Continuous Delivery of Hyperpolarized Xenon-129 Gas Using a “Stopped-Flow” Clinical-Scale Cryogen-Free Hyperpolarizer

Abstract

In 2022, the FDA approved hyperpolarized (HP) ¹²⁹Xe gas as an inhalable contrast agent for functional lung imaging. For clinical imaging, HP ¹²⁹Xe is usually given as a bolus inhalation. However, for preclinical applications (e.g., pulmonary imaging in small rodents), continuous delivery of HP ¹²⁹Xe is greatly desired to enable MRI scanning under conditions of physiological continuous animal breathing patterns. Moreover, HP ¹²⁹Xe gas can be utilized for other applications including materials science and bioanalytical chemistry, where continuous flow of hyperpolarized gas through an NMR sample over several minutes is also desired for sensing of ¹²⁹Xe inside an NMR spectrometer. ¹²⁹Xe is often hyperpolarized using continuous-flow spin exchange optical pumping, which employs a lean (1-2%) mixture of Xe and a carrier gas (e.g., He and N₂). The low Xe concentration in the produced output reduces the NMR detection sensitivity, and thus Xe cryo-collection is typically employed to achieve near-100% pure gas-phase Xe before administration to the sample or subject. However, the need for cryo-collection undermines a key advantage of continuous-flow production, i.e., the continuously flowing in a hyperpolarizer HP ¹²⁹Xe gas is trapped inside the hyperpolarizer, and the produced HP ¹²⁹Xe gas is released at once when the production cycle (30-60 mins) is completed. An alternative HP ¹²⁹Xe production technology employs a “stopped-flow” approach, where a batch of HP gas is hyperpolarized over time and quickly released from a hyperpolarizer. Here, a clinical-scale “stopped-flow” ¹²⁹Xe hyperpolarizer was employed to hyperpolarize a 1.3 liter-atm batch of 50:50 Xe:N₂ gas mixture inside a glass cell with an ultra-long lifetime of the HP ¹²⁹Xe state (T₁ > 2 hours). The produced HP ¹²⁹Xe gas was slowly delivered into a 5-mm NMR tube via PEEK tubing under a wide range of gas flow rates: 3-180 standard cubic centimeters per minute (sccm). The polarization of the gas ejected from the hyperpolarizer was quantified using in situ low-field NMR polarimetry and additionally verified using a 0.35 T clinical MRI scanner. Continuous-flow delivery of HP ¹²⁹Xe was demonstrated for up to 15 minutes with a gas flow rate of 45-150 sccm over a 2.5-meter length of PEEK tubing, suffering only small losses in ¹²⁹Xe polarization. These observations are additionally supported by ¹²⁹Xe relaxation measurements inside the PEEK tubing employed for gas delivery and the 5-mm NMR tube employed for polarimetry. ¹²⁹Xe polarization of 16-19% was obtained in the delivered gas, starting with an “in-polarizer” ¹²⁹Xe polarization of 19%. We envision that this method can be employed for on-demand cryogen-free delivery of hyperpolarized gas using “stopped-flow” ¹²⁹Xe hyperpolarizers for a broad range of applications, from pre-clinical imaging, to biosensors, to spectroscopy of materials surfaces.

Graphical Abstract

INTRODUCTION

Nuclear spin polarization (P), or the degree of nuclear spin alignment with applied magnetic field, can be significantly enhanced beyond its thermal equilibrium level using NMR hyperpolarization.^1-2 Since the NMR signal is directly proportional to P,³ the orders-of-magnitude polarization gain obtained through hyperpolarization results in correspondingly substantial gains in NMR and MRI detection sensitivity.^{1, 4-5}

Hyperpolarized (HP) NMR-active noble gases such as ³He, ⁸³Kr, and ¹²⁹Xe can be utilized as inhalable MRI contrast agents to assess lung function,^6-13 probe the metabolic status of brown fat,¹⁴ image the brain,¹⁵ and perform hyperpolarized Chemical Exchange Saturation Transfer (hyper-CEST)^16-21 for sensing²² and imaging the interactions of receptors and drugs.^23-25

HP ¹²⁹Xe gas was recently approved by the FDA as a safe contrast agent for lung imaging in December 2022, making it the leading HP contrast agent of any kind, and certainly the key inhalable HP gas contrast agent.^26-30 In this new functional imaging modality, the HP ¹²⁹Xe gas is typically inhaled to enable in vivo administration³⁰; the corresponding MRI scan is thus recorded after inhalation—typically on a single patient breath hold. Another clinical MRI scanning approach is to perform MR image acquisition during multiple subsequent inhalations of HP ¹²⁹Xe gas (albeit with lower volumes of HP ¹²⁹Xe gas administered during each inhalation). This approach offers a number of advantages including providing more detailed gas dynamics information and lower Xe dose per individual inhalation; moreover, free-breathing administration facilitates subject (or patient) participation and is more natural—potentially providing a better reflection of physiological conditions. However, unlike a single-dose protocol, multi-breath protocols require continuous delivery of HP ¹²⁹Xe gas to a patient or subject. Finally, the development of biosensors enabled by HP ¹²⁹Xe also benefit substantially from continuous delivery of HP ¹²⁹Xe, as hyper-CEST^16-21 experiments typically require multiple repetitions with signal saturation over a range of frequencies in a pseudo-2D acquisition mode.²⁵ Beyond the realm of medical diagnostics, HP ¹²⁹Xe has the potential to be used in various materials science applications such as porous materials characterization, surface chemistry analysis, polymer membrane studies, magnetometry,³¹ etc.^32-37 These applications also benefit substantially by having a continuous supply of highly concentrated and highly polarized HP ¹²⁹Xe gas.

HP ¹²⁹Xe is typically produced via spin-exchange optical pumping (SEOP).³⁸ The SEOP process consists of two steps: Firstly, laser photons of the required wavelength (for example, the D₁ transition wavelength of rubidium at ~794.8 nm) supplied by a circularly polarized laser are absorbed by the outer-shell electron of an alkali metal (rubidium in this study), causing a highly spin-polarized electronic ground state of Rb to be achieved over time.³⁹ This electron spin angular momentum is then subsequently transferred to the ¹²⁹Xe nuclei through gas-phase collisions of Rb and ¹²⁹Xe via Fermi contact interactions.⁴⁰ Typically, the production of HP ¹²⁹Xe using the SEOP process is performed either in a continuous-flow^41-43 mode or in a stopped-flow (a.k.a. batch-) mode.^{29, 44-47} A recirculating variant of the continuous-flow method has also been reported for materials science applications.⁴⁸

As the name may imply, the continuous-flow SEOP methodology should be ideally suited for continuous delivery of produced HP ¹²⁹Xe gas exiting the hyperpolarizer device:⁴⁹ Indeed, the flow rate can be readily adjusted for the desired applications.⁵⁰ However, this method usually utilizes a lean (ca. 2% Xe and the rest is carrier) gas mixture as a hyperpolarizer feedstock; richer Xe mixtures are possible for some applications⁵¹ but generally result in much lower Xe polarization levels. Because NMR signal is directly proportional to the concentration or partial pressure of the HP Xe analyte,³ and the corresponding goal of maximizing the signal-to-noise ratio (SNR) of the HP ¹²⁹Xe analyte, it usually becomes necessary to increase the concentration of the produced HP ¹²⁹Xe via cryo-collection⁴⁹ (while other workarounds have been shown, they have their own additional disadvantages^52-53). During cryo-collection, the produced ¹²⁹Xe is cryogenically trapped, and at completion, the frozen HP ¹²⁹Xe is rapidly thawed to produce a bolus of pure HP Xe gas.⁵⁴ The cryo-collection requires the use of liquid N₂, which complicates the production process and defies a key intrinsic advantage of continuous-flow production technology as no HP Xe gas exits the hyperpolarizer during the production process. Moreover, once the HP gas is cryogenically trapped, there is no practical way to cancel the experiment—thus requiring careful planning of such studies and leaving virtually no room for error. Of note, ¹²⁹Xe is expensive ($200-300 per liter*atm in isotopically enriched form and $20-$200 per liter*atm in the non-enriched 26%-natural abundance form).

Batch-mode SEOP production of HP ¹²⁹Xe usually operates in the Xe-rich regime: indeed, since P_Xe over 50% has been demonstrated on a clinical scale^{44, 55-56} in gas mixtures with 50% or greater Xe content, the cryo-collection of HP Xe can be obviated (presenting a key advantage of this method),^{11, 57} therefore making batch-mode SEOP process completely cryogen-free.^{11-12, 55, 58-64} However, this method historically was employed to produce a bolus of HP Xe gas, which is typically ejected into a storage Tedlar bag as a single bolus. Although the ejected gas dosing can be adjusted (to tailor the needs to the desired experimental goal), the produced bolus is poorly suited for continuous-HP-Xe delivery experiments. Due to this gap in the field, a number of continuous delivery devices have been designed to utilize a bolus of HP ¹²⁹Xe gas (as a source) with the goal of performing continuous delivery of HP gas for the desired application.^{58, 65} While these designs have their merits, they inevitably add additional experimental complexity and cost, and require custom hardware.

In this study, we explored the utility of continuous delivery of HP gas from a hyperpolarizer otherwise operating in a “batch mode” of ¹²⁹Xe gas hyperpolarization. Specifically, the produced HP gas is stored inside the hyperpolarizer production glassware. This stored HP gas is released continuously for the delivery of HP ¹²⁹Xe to a 5-mm NMR tube with the flow rate ranging from 3 to 180 standard cubic centimeters per minute (sccm) for up to 15 min. This efficient process becomes possible due to (1) the long lifetime of the HP state in the SEOP cell employed for transient storage inside the hyperpolarizer (T₁ ≅ 2 h), and (2) the short travel time of HP ¹²⁹Xe through the microfluidic PEEK tubing, employed for the continuous gas delivery into standard 5-mm NMR tubes. The utility of the method has been verified by in situ low-field polarimetry performed on the SEOP cell and ex situ polarimetry performed using the 5-mm NMR tube, where the HP ¹²⁹Xe was delivered from the SEOP cell. The P_Xe was additionally verified via ex situ polarimetry of the ejected HP ¹²⁹Xe gas using a 0.35 T clinical MRI scanner. The obtained results are also supported by the HP ¹²⁹Xe relaxation dynamics performed inside the transfer PEEK tubing, glass NMR tube, and Tedlar bag in low magnetic fields. The ejection process was performed using P_Xe-safe solenoid valves and a mass-flow controller that can be readily integrated in the automated workflow of high-resolution NMR experiments using transistor-transistor logic (TTL) control lines of the NMR spectrometer and MRI scanners (work in progress in our laboratory). The presented method provides a simple, cost-efficient approach for continuous delivery of HP ¹²⁹Xe gas produced via the batch-mode SEOP technique.

EXPERIMENTAL SECTION

HP ¹²⁹Xe gas production and in situ polarimetry.

HP ¹²⁹Xe gas mixture was produced by using a batch-mode generation-3 (GEN-3) hyperpolarizer.^66-67 Briefly, hyperpolarization was performed inside a Pyrex 0.5-L 2-inch-inner diameter SEOP cell at an optimized temperature of 75 °C. Once P_Xe buildup was completed (in ~30-60 min), an automated sequence was used to cool down the SEOP cell (in ~5 min) to eject the HP gas at a temperature below 42 °C (near the Rb metal melting point of 39.5 °C, where the Rb vapor pressure is very low). The SEOP cell was filled with a 2000 torr gas mixture: 50% Xe and 50% N₂. Unfortunately, the LDA laser employed in the reported experiments was no longer within the vendor specifications: FWHM ~0.5 nm (versus 0.2 nm FWHM) and output power of <100 W (versus 170 W), resulting in reduced SEOP efficiency: indeed, P_Xe was limited to 30-40% upon completion of the buildup procedure (e.g., Figure S13a) versus P_Xe of over 50% that was routinely achieved previously with an LDA exhibiting normal performance.^66-67 Following the ejection of HP gas, the SEOP cell was reloaded with unpolarized gas mixture for the next SEOP cycle.^66-67 In situ HP ¹²⁹Xe polarimetry (i.e., NMR spectroscopic P measurement of HP species) was performed as described previously (see SI for details).⁶⁸

HP ¹²⁹Xe gas delivery in 5-mm NMR tube and ex situ low-field NMR polarimetry.

Following the cooldown of the SEOP cell below 42 °C, the pressurized HP gas mixture (2000 torr, 50% Xe and 50% N₂) was ejected from the hyperpolarizer via polyetheretherketone (PEEK) tubing line (1/16-in. OD, 0.040-in ID, Idex, P/N 1538XL). The ejection of the HP gas from the hyperpolarizer was done over the course of 15 min of gas delivery into 5-mm NMR tube that was situated inside the polarimetry station operating at ~3.5 mT.⁶⁸ HP ¹²⁹Xe polarimetry in the polarimetry station was performed using Eq. S1, and the corresponding-size reference sample of thermally polarized water doped with CuSO₄ (SI).

The ejected HP gas from the polarizer flows to the NMR tube through PEEK tubing, and then passes through a one-way check valve (to prevent any gas backflow that could result in SEOP cell poisoning), and through a 1/16-in. OD (1/32-in. ID, 1 ft long) Teflon catheter that guides the incoming HP Xe gas to the bottom of the NMR tube. A non-magnetic push-to-connect “T” shaped reducer allows mating (i) the 5-mm NMR tube (via 1/4-in. end), (ii) the incoming HP Xe gas (via 1/8-in. input port), and (iii) exhausting Xe gas mixture (via 1/8-in. exit port). The 1/8-in. terminals of the push-connect adapter use 1/8-in. Teflon tubing lines. The NMR tube is jacketed with 1/4-in. OD Teflon tubing, with the overall approach that follows closely to our previous designs.^69-70 The exit line is then connected to a pressure gauge and to a needle valve (for variable-flow delivery). The exhaust of the setup is open to atmosphere, and it was employed for real-time flowrate measurement of the exiting gas using water displacement in a graduated cylinder, Figure 1.

Figure 1.

a) Schematic diagram of continuous HP ¹²⁹Xe gas delivery from the hyperpolarizer to a 5-mm NMR tube located inside the polarimetry station using a needle valve. P_Xe sensing was performed in-situ within the ¹²⁹Xe polarizer and in the 5-mm NMR tube throughout the delivery process. The HP gas flow control is enabled either by a needle valve or by a MFC equipped with additional solenoid valves and a coupler.

The ~8-cm-long 5-mm NMR tube (washed extensively with methanol) was employed for collecting and sensing of HP ¹²⁹Xe gas. This NMR tube resided inside a homogeneous B₀ electromagnet of the 3.5 mT polarimeter.⁶⁸ The NMR tube was placed next to a surface RF coil that was built similarly to the RF coil for in situ polarimetry, except that it was tuned to 41.9 kHz compared to 40.8 kHz in the hyperpolarizer, Figure 1. This coil was connected to a second XeUS NMR spectrometer⁶⁸ used for ex situ polarimetry on the NMR tube.

The HP ¹²⁹Xe gas ejection was realized in two ways: variable-flow rate release using a needle valve and a continuous-flow rate using a mass flow controller (MFC) and the additional valve and a coupler shown in Figure 1. For variable rate ejection, before the hyperpolarizer enters the 15-minute-long phase of gas ejection, the two-way valve shown in Figure 1 was closed, and three purge cycles of vacuum and N₂ gas application on the hyperpolarizer effectively purged the gas manifold (all the way to the two-way valve) of residual trapped air. When the polarizer started the HP gas ejection, this two-way manual valve was opened, thus, enabling the continuous delivery of HP gas over the 15-min period. When using the needle valve, the gas flow rate is controlled by a pressure gradient, and the flow rate changed from ~3 standard cubic centimeter per second (sccs) at the start of the gas delivery (the highest pressure gradient) down to 0.05 sccs at the end of the delivery period (the smallest pressure gradient between the SEOP cell and the ambient pressure). The following data was sampled every 20 s during the 15-min delivery process: P_Xe in the SEOP cell (e.g., Figure 2a) and in the 5 mm NMR tube (e.g., Figure 2b), gas pressure, and gas flow rate (by monitoring the amount of the total delivered gas), Table S9.

Figure 2.

a) In-situ NMR polarimetry immediately before the gas delivery to 5-mm NMR tube. b) Ex-situ NMR polarimetry of the ejected HP ¹²⁹Xe gas inside the NMR tube at 41.9 kHz. c) T₁ decay of stopped HP ¹²⁹Xe gas in a 5 mm NMR tube. g) T₁ decay of HP ¹²⁹Xe gas inside PEEK tubing. e) P_Xe, flow rate (sccs), and gas pressure (P, atm) during the process of HP gas delivery.

After closing the two-way valve of the delivery setup and the eject valve inside the hyperpolarizer, the gas flow was effectively stopped. Acquisition of NMR signals from stopped HP gas allowed us to measure the HP ¹²⁹Xe T₁ decay, primarily due to relaxation via collision with the glass walls of the NMR tube, Figure 2c. In a separate experiment, the setup was shifted by placing a section of PEEK tubing in front of the RF coil (instead of the NMR tube), allowing us to measure ¹²⁹Xe relaxation inside the PEEK tubing, Figure 2d. Finally, after completion of the HP gas delivery, the SEOP cell was reloaded with Xe:N₂ gas mixture, repolarized, and T₁ was measured again; this measurement was performed to confirm that no Rb metal poisoning occurred during the gas delivery process (which typically manifests itself as a substantial reduction in ¹²⁹Xe T₁ inside the SEOP cell). A series of T₁ data obtained ‘before’ and ‘after’ multiple HP gas ejections is presented in Figure S8.

In a separate series of experiments shown in Figure 3, the experimental setup of Figure 1 was modified as shown in Figures S11 and S12 via two important changes: First, two solenoid valves (Burkert, type 6126) were added before and one after the NMR tube (i.e., both valves denoted by ⊗ were realized by the solenoid valves). An additional PEEK coupler was also added to connect the inlet 1/16” PEEK tube to the 1/16” Teflon catheter: as a result, the total length of PEEK path was increased to 8 ft (versus 6 ft used in the needle-valve configuration described above). Second, an MFC (Sierra Instruments, Monterey, CA, P/N C100L-DD-OV1-SV1-PV2-V1-S0-C0) replaced the needle valve, allowing the setup to maintain a constant HP gas mixture flow during gas delivery from the hyperpolarizer to the NMR polarimetry station.

Figure 3.

Graphs showing time-dependent plots of gas-phase P_Xe, flow rate (sccs), and gas pressure (P, atm) during the process of HP gas delivery using continuous HP ¹²⁹Xe gas-mixture flow rate settings of 150 sccm (a), 90 sccm (b), 60 sccm (c); 1 sccm = 60 sccs.

In a separate experiment—the results of which are presented in the SI—the MFC-based setup was employed (as described above), but ~0.4 mL of HPLC-grade methanol was loaded in the NMR tube, thus allowing HP ¹²⁹Xe signal to be detected from the dissolved phase. Note that P_Xe quantification in the dissolved phase was not performed because the ¹²⁹Xe concentration was not independently determined.

HP ¹²⁹Xe Tedlar bag delivery and ex situ polarimetry.

The P_Xe level measured inside our stopped-flow hyperpolarizers was verified by both low-field polarimetry and polarimetry inside an MRI scanner, providing additional validation for the P_Xe level measured in situ, and ¹²⁹Xe T₁ measurements in Tedlar bag at 3.5 mT and 0.35 T (see SI for details).

3. RESULTS AND DISCUSSION

Continuous delivery of HP ¹²⁹Xe using variable gas flow.

The efficiency of batch-mode hyperpolarizers relies heavily on the relatively long in-cell ¹²⁹Xe T₁ (≥2 h).^{57, 71-73} The key SEOP cell degradation pathway is the oxidation of Rb metal by residual oxygen and air moisture that may enter the cell in trace quantities, typically during the gas handling process. Following Rb oxidation, ¹²⁹Xe in-cell T₁ is reduced substantially (to less than 30 min), resulting in reduced P_Xe. Therefore, the ¹²⁹Xe T₁ provides a “good SEOP-cell health” indicator in batch-mode ¹²⁹Xe SEOP hyperpolarization.^{66, 74} Thus, before proceeding with systematic studies, we performed a series of HP ¹²⁹Xe gas ejections from the SEOP cell under conditions of continuous delivery using the setup shown in Figure 1, with the overall rationale to ensure that the SEOP cell is not being poisoned through this new gas delivery process. Indeed, Figure S14b shows that no significant change in T₁ was seen before versus after the HP gas continuous delivery: the T₁ values were 146 ± 20 and 144 ± 9 min, respectively (a more rigorous testing (Figure S8) showed no substantial T₁ changes after the 11^th HP gas ejection from the hyperpolarizer, additionally confirming that this HP gas ejection procedure is safe for the Rb contained in the SEOP cell).

Encouraged by these pilot findings, we proceeded with systematic HP production studies using needle valve control of the flow rate, Figures 1 and S14. For each production cycle, P_Xe build-up was established for 1 hour at 75 °C on a freshly reloaded SEOP cell (with 2000 Torr Xe:N₂ 50:50 gas mixture), Figure S14c: in one illustrative example, P_Xe of 31±2% was obtained via the optimized process, with a buildup time constant T_b of 24 ± 3 min—corresponding to a buildup rate γ_SEOP of 0.041 ± 0.0006 min⁻¹. Next, the SEOP cell was cooled down to 42 °C, and an in-situ P_Xe value was measured one last time immediately before the HP gas ejection from the SEOP cell: Figure 2a illustrates a representative in-situ NMR spectrum, confirming P_Xe of 26% (a minor reduction of P_Xe during the SEOP cell cool-down is attributed to T₁ relaxation decay of the HP state⁷⁵). Following the HP ¹²⁹Xe gas delivery into the NMR tube located inside the nearby polarimeter, the NMR signal recorded at 41.9 kHz (Figure 2b), revealing P_Xe of 22%. The minor decrease of P_Xe (from 26% measured inside the hyperpolarizer) is rationalized via an accelerated T₁ relaxation due to HP ¹²⁹Xe collision with the walls of the NMR tube and PEEK tubing that Xe gas experiences during the flow process: T₁ = 5.3 ± 0.9 min (Figure 2c) and 5.5 ± 3.3 min (Figure 2d), respectively. The substantial decrease in T₁ is rationalized by the substantial increase in the wall collision rate due to an increase in surface-to-volume ratio. These results are important as they provide guidance for the limits of HP ¹²⁹Xe delivery through narrow-bore tubing: in order to retain the bulk of P_Xe, the travel time of HP gas must be substantially shorter than T₁. This notion is semi-quantitively supported by the continuous HP ¹²⁹Xe gas delivery through the NMR tube, Figure 1h, where the gas flow rate varied from ~3 sccs (at the beginning) down to 0.05 sccs (at the end of a 900-s-long ejection process). The confined volume of the glass and PEEK tubing (i.e., inside the SEOP cell stem, NMR tube path from the bottom to the detection region, and PEEK path) was estimated to be on the order of 10 cc. Therefore, the HP gas travel time at the beginning of the ejection process (P = 2.5 atm) was estimated to be 8 s, which is relatively inconsequential with respect to the T₁ value of approximately 5 min. On the other hand, the HP gas travel time at the end of the ejection process (P = 1 atm) was estimated to be over 3 min, which can no longer be neglected, resulting in substantial P_Xe decrease at the end of the HP gas delivery procedure. The overall contribution of the in-cell T₁ was small, as the 15-min delivery process resulted in P_Xe decreasing by a factor of 1.1. The HP gas delivery process was reproducible: indeed, Figure S9 exhibited overall the same trends.

Continuous delivery of HP ¹²⁹Xe gas using mass-flow control (fixed HP gas flow rate).

Although the use of a needle valve allowed testing of the efficiency of HP gas delivery under a wide range of flow rates and pressures, a fixed flow rate may be desired where steady-state delivery is required, e.g., for biosensor applications.^{18-19, 32, 52-53, 76-77} Thus, we replaced the needle valve by an MFC, and also added two solenoid valves to gate the process of HP gas delivery, Figure 1 and Figure S16a. While the solenoid valves were not required for our experiments, they are greatly desired for the process of automated gas flow cessation during NMR acquisition to minimize susceptibility-induced B₀ field gradients at high magnetic fields. The HP gas delivery experiments were performed at three fixed (as opposed to variable described in the above sub-section) flow rate values: 150 sccm (Figure 3a), 90 sccm (Figure 3b) and 60 sccm (Figure 3c)

A flow rate of 150 sccm (Figure 3a) resulted in the delivery of ~800 scc HP gas in ~320 s, after which the flow rate could no longer be controlled due to lack of back pressure in the SEOP cell. The initial P_Xe oscillation at the beginning of the ejection process is explained by initial oscillation in the MFC output. Following the flow rate stabilization, the P_Xe (18-19%) and the flow rate (150 sccm) remained steady throughout the process, until the HP gas overpressure in the cell was exhausted. Of note, P_Xe “paradoxically” exhibited no decrease between t = 100 s to t = 300 s, which is discussed in more detail below.

The experiments using a flow rate of 90 sccm (Figure 3b) resulted in an extended gas flow (up to 600 s) with consistent P_Xe of 16-17%, and also exhibited a small but “paradoxical” growth in P_Xe despite T₁ relaxation in the SEOP cell. The reader is reminded that a small decrease in P_Xe (by a factor of 1.1) is anticipated during the 900-second-long ejection process due to the in-cell T₁ of ~140 min, Figure S14b. Further reduction of the flow rate to 60 sccm (Figure 3c) enabled a stable flow to be maintained for 800 s, as expected. Moreover, the “paradoxical” trend of P_Xe “growth” (from 16% to 19%) between t = 0 s and t = 400 s was more pronounced in this dataset. We rationalize this counter-intuitive observation as possibly originating from two potential contributions. First, the P_Xe throughout the SEOP cell may not be uniform thus resulting in the change of the detected P_Xe (downstream in the NMR tube after the HP gas delivery) as the gas is released from the SEOP cell. Second, while the flow rate is fixed throughout the process of HP gas delivery (blue curve), the pressure was decreasing (green curve). The pressure decrease is important as it results in the effectively reduced gas transfer time, potentially resulting in reduced T₁ relaxation losses during gas passage through the tubing. As a result of this counter-effect, the in-cell T₁ decay would be mitigated or in some cases (Figure 3c) would be overpowered by the gains manifested in reduced gas travel time and reduced T₁ losses during gas delivery—thereby resulting in a smoother trend of the detected P_Xe inside the 5 mm NMR tube versus initial estimates.

When the HP gas flow rate was reduced to 45 sccm (Figure S15), and the ejected gas was bubbled through an NMR tube filled with 0.4 mL methanol, a steady flow of HP Xe gas was maintained throughout the 900-s ejection period, revealing a nearly constant signal from the dissolved-phase HP Xe in methanol (Figure S15b). The P_Xe quantification was not possible because concentration of Xe in methanol was not measured.

All in all, the results reported in Figures 2-3 clearly demonstrated a remarkable utility of the stopped-flow hyperpolarizer to create a continuous delivery of a stream of HP Xe gas for up to 15 min, with small variation in P_Xe throughout the process. The key disadvantage of the reported approach is the requirement to produce a batch of HP gas prior to delivery. As a result, it may take a 30-60 min before the process can be initiated. The primary advantages of this new approach are: 1) cryogen-free operation; 2) a wide dynamic range of gas flow rates; 3) opportunities for automation and integration in the gated workflow of the NMR spectrometer or MRI scanner (e.g., using TTL control lines to synchronize gas ejection with pulse sequence events using solenoid valves and the MFC). Moreover, in case the user decides to abort the procedure or not use the produced HP Xe batch, there is little wasted Xe gas, as the bulk of the unused Xe gas is retained in the SEOP cell and can be re-used. The latter point is an important consideration because Xe is expensive: up to $300/liter.

Additional P_Xe validation via 0.35 T MRI.

We have performed additional studies to validate the obtained P_Xe levels using a 0.35 T MRI scanner, with the overall goal to enhance the scientific validity of the reported findings and brings this study one step closer to an anticipated real-world application. For these studies, the experimental setup was truncated with the exit path connected directly to the Tedlar bag via PEEK/Tygon line and check valve, Figure S16a. Following the collection of ~0.8 L of HP gas in a one-liter Tedlar bag, P_Xe assessment at 3.5 mT field of the polarimetry station and at 0.35 T field of MRI scanner were performed. In this experiment, in-situ P_Xe of 25% was measured in the SEOP cell of the hyperpolarizer (Figure S16b) immediately before HP gas delivery into a Tedlar bag placed inside the polarimetry station. Immediately following the ejection, P_Xe of 21% was recorded from the Tedlar bag at 41.9 kHz, Figure S16c. Following the HP gas transfer to the MRI scanner facility in the same Tedlar bag (total delivery time of ~12 mins), P_Xe was reduced to 19% as measured inside the 0.35 T MRI scanner, Figure S16d. The experimental details on the transfer process and the transfer device, including bill of materials (BOM) are provided in the SI.

Outlook.

A number of additional efforts could be employed to improve overall performance. For example, using glass wall sealing with SurfaSil^™ for coating of the detection reservoir could improve the T₁ relaxation within the NMR tube. Reduction of T₁ transfer losses may also allow one to perform HP ¹²⁹Xe gas delivery with even lower flow rates, without substantial P_Xe losses. The reported studies clearly indicate that HP ¹²⁹Xe can be delivered continuously from a stopped-flow hyperpolarizer with steady P_Xe levels over significant periods of time, spanning 15 min. We anticipate the 0.8 L volume should be sufficient for gas delivery of up to 60 min, albeit at reduced flow rate and P_Xe values (although this could be augmented by using higher in-cell pressures).

CONCLUSION

In conclusion, we have developed a robust method for continuous delivery of HP ¹²⁹Xe gas using an automated cryogen-free batch-mode clinical-scale hyperpolarizer. It was possible to deliver HP gas to a 5-mm NMR tube situated approximately 1.5 m from the hyperpolarizer (over ~2.5-meter total path length), and we envision the length can be substantially increased based on the semi-quantitative T₁ relaxation model. P_Xe of up to 25% was demonstrated after the delivery, which can be further improved using a pristine high-power laser for more optimal SEOP performance, as has been previously demonstrated.^{56, 66, 74} The continuous gas delivery was demonstrated using a wide range of flow rates: 0.05-3 sccs (albeit with P_Xe reduction at slower rates), clearly demonstrating the versatility of this approach. Constant-flow durations of up to 15 min were shown using 45 sccm HP gas flow rate, which potentially can be further increased. The capability of controlling the continuously-delivered HP gas flow rate using a MFC with solenoid valve gating of the gas flow was also demonstrated, which is desired for automated multi-scan experiments using the control interface of an NMR spectrometer, MRI scanner, or a stand-alone controller. The obtained P_Xe values of the delivered HP gas were measured by low-field polarimetry at 3.5 mT and additionally confirmed by polarimetry using a 0.35 T clinical open MRI scanner. We also report pilot feasibility MRI experiments employing a 0.35 T scanner with a large-volume transmit RF coil that was integrated in the poles of the open MRI magnet, paving the way for streamlined studies, employing HP ¹²⁹Xe gas contrast agents in rodents and in human volunteers. Taken together, the results reported here have the potential to expand the utility of the recently FDA-approved HP ¹²⁹Xe gas contrast agent for a wide range of complementary pre-clinical and bioanalytical applications.