唾液1,5-脱水葡萄糖醇检测方法的建立

Establishment of a method to detect the level of saliva 1,5-anhydroglucitol

目的建立唾液1,5-脱水葡萄糖醇(1,5-AG)的检测方法。方法2017年6月至2018年3月于上海交通大学附属第六人民医院招募正常糖代谢(NGT)者和糖尿病患者共100例(NGT 69例,糖尿病31例,男、女各50例),咀嚼Salivette唾液采集管匹配的棉棒收集唾液。采用酶法检测探究检测唾液1,5-AG样本留取及储存方法。采用质谱法检测唾液1,5-AG水平,分析其在NGT和糖尿病患者之间的差异。组间比较采用t检验及Mann-Whitney U检验,相关性分析采用Spearman相关法。结果(1)唾液采集前刷牙与否不影响唾液1,5-AG水平;1 min咀嚼40~50、50~60和60~70次采集的唾液量差异均无统计学意义(均P>0.05);与采集后立即检测相比,唾液采集后常温或4℃储存2 h检测的唾液1,5-AG水平差异无统计学意义(均P>0.05)。(2)与NGT相比,糖尿病患者唾液1,5-AG[(1.17±0.24)比(0.94±0.19)μg/ml,t=4.084]及血清1,5-AG水平[22.0(17.6,28.5)比4.6(2.8,8.2)μg/ml,Z=-5.421]均显著降低(均P<0.05)。唾液1,5-AG水平与血清1,5-AG水平呈显著正相关(r=0.528,P<0.05),与餐后2 h血糖、糖化血红蛋白、糖化白蛋白均呈显著负相关(分别为r=-0.366、-0.543、-0.444,均P<0.05)。结论唾液采集过程简单,可于常温或4℃短时储存。唾液1,5-AG水平可显著区分NGT人群与糖尿病患者,有望成为评估糖尿病患者短期血糖控制水平的补充指标。

Objective To establish a method to detect the level of saliva 1,5-anhydroglucitol(1,5-AG).Methods From June 2017 to March 2018,100 subjects(50 men and 50 women)with normal glucose tolerance(NGT group,n=69)or diabetes mellitus(DM group,n=31)were enrolled in the Sixth People′s Hospital affiliated to Shanghai Jiao Tong University.All subjects chewed swabs in Salivette saliva collection tubes to collect saliva.Enzymatic assay was used to explore the collection and storage methods for saliva 1,5-AG measurements.The mass spectrometry method was explored to analyze the saliva 1,5-AG levels.Differences were compared between NGT subjects and DM patients.Methods of t test and Mann-Whitney U test were used for comparison between groups,and the Spearman correlation was used for correlation analysis.Results(1)Brushing teeth before saliva collection had no effect on saliva 1,5-AG levels.There was no significant difference in the volume of saliva between chewing 40-50 times,50-60 times,and 60-70 times per minute(all P>0.05).The 1,5-AG levels in saliva stored at room temperature or 4℃for 2 hours were not different from those detected immediately after collection(both P>0.05).(2)Compared with NGT subjects,DM patients had lower saliva 1,5-AG levels[(1.17±0.24)vs(0.94±0.19)μg/ml,t=4.084]and serum 1,5-AG levels[22.0(17.6,28.5)vs 4.6(2.8,8.2)μg/ml,Z=-5.421](both P<0.05).The saliva 1,5-AG levels were positively correlated with serum 1,5-AG levels(r=0.528,P<0.05)and negatively correlated with 2 hour plasma glucose,glycated hemoglobin and glycated albumin(r=-0.366,-0.543,-0.444,respectively;all P<0.05).Conclusions Saliva collection is simple and the sample can be stored at room temperature or 4℃for a short time.Saliva 1,5-AG levels can significantly differentiate DM patients from NGT subjects,which is expected to be a supplementary indicator for evaluating short-term glycemic control in DM patients.