Change of Alpha-Amylase Activity in Saliva in Response to Blood Glucose After A High Carbohydrate Ingestion

Salivary α-amylase activity (SAA) may be correlated with blood glucose levels. However, change of SAA in response to blood glucose after carbohydrate diet remains unknown. In 17 healthy young adults, SAA was inversely correlated with blood glucose levels (r = −0.57, P = 0.02) at 1 h after a high carbohydrate ingestion, corresponding to a nadir in salivary α-amylase activity and a peak in blood glucose, suggesting a close link between these factors.


Introduction
Early clinical studies have shown that glucose load in the form of either oral or intravenous route mostly reduces serum and pancreatic amylase, which is greatly manifested around 1h after the load [1][2][3][4][5]. Kayashima et al. [3] and Otsuki et al. [4] found that the obvious reduction in serum (total) and serum pancreatic amylase after glucose load was much prevalent in healthy persons without inflammatory disease such as pancreatitis and hepatitis, although underlying mechanism is unknown [3][4][5]. Some recent studies have reported inverse associations between salivary αamylase activity and glucose homeostasis [6][7][8].
Although serum amylase consists of salivary and pancreatic amylase with almost equal ratio [9], these studies have not revealed the effect of such glucose or starch load on the amylase activity in saliva, which initiates carbohydrate digestion in the oral cavity but the clinical implications is still unclear [10]. For the last decade, in terms of glucose homeostasis and obesity, the clinical relevance of saliva and salivary amylase has been paid much attention than ever before. High salivary amylase, which can be positively associated with high copy number variations in the salivary amylase gene (AMY1) [11,12], appears to be associated with lower risk for diabetes and obesity compared with low copy number variations [7,13,14].
Here, in the light of these backgrounds, I report a preliminary result of single arm test in healthy young Japanese persons that shows an acute effect of a high carbohydrate diet on α-amylase activity in saliva (salivary amylase activity) and a significant inverse relationship between salivary amylase activities and blood glucose levels during the carbohydrate load.

Methods
This study was approved by the Ethics Committee of Josai University, Saitama, Japan. All of the participants provided written informed consent for the study. In November and December 2013, we recruited 20 healthy young nonsmoker university students aged 20-22 years. The subjects had no medical history of cardiometabolic diseases or oral diseases. Anthropometric and laboratory tests were performed in the morning after an overnight fast. The subjects were allowed to drink just one cup of water (< 200 mL) 2 h before the tests. Salivary amylase activity in the oral cavity was measured using the Dry Chemistry System (NIPRO, Osaka, Japan). The procedure, which is described in more detail elsewhere [15,16], takes 30 s and is painless. Blood glucose levels were measured using a glucose meter (ACCU-CHEK Aviva; Roche Diagnostics K.K., Tokyo, Japan). Immediately after baseline measurements, the subjects consumed a high carbohydrate diet, comprising two warm balls of white Nakajima K. Change of Alpha-Amylase Activity in Saliva in Response to Blood Glucose After A High Carbohydrate Ingestion. Diabetes Obes Int J 2017, 2(2): 000150. Copyright© Nakajima K. rice (200 g; energy 295 kcal, carbohydrate 68 g, lipid 0 g, protein 4.2 g, trace minerals and vitamins, and water) with salt (0.8 g) without drinking any fluid. The rice balls were prepared in our laboratory by the stuff members. Salivary amylase activity and blood glucose levels were measured at baseline (i.e., in the fasting state) and at 15, 30, 60, 120, and 180 min after the carbohydrate ingestion.
Data are expressed as the mean ± standard deviation and standard error in the table and figure, respectively. Changes in salivary amylase activity and blood glucose were assessed by repeated-measures analysis of variance. Post hoc analysis was conducted by Bonferroni test. Correlations between clinical variables were examined using Pearson correlation coefficients. Statistical analyses were performed using Statview version 5.0 (SAS Institute, Cary, NC, USA). Values of P < 0.05 were considered statistically significant (Bonferroni test, P < 0.0033).

Results and Discussion
The 17 subjects completed the carbohydrate load test, although salivary amylase at 30 and 180 min was only measured in 14 subjects. (Table 1) shows the clinical characteristics of subjects at baseline. The baseline mean salivary amylase activity was consistent with the results of previous studies that measured salivary amylase using the same method [16,17].
As shown in (Figure 1) blood glucose levels increased for the first 60 min after starch ingestion and gradually decreased thereafter. By contrast, salivary amylase activity decreased reaching a nadir at 60 min (not statistically significant, P = 0.009, Bonferroni test) with the smallest standard error, but then increased thereafter. The correlation coefficients between salivary amylase activity and blood glucose levels were −0.33 (P = 0.19), −0.33 (P = 0.19), −0.21 (P = 0.48; n = 14), −0.57 (P = 0.02), 0.10 (P = 0.68), and −0.11 (P = 0.70; n = 14) at baseline, 15, 30, 60, 120, and 180 min, respectively. Therefore, salivary amylase activity was significantly and inversely correlated with blood glucose levels at 60 min, corresponding a peak in blood glucose and a nadir in salivary amylase activity. Intriguingly, the blood glucose level at 15 min was significantly and inversely correlated with the salivary amylase activity at 30 min (r = −0.72, P = 0.004, n = 14). Values are expressed as the mean and standard error. Repeated-measures analysis of variance and post hoc test did not reveal a clear reduction in salivary amylase activity until and at 60 min (P = 0.06 and P = 0.009, respectively), using data obtained at baseline, 15 min, and 60 min. *Blood glucose levels and salivary αamylase activity were measured in 14 subjects at 30 and 180 min.
Current study demonstrates that α-amylase activity in saliva may decrease around 1h after a carbohydrate diet in healthy persons, which is not inconsistent with previous studies that shows the reduction in serum and pancreatic amylase after glucose load [1][2][3][4][5]. Minimum insulin is considered to be necessary for the production of pancreatic enzymes via insulin receptors [10,18,19] and impaired insulin action due to insulin deficient and/or insulin resistance can cause reduced pancreatic Nakajima K. Change of Alpha-Amylase Activity in Saliva in Response to Blood Glucose After A High Carbohydrate Ingestion. Diabetes Obes Int J 2017, 2(2): 000150. Copyright© Nakajima K. 3 amylase. Current results suggest that transient high blood glucose, even within the normal response after a meal, can also reduce salivary amylase irrespective of intact insulin action and that such plausible tight mechanism may be applicable to not only pancreatic but also salivary amylase.
In this study, during the first 30 min, a change in blood glucose level (at 15 min) likely proceeded to a change in salivary α-amylase activity. It is noteworthy that the significant correlation between salivary amylase and blood glucose was observed at 1 h after carbohydrate ingestion, but not at fasting, suggesting a possibility that multiple factors including stress or hungry may affect the level of serum salivary amylase in the fasting state [16,17,20], but once blood glucose is elevated, serum salivary amylase level might be restricted in response to the individual's level of blood glucose.
Taken together, these preliminary findings suggest that salivary α-amylase activity is linked to glucose homeostasis possibly through insulin receptors and the sensitivity for blood glucose concentration in the salivary gland. Therefore, although the baseline salivary amylase level may be primarily determined by the AMY1 gene [11,12], other factors, including glucose homeostasis or dietary factors, might influence the fluctuations of pancreatic and salivary α-amylase activity.
In this study, the effects of proteins or lipids on salivary α-amylase were not examined because of a preliminary study. Otsuki et al. [4] found that serum amylase increased 2h after a high protein diet without changes in blood glucose level in 35 healthy adults aged 26-40 years. Then, although high protein diet is unlikely to elicit reduced salivary amylase, future studies is needed to warrant the effects of kinds of amino acids, lipids, or other nutrients such vitamins and minerals on salivary α-amylase.
In conclusion, this study demonstrates a possible reduction in salivary α-amylase activity after carbohydrate ingestion that was correlated with simultaneous changes in blood glucose levels, suggesting a close link between the activity level of salivary amylase and glucose homeostasis.