LC/MS/MS方法筛查新生儿苯丙酮尿症

苯丙酮尿症 [1～ 4 ] ( PKU )发病原因是患者基因缺陷使肝脏不能合成苯丙氨酸羟化酶而导致体内苯丙氨酸 ( Phe)不能正常代谢为酪氨酸 ( Tyr) ,前者在体内大量堆积并氧化为对人体有害的苯丙酮酸 . PKU是目前筛查范围最广的氨基酸代谢遗传疾病 ,在全世界每年 [5]约有一千万婴儿接受 PKU筛查 ;在我国 ,PKU也是卫生部要求重点筛查的病种 .Chace[5]等在 1 993年报道了 MS/ MS方法筛查新生儿PKU:直接使用 MS/ MS的中性碎片丢失扫描方式检测 Phe和 Tyr,通过氘代内标与待测氨基酸的质谱峰高比来定量 .MS/ MS方法速度快、准确性好、可以…

Newborn Screening of Phenylketonuria by LC/MS/MS

The application of MS/MSin newborn screening is one of the most important branches of MS/MS's current clinical applications. In this paper, we firstly reported a new method of newborn screening for Phenylketonuria(PKU) by coupling LCwith MS/MS, which was relatively cheap, fast, sensitive, and accurate. In this method, only a few steps of sample preparation were required and no isotopically labeled internal standard was required. Phe and Tyr were extracted from dry blood spots with 0.1 mol/L HCl-methanol and reacted with 2.8 mol/L HCl-butanol at 68 ℃ for 35 min. MS/MSwas used as a detector for LCand multi reactions monitoring (MRM) scan mode was employed to determine phenylalanine and tyrosine in blood samples. Among the product ions of Phe and Tyr butyl esters, [M+H-102]⁺was the most abundant in intensity and unique to α-amino acids, therefore the precursor-daughter ion pairs of m/z 222-120 and m/z 238-136 were selected as MRMion pairs. The contents of these two amino acids in blood samples were determined by calculating the areas of their ionic flow peaks. Finally, the molar ratios of Phe to Tyr in blood samples were calculated as the basis of PKUdiagnosis. The curve for Phe showed excellent linearity over the concentration range of 24_1 152 μmol/L(r²=0.999). As to Tyr, the range of linearity was 22_1 056 μmol/L(r²=0.998). The average analytical recoveries of added Phe and Tyr from blood were 98% and 96%, respectively. The limits of detection for Phe and Tyr were respectively 0.5 pg and 2.0 pg , which were 10³times lower than that of reported MS/MSmethod. About 40 blood samples were analyzed by this method, and the results proved that this method could successfully discriminate between normal infants and PKUpositive ones. Each sample's analysis time was about 4 minutes; therefore it was capable for large-scale screening.