Photoacoustic spectroscopy in a differential Helmholtz resonator has been employed with near-IR and red diode lasers for the detection of CO 2 , H 2 S and O 2 in 1 bar of air/N 2 and natural gas, in static and flow cell measurements. With the red distributed feedback (DFB) diode laser, O 2 can be detected at 764.3 nm with a noise equivalent detection limit of 0.60 mbar (600 ppmv) in 1 bar of air (35-mW laser, 1-s integration), corresponding to a normalised absorption coefficient α = 2.2 × 10 −8 cm −1 W s 1/2 . Within the tuning range of the near-IR DFB diode laser (6357–6378 cm −1 ), CO 2 and H 2 S absorption features can be accessed, with a noise equivalent detection limit of 0.160 mbar (160 ppmv) CO 2 in 1 bar N 2 (30-mW laser, 1-s integration), corresponding to a normalised absorption coefficient α = 8.3 × 10 −9 cm −1 W s 1/2 . Due to stronger absorptions, the noise equivalent detection limit of H 2 S in 1 bar N 2 is 0.022 mbar (22 ppmv) at 1-s integration time. Similar detection limits apply to trace impurities in 1 bar natural gas. Detection limits scale linearly with laser power and with the square root of integration time. At 16-s total measurement time to obtain a spectrum, a noise equivalent detection limit of 40 ppmv CO 2 is obtained after a spectral line fitting procedure, for example. Possible interferences due to weak water and methane absorptions have been discussed and shown to be either negligible or easy to correct. The setup has been used for simultaneous in situ monitoring of O 2 , CO 2 and H 2 S in the cysteine metabolism of microbes ( E . coli ), and for the analysis of CO 2 and H 2 S impurities in natural gas. Due to the inherent signal amplification and noise cancellation, photoacoustic spectroscopy in a differential Helmholtz resonator has a great potential for trace gas analysis, with possible applications including safety monitoring of toxic gases and applications in the biosciences and for natural gas analysis in petrochemistry. Graphical abstract

We introduce and compare two powerful new techniques for headspace gas analysis above bacterial batch cultures by spectroscopy, Raman spectroscopy enhanced in an optical cavity (CERS), and photoacoustic detection in a differential Helmholtz resonator (DHR). Both techniques are able to monitor O2 and CO2 and its isotopomers with excellent sensitivity and time resolution to characterize bacterial growth and metabolism. We discuss and show some of the shortcomings of more conventional optical density (OD) measurements if used on their own without more sophisticated complementary measurements. The spectroscopic measurements can clearly and unambiguously distinguish the main phases of bacterial growth in the two media studied, LB and M9. We demonstrate how 13C isotopic labeling of sugars combined with spectroscopic detection allows the study of bacterial mixed sugar metabolism to establish whether sugars are sequentially or simultaneously metabolized. For E. coli, we have characterized the shift from glucose to lactose metabolism without a classic diauxic lag phase. DHR and CERS are shown to be cost-effective and highly selective analytical tools in the biosciences and in biotechnology, complementing and superseding existing conventional techniques. They also provide new capabilities for mechanistic investigations and show a great deal of promise for use in stable isotope bioassays.

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