samples in the presence of Neonol-10 and medical purity grade pancreatin with further removal of nonbacterial ATP by filtration through a membrane filter, cell disruption by treatment with dimethyl sulfoxide, and measurement of ATP concentration in a reaction with the bioluminescent reagent Immolum. The TBC detection threshold is 0.5 x 10(5) colony-forming units (CFU) per ml milk. Coefficients of correlation between the standard plate count method and bioluminescence assay (R) and residual standard deviations (Sxy) in raw milk samples (n = 140) were 0.83 and 0.54, respectively. In sterilized milk samples artificially contaminated with pure cultures of the main representatives of milk microflora (coli-forms, Staphylococcus aureus, Streptococcus thermophilus, and Streptococcus group D), these values were 0.89-0.99 and 0.09-0.29, respectively. The specific content of ATP was found to be (0.8 +/- 0.1) x 10(-18) mol/CFU in coli-forms; (12.0 +/- 8.1) x 10(-18) mol/CFU in S. aureus; (35.2 +/- 16.9) x 10(-18) mol/CFU in S. thermophilus; and (42.5 +/- 1.3) x 10(-18) in Streptococcus group D.
一种快速（30 - 35分钟）检测生乳中总细菌污染（TBC）的生物发光测定法得到了优化。该方法包括将牛奶样品在Neonol - 10和医用纯度级别的胰蛋白酶存在下孵育，然后通过膜过滤进一步去除非细菌ATP，用二甲基亚砜处理使细胞破裂，并与生物发光试剂Immolum反应测定ATP浓度。TBC检测阈值为每毫升牛奶0.5×10⁵菌落形成单位（CFU）。生乳样品（n = 140）中，标准平板计数法与生物发光测定法之间的相关系数（R）和剩余标准差（Sxy）分别为0.83和0.54。在人工接种了牛奶微生物主要代表纯培养物（大肠菌群、金黄色葡萄球菌、嗜热链球菌和D群链球菌）的灭菌牛奶样品中，这些值分别为0.89 - 0.99和0.09 - 0.29。发现大肠菌群中ATP的比含量为（0.8 ± 0.1）×10⁻¹⁸摩尔/CFU；金黄色葡萄球菌中为（12.0 ± 8.1）×10⁻¹⁸摩尔/CFU；嗜热链球菌中为（35.2 ± 16.9）×10⁻¹⁸摩尔/CFU；D群链球菌中为（42.5 ± 1.3）×10⁻¹⁸。
