# Unstimulated Parotid Saliva Is a Better Method for Blood Glucose Prediction
## 4 Discussion  
Currently, non-invasive detection techniques based on saliva samples are basically targeted at DM patients. In this study, the glucose oxidase method with TBHBA as the chromogen has been improved to be suitable for healthy people. This method was used to compare the saliva glucose concentration of six different saliva collection methods for the first time. This study found that the product has a maximum absorption at a wavelength of 520 nm, the optimal pH is 6.5, and the optimal NaCl concentration is 5 mg/dL. The environmental pH value will change or affect the dissociation state of the enzyme and the substrate to increase or decrease the enzyme activity. Therefore, the maximum activity of the enzyme requires the corresponding optimum pH value. The main catalytic enzymes in the test solution are GOD and POD. The activity of the former was in the range of pH 4.0 to 7.0, and the activity of the latter was in the range of pH 5.0 to 9.0. The optimal pH value of the reaction test solution measured in the experiment was 6.5, which was within the range of GOD activity and POD activity. Therefore, it was theoretically speculated that the pH value of 6.5 was the comprehensive optimal pH value of the same system where the two enzymes were located. Besides, it is found that its linear range, accuracy, and precision can meet the requirements of detection, and more importantly, makes the measurement process more standardized, reduces errors, and is simple and easy to implement. Saliva, like plasma or serum, is a unique and complex body fluid. Sufficient saliva secretion is essential for maintaining oral health. The advantages of saliva assessment include the cost-effectiveness of non-invasive collection and screening of large populations [25]. Saliva is currently considered to be an excellent diagnostic biomarker for human characteristics [26]. In the present study, we found that the unstimulated saliva glucose levels were higher than in stimulated saliva, that unstimulated parotid saliva glucose level was higher than unstimulated sublingual/submandibular saliva, and that unstimulated whole saliva had the lowest level. The blood glucose and unstimulated parotid salivary glucose levels were significantly higher than levels in unstimulated sublingual/submandibular saliva and unstimulated whole saliva, and the glucose levels in parotid saliva were strongly correlated with blood glucose in healthy people. However, in another study, no correlation was found between saliva and plasma glucose levels [27]. Nonetheless, our study revealed a significant, strong correlation between parotid salivary and blood glucose levels but not between  
Figure 6: The correlation between saliva glucose and blood glucose: (**a**) unstimulated parotid saliva (UPS), (**b**) unstimulated sublingual/submandibular saliva (USS), (**c**) unstimulated whole saliva (UWS). mixed salivary and blood glucose levels, unstimulated sublingual/submandibular saliva and blood glucose; the glucose level in parotid saliva, but not that in unstimulated sublingual/submandibular saliva and unstimulated whole saliva, may thus reflect the blood glucose level. The different conclusions are mainly caused by different saliva collection methods. The six saliva collection methods in this study can make up for the shortcomings of existing research and provide the next step for the concentration of glucose in saliva. The determination of saliva has laid a good foundation and pointed out the direction for finding the most suitable method to collect saliva glucose. The determination of saliva glucose concentration is a prerequisite for the development of saliva as a diagnostic and prognostic tool for DM biomarker discovery. In this case, it is important to keep the technical variability caused by sample collection and processing to a minimum so that inter-subject variability in health and disease states can be assessed reproducibly [28]. Single or mixed saliva can be collected.