1. Introduction {#sec1-sensors-17-02784}
===============

The carbon cycle in the ocean is a dynamic component of the global carbon budget, but the diverse sources and carbon sinks as well as their complex interactions in the ocean remain poorly understood \[[@B1-sensors-17-02784],[@B2-sensors-17-02784]\]. The dissolved inorganic carbon (DIC) in the ocean is the sum of HCO~3~^−^, CO~3~^2−^, H~2~CO~3~ and CO~2~ \[[@B3-sensors-17-02784]\]. The concentration of DIC is about 2.30 mmol/L, with ${HCO}_{3}^{–}$ (about 1.95 mmol/L) making up the majority of it \[[@B4-sensors-17-02784]\]. Therefore, bicarbonate is a key factor in carbon cycle research. The modern methods for oceanic carbonate system measurement are mainly involved in total carbon and CO~2~ partial pressure (pCO~2~) investigations \[[@B5-sensors-17-02784],[@B6-sensors-17-02784]\]. However, the direct detection of HCO~3~^−^ and CO~3~^2−^ in seawater it is still a huge challenge due to their low concentrations. Raman spectroscopy is potentially capable of overcoming this challenge on the condition of a significant ensitivity enhancement \[[@B7-sensors-17-02784]\].

In 2004, scientists reported the first attempt to detect HCO~3~^−^ in water solutions directly by using Raman spectroscopy \[[@B8-sensors-17-02784]\], with a limit of detection (LOD) of 7.5 mmol/L after an integration time of 20 min. They had also tried to enhance the HCO~3~^−^ signal with an introduced liquid core waveguide \[[@B9-sensors-17-02784]\], eventually obtaining an amplification ratio of 7.8× in the peak intensity. All those efforts still do not make the direct Raman detection of HCO~3~^−^ in seawater practical due to the fact its concentration is as low as 2 mmol/L. The strong background signal resulting from SO~4~^2−^, which has a concentration of \~28 mmol/L in seawater \[[@B10-sensors-17-02784]\], makes direct detection of HCO~3~^−^ even more difficult. With a similar waveguide-based enhancement mechanism, the multi-pass cavity concept has been presented as an effective approach in gas Raman detection with desirable enhancement \[[@B11-sensors-17-02784],[@B12-sensors-17-02784],[@B13-sensors-17-02784],[@B14-sensors-17-02784]\]. Taking such an approach in direct seawater Raman detection is the motivation for the work presented in this article. A near-concentric cavity-enhanced Raman spectroscopy system was specially built for liquid sample detection. With the aid of this system, direct HCO~3~^−^ Raman detection in seawater becomes readily feasible as expected.

2. Experiment Setup {#sec2-sensors-17-02784}
===================

As shown in [Figure 1](#sensors-17-02784-f001){ref-type="fig"}, the near-concentric cavity-enhanced Raman spectroscopy system is adapted on the basis of a formerly reported design \[[@B14-sensors-17-02784]\]. In order to detect liquid samples, a sample cell of fused silica with 10 mm × 10 mm × 40 mm inner size and 1 mm wall thickness is used and placed at the center of the cavity. In this system, a diode-pumped, frequency-doubled Nd:YAG CW laser (532 nm) with a power of 300 mW is used as the Raman excitation laser source. The polarization of the laser beam is rotated by 90° through passing a half wave plate (HWP), and then the laser beam is compressed and focused into the chamber by a couple of lens (a planoconvex lens with a focal length of 100 mm and a planoconcave lens with a focal length of −75 mm). The near concentric cavity is composed of two identical spherical mirrors M1 with 25.4 mm diameter and 25 mm focal length. The reflectivity of these two mirrors is over 99% from 500 to 700 nm. These two mirrors are spaced face to face at a distance of 104.2 mm, and the mirror M1-2 is clockwise rotated about 0.04° to create the near concentric cavity reflection mode. Because of the strong absorption of water, the laser beam number in the cavity is 18, which is less than the number in the cavity of a gas sample cell. The scattering signal from the center of cavity is collected by an achromatic doublet lens L1 (f = 30 mm) with a diameter of 25.4 mm. A mirror M2 (f = 12.5 mm), placed on the opposite side beyond the sample chamber, is used to enhance the collection efficiency of the signal. Another lens L2 (f = 30 mm) is used to couple the Raman signal into the delivery optical fiber bundle (19 μm × 100 μm, NA = 0.22), with a long pass filter placed on its front to get rid of Rayleigh scattering from the collected signal. From the other end of the fiber, the signal is then coupled into a spectrometer (Acton SP2500i, Princeton Instruments, Trenton, NJ, USA), with 1200 g/mm grating and 10 μm entrance slit width. The Raman spectra are recorded by a CCD detector (SPEC-10: 400B, Princeton Instruments, Trenton, NJ, USA) with a 1340 × 400 imaging array and 20 μm × 20 μm pixel size operating at −40 °C. The resulted spectral range is 800\~2000 cm^−1^ with a resolution of 2 cm^−1^. To get a relative desirable Raman signal and prevent the CCD from overexposure, each spectrum is accumulated five times with an integration time of 10 s, so the total acquisition time is 50 s for one sample. Five measurements are taken for each sample to evaluate the repeatability.

3. Results {#sec3-sensors-17-02784}
==========

The reported LOD for SO~4~^2−^ by using a liquid core optical fiber Raman system is about 1.5 mmol/L \[[@B15-sensors-17-02784]\]. As a comparison, we prepared 15 Na~2~SO~4~ solutions (0.10 mmol/L--2.00 mmol/L) to evaluate the LOD of the NC-CERS system. The LOD is defined as the ratio of three times the noise intensity (σ) to the slope of calculation curve. The spectra of four solutions are shown in [Figure 2](#sensors-17-02784-f002){ref-type="fig"}a, and the Raman signal of SO~4~^2−^ can be observed clearly in the spectrum of the 0.10 mmol/L solution. Given this, the NC-CERS system has a LOD less than 0.10 mmol/L for SO~4~^2−^ which is 15 times lower than that reported before. The linear relationships between concentrations and peak intensity of the SO~4~^2−^ are shown in [Figure 2](#sensors-17-02784-f002){ref-type="fig"}b and the response of the NC-CERS system for liquid detection has good linearity with R^2^ = 0.996 over the whole range.

To evaluate its performance for HCO~3~^−^, we prepared nine NaHCO~3~ solutions with concentrations ranging from 0.40 mmol/L to 4.00 mmol/L. The corresponding concentrations of HCO~3~^−^ are from 0.37 mmol/L to 3.79 mmol/L calculated by using the carbon balance mode. All of the concentrations of HCO~3~^−^ are corrected by using the equilibrium mode of carbon components in water. [Figure 3](#sensors-17-02784-f003){ref-type="fig"}a shows the spectra of five of these concentrations. The peak intensity of the HCO~3~^−^ Raman signal increases as a function of the concentration, and the Raman signal of HCO~3~^−^ can be distinguished in the solution with 0.37 mmol/L NaHCO~3~. It is proved that the LOD of the NC-CERS system for HCO~3~^−^ is less than 0.37 mmol/L, which is much lower than its concentration in seawater. The linear relationship between concentrations and peak intensities of the HCO~3~^−^ is shown in [Figure 3](#sensors-17-02784-f003){ref-type="fig"}b and it shows good linearity with R^2^ = 0.978.

Furthermore, we prepared simulated seawater with fixed a NaSO~4~ concentration (28.00 mmol/L) and different NaHCO~3~ concentrations (0.00, 2.00, 4.00 mmol/L) to evaluate its ability to quantify HCO~3~^−^ in seawater. The corresponding concentrations of HCO~3~^−^ are 0.00, 1.93 and 3.79 mmol/L. The corresponding spectra are shown in [Figure 3](#sensors-17-02784-f003){ref-type="fig"}c, and the detailed spectra are shown in [Figure 3](#sensors-17-02784-f003){ref-type="fig"}d. Compared with the intensity of SO~4~^2−^, the intensity of the HCO~3~^−^ signal is much weaker. Even so, the Raman signal of HCO~3~^−^ can be detected in the spectrum of the mixed solution with 1.93 mmol/L NaHCO~3~. This shows the ability of the NC-CERS system for direct HCO~3~^−^ detection in seawater, but the strong and adjacent SO~4~^2−^ signal of seawater has an impact on the quantitative analysis of HCO~3~^−^.

In order to reduce this impact, we developed an adaptive signal extraction method for HCO~3~^−^ and the process is shown in [Figure 4](#sensors-17-02784-f004){ref-type="fig"}a--c. The spectrum is obtained from a mixed solution with 28.00 mmol/L Na~2~SO~4~ and 1.93 mmol/L NaHCO~3~. In this spectrum, the Raman peaks of SO~4~^2−^ (981 cm^−1^) and H~2~O (1640 cm^−1^) can be clearly recognized, while the peak of HCO~3~^−^ (1021 cm^−1^) is almost submerged in the background. Given this, the original spectrum is normalized according to the peak intensity of H~2~O to reduce the impact of laser power fluctuation as shown in [Figure 4](#sensors-17-02784-f004){ref-type="fig"}a. The next step is baseline correction. We have compared four baseline fitting results with four methods (Voigt, Gaussian, Lorentzian and bi-exponential fitting methods), and the polynomial function shows the best performance among them. The baseline corrected result is shown as a blue line in [Figure 4](#sensors-17-02784-f004){ref-type="fig"}b. The last step is the weak HCO~3~^−^ signal extraction from the neighboring strong SO~4~^2−^ signal. The right wing of the SO~4~^2−^ signal is regarded as the baseline of the HCO~3~^−^ signal and is fitted by using a bi-exponential function in the region of 1000--1050 cm^−1^. We again tried four baseline fitting methods (Voigt, Gaussian, Lorentzian and bi-exponential fitting methods), and the polynomial function showed the best performance among them. The fitted baseline is shown as the green dotted line, and the corrected spectrum is shown as the black circle dots in [Figure 4](#sensors-17-02784-f004){ref-type="fig"}c. The Raman peak of HCO~3~^−^ is extracted and fitted by using a Gaussian function which is shown as the red line in [Figure 4](#sensors-17-02784-f004){ref-type="fig"}c.

Based on this data processing method, the Raman signals of HCO~3~^−^ in simulated seawater solutions are extracted and shown in [Figure 4](#sensors-17-02784-f004){ref-type="fig"}d. The signal of a deep seawater sample is also shown as the cyan line in [Figure 4](#sensors-17-02784-f004){ref-type="fig"}d. The linear relationship between concentrations and peak intensities of fitted HCO~3~^−^ is shown in [Figure 4](#sensors-17-02784-f004){ref-type="fig"}e and it shows high linearity with R^2^ = 0.951 over the range. We chose a mixed solution with 28.00 mmol/L NaSO~4~ and 1.50 mmol/L NaHCO~3~ as a blind sample. The test result is 1.35 mmol/L with 5.0% relative error, which is shown as the pink point in [Figure 4](#sensors-17-02784-f004){ref-type="fig"}e. Furthermore, we tested deep seawater, knowing the concentration of HCO~3~^−^ is 1.91 mmol/L which is shown as a blue point in [Figure 4](#sensors-17-02784-f004){ref-type="fig"}e. Compared with the reported HCO~3~^−^ concentration value of 1.95 mmol/L in seawater \[[@B4-sensors-17-02784]\], the relative error was about 2.1%.

4. Conclusions {#sec4-sensors-17-02784}
==============

In this paper, we developed a near-concentric cavity-enhanced Raman spectroscopy system for liquid sample detection based on a near-concentric cavity and directly detected bicarbonate in seawater for the first time. Systematic experiments have been carried out for system evaluation. As shown by the NC-CERS system tests, the LOD for SO~4~^2−^ is less than 0.10 mmol/L, which is 15 times lower than the result reported before. The LOD for ${HCO}_{3}^{–}$ is less than 0.37 mmol/L, which is much lower than its concentration in seawater (1.95 mmol/L). The detection performance of the NC-CERS system for HCO~3~^−^ in seawater is evaluated by analyzing mixed solutions of fixed 28.00 mmol/L NaSO~4~ and different NaHCO~3~ concentrations. The Raman signal of 1.93 mmol/L HCO~3~^−^ in mixed solution can be clearly detected, which shows the ability of direct HCO~3~^−^ detection in seawater. To further realize the quantitative analysis for HCO~3~^−^, a specially developed data processing scheme is introduced. With this data processing method, the weak HCO~3~^−^ signal is extracted from the neighboring strong SO~4~^2−^ signal, hence a quantitative analysis with R^2^ = 0.951 is made possible. Based on this calibration curve, the HCO~3~^−^ concentrations of a blind sample and a seawater sample are calculated. The calculated values are 1.43 mmol/L and 1.91 mmol/L with 4.9% and 2.1% relative error, respectively. It is hoped that this NC-CERS system could be fully developed for field ocean observations in the near future. CO~3~^2−^ with a concentration down to 1/10 of the HCO~3~^−^ in seawater will be the next detection object.

This work was supported by National Natural Science Foundation of China (61575181), Fundamental Research Funds for the Central Universities (201562008), National Key Research and Development Program of China (2016YFC0302101), the Strategic Priority Research Program, CAS (XDA11040301). The author (DewangYang) would like to appreciate Zhao Luo and Tengyue Wang's language revision.

Jinjia Guo and Ronger Zheng initiated and supervised the research project. Dewang Yang and Jinjia Guo proposed the idea, conceived and designed the experiments. Dewang Yang, Jinjia Guo, Chunhao Liu and Qingsheng Liu performed the experiments. Dewang Yang processed the data and wrote the initial draft. Ronger Zheng revised the manuscript and figures. The final revisions and writing were done by Jinjia Guo.

The authors declare no conflict of interest.

![Schematic of the near-concentric cavity-enhanced Raman spectroscopy system for liquid sample detection.](sensors-17-02784-g001){#sensors-17-02784-f001}

![Raman spectra of a range of Na~2~SO~4~ solution (0.10 mmol/L to 2.00 mmol/L). (**a**) Four spectra of typical concentration. (**b**) The linear relationship between concentrations and peak intensities of the SO~4~^2−^ signal.](sensors-17-02784-g002){#sensors-17-02784-f002}

![The detection ability of the NC-CERS system for HCO~3~^−^. (**a**) The spectra of 5 NaHCO~3~ aqueous solutions with different concentrations. (**b**) The linear relationship between concentrations and peak intensities of the HCO~3~^−^ signal. (**c**) The Raman spectra of three mixed solutions which contains fixed NaSO~4~ concentration (28.00 mmol/L) and different NaHCO~3~ concentrations. (**d**) The detailed spectra (1000--1050 cm^−1^) of mixed solutions.](sensors-17-02784-g003){#sensors-17-02784-f003}

![The signal extraction method for quantitative analysis of HCO~3~^−^. (**a**) The normalized spectrum of mixed solution with 28.00 mmol/L Na~2~SO~4~ and 1.93 mmol/L NaHCO~3~. (**b**) Baseline correction. The red dotted line represents the baseline curve fitted by a polynomial function, and the blue line is the baseline corrected curve. (**c**) The signal extraction method for HCO~3~^−^. The green dotted line represents the baseline B fitted by a double exponential function. The extracted signal is shown as black circle dots and fitted by using a Gaussian function (red line). (**d**) Fitted HCO~3~^−^ Raman signals of 1000 m depth seawater and four mixed solutions with fixed 28.00 mmol/L Na~2~SO~4~ and different concentrations of NaHCO~3~. (**e**) The calibration curve of HCO~3~^−^. The pink point is the result of a blind sample, and the blue point is the result of a seawater sample.](sensors-17-02784-g004){#sensors-17-02784-f004}
