Diabetes mellitus is a widespread human disease with worldwide 346 million persons concerned and an estimated 3.4 million deaths due to high blood glucose level per year [1
]. Currently no treatment exists or is under development which could possibly cure this illness in the near future. The therapy of diabetes mellitus so far consists in monitoring the blood glucose (BG) level of a patient to avoid the danger of hypo- and hyperglycemia and to assist in adjusting the diet and medical treatment. To monitor the blood sugar level as accurate as possible frequent measurements are required. Until today this involves puncturing the fingertip with a lancing device to obtain a drop of blood. The blood sample is placed on a test strip and usually analyzed via an electrochemical reaction. This expensive procedure is uncomfortable especially if frequently performed, it bears the risk of infections and does not represent a continuous measurement technique, which would be ideal for glycemic control [2
]. Hence a non-invasive glucose sensor would greatly increase the quality of life of diabetes patients. Despite intensive research towards a non-invasive glucose monitoring method since more than 25 years still no reliable commercial sensor exists, which circumvents the need of blood sample taking. An overview of the broad research activity and numerous companies involved in this area is given in review articles [3
]. summarizes the vast field of glucose measurement techniques and distinguishes three different categories: invasive, minimal invasive and non-invasive approaches. Optical techniques like polarimetry [7
], Raman spectroscopy [9
], diffuse reflection spectroscopy [11
], absorption spectroscopy [13
], thermal emission spectroscopy [16
], fluorescence spectroscopy [18
] and photoacoustic (PA) spectroscopy [20
] have been used to sense glucose with respect to non-invasive monitoring.
Overview of possible techniques and active research areas for in-vivo glucose measurements.
Most of the optical attempts use near-infrared (NIR) light because it can penetrate up to several mm into human tissue. Unfortunately, glucose absorption in this wavelength region is weak and interferes strongly with other blood and tissue components [23
], which hampered a breakthrough to non-invasive glucose monitoring. Attempts using NIR PA spectroscopy [21
] mainly employ pulsed lasers as excitation sources.
On the contrary, glucose shows strong characteristic absorption and interferes less with other tissue components in the mid-infrared (MIR) spectral region [5
]. However, MIR light only penetrates up to 100 μ
m into human skin due to the strong water absorption [30
]. As a consequence glucose has to be sensed within the interstitial fluid (ISF) of the epidermis since blood capillaries are not reached. Metabolites and proteins diffuse into the ISF on their way from capillaries to cells. This leads to a strong correlation of BG levels and ISF glucose concentration within the physiological range as confirmed in clinical trails [32
]. In the ISF small-to-moderate sized molecules, like glucose (or ethanol), are present in the same proportion as in blood. Hence a frequent calibration with blood measurements is not necessary [33
]. The diffusion process leads to a delayed increase of the glucose concentration in the ISF, which is stated to be between 5 to 15 minutes [13
]. In general the outer skin layers have a greater time delay and smaller glucose concentration maxima. Concerning the correlation for decreasing glucose concentration some uncertainty persists, but there is most likely no time delay due to the high glucose clearance from epidermal ISF [35
]. The outer most layer of the skin, the stratum corneum (SC) - consisting of cell remains (i.e. dead cells) - acts as a barrier to protect the human body from mechanical, chemicals or microbiological impacts from the surrounding [36
]. Moreover the SC is responsible to prevent transepidermal water loss. This layer is only 10 – 20 μ
m thick (except at the sole of foot and the palm, where it can reach up to several mm), has a water content of approximately 10 % [31
] and contains marginal amounts of glucose. Thus in vivo
glucose sensing has to involve skin layers deeper than the SC. For in vitro
studies of blood samples or homogeneous tissue phantoms, the penetration depth is not important since glucose can be detected at the surface. In such experiments glucose concentrations can be readily tracked using MIR light. Some research groups employed a quantum cascade laser (QCL) or a FTIR spectrometer to perform transmission measurements with glucose detection limits of 13.8 mg/dl (in whole blood) [38
], 9.4 mg/dl [39
] and 4 mg/dl [13
] (both in aqueous solution). Unfortunately, these sensitive measurements are hardly convertible to in vivo
sensing and therefore of little benefit to non-invasive glucose sensing. Guo et al. used wavelength-modulated differential laser photothermal radiometry with two QCLs at 9.5 and 10.4 μ
m (i.e., on and off a glucose absorption peak) to measure glucose concentrations (0 – 440 mg/dl) in an homogeneous aqueous phantom [17
]. With a pulsed CO2
laser and a PA detection Christison et al. measured glucose concentrations in aqueous solutions and whole-blood (18 – 450 mg/dl) [40
]. These two approaches have the potential of adapting from a laboratory setup to a small-sized portable sensor in the future. However, both approaches sense glucose concentrations at the sample surface and not in deep epidermal layers as required for in vivo
studies. An attempt to apply MIR PA measurements to in vivo
glucose sensing was recently reported by Lilienfeld-Toal at al. [41
] but could only indicate a qualitative correlation between BG and ISF glucose concentration.
We developed a laser PA detection scheme using an external-cavity quantum cascade laser (tuning range 1010 – 1095 cm−1) as light source and a small-volume PA cell for detection. This setup bears the potential to shrink from a table size apparatus to a handheld device. When investigating epidermal skin samples having a high water content, challenges are the detection of a weak PA signal due to glucose on a strong water background and humidity variations in the PA chamber due to the evaporation of water. To stabilize the conditions in the PA chamber (i.e., to maintain a constant relative humidity) for sensitive measurements, a constant N2 flow is applied to ventilate the cell. Here we report on the performance of this setup by measuring glucose concentration changes through the SC in human epidermis in vitro. This is a significant step compared to former measurements since it demonstrates the tracking of glucose by MIR light not only at the surface but in lower epidermal skin layers. Furthermore by tuning the QCL a spectrum of human epidermis with and without the presence of glucose could be recorded.