Laëtitia Cesari
email: laetitia.cesari@univ-lorraine.fr

Christophe Castel

Eric Favre

Membrane contactors for intensified gas-liquid absorption processes with physical solvents: a critical parametric study

Keywords: Membrane contactors, gas-liquid absorption, gas separation, physical solvents, intensification

HAL is

List of parameters

Uppercase parameters

𝐶 Concentration

Cp heat capacities at constant pressure Cv heat capacities at constant volume

Introduction

Membrane contactors are considered as one of the most promising technology for the intensification of gas-liquid absorption processes [START_REF] Feron | CO2 separation with polyolefin membrane contactors and dedicated absorption liquids: performances and prospects[END_REF]. The use of modules equipped with highly gas permeable membranes and placed between the gas and liquid phase indeed offers unique possibilities compared to packed columns. The much larger specific surface area of membrane modules (5000 to 10 000 m -1 ) compared to column packings (200-500 m -1 ) opens the possibility to significantly decrease the size of the installation. The so-called intensification factor (I) which results from the volume reduction is one of the key interest of the technology [START_REF] Gabelman | Hollow fiber membrane contactors[END_REF][START_REF] Bounaceur | Membrane contactors for intensified post combustion carbon dioxide capture by gas-liquid absorption in MEA: A parametric study[END_REF][START_REF] Zhao | Status and progress of membrane contactors in post-combustion carbon capture: A state-of-the-art review of new developments[END_REF][START_REF] Favre | Membrane contactors for intensified post-combustion carbon dioxide capture by gas-liquid absorption processes[END_REF]. A significant economy in terms of weight, because of the low density of polymers, can also be obtained; this characteristic can be of interest for off shore applications or mobile systems. Additionally, because no direct contact takes place between phases in a membrane contactor, the gas and liquid flowrates can be independently chosen. Consequently, a much larger ratio of gas / liquid velocities is possible and weeping or flooding phenomena are prevented. Nevertheless, several problems can limit the interest of membrane contactors; a low gas permeability material (such as a wetted hydrophobic membrane) and / or a low specific surface area module (i.e. large membrane diameter or low packing fraction) can lead a specific size of the unit which is larger than the baseline packing column case (i.e. I < 1). Moreover, pressure drop effects in hollow fiber modules are likely to significantly increase the energy requirement of the gas absorption process; a trade-off between process intensification and energy requirement has thus to be taken into account, but few studies addressed this point [START_REF] Abu-Zahra | CO2 capture from power plants: Part I. A parametric study of the technical performance based on monoethanolamine[END_REF][START_REF] Chabanon | Pushing the limits of intensified CO2 post-combustion capture by gas-liquid absorption through a membrane contactor[END_REF]. Membrane stability issues are also of importance, especially for harsh environments, such as chemical solvents.

Up to now, much effort has been made to investigate the potential of membrane contactors for gas-liquid processes based on chemical solvents, especially for carbon capture applications. The possibility to decrease the size of CO2 postcombustion capture units is effectively of major interest. Numerous studies have been reported in order to evaluate the possibilities and limitations of membrane contactors for CO2 capture with solvents such as MEA and MDEA, among others. The key role of membrane mass transfer performance, the strong impact of wetting effects and the materials stability issues have been highlighted. Through a series of modelling and simulation investigations, post-combustion membrane contactors can now be rigorously designed, with several pilot units tested with real flue gases.

Surprisingly, the same situation does not hold for gas liquid absorption processes based on physical solvents [START_REF] Chabanon | Gas-liquid separation processes based on physical solvents: opportunities for membranes[END_REF]. A large number of industrial units already make use however of physical solvents for a variety of gas purification applications, such as CO2 removal from a pressurized gas mixture (hydrogen purification, precombustion carbon capture [START_REF] Chabanon | Membrane contactors for intensified gas-liquid absorption processes[END_REF]). Absorption in a physical solvent is classically considered as the baseline technology for high pressure gas separation applications, with a pressure swing regeneration process to enable solvent recycling. Physical solvent processes are most often operated at high pressure in order to enhance the solubility of gases in the solvent and therefore generate a higher driving force. Even if the gas solubility decreases with a rise of temperature, physical solvents are also often operated at high temperature in order to reduce viscosity and pressure drop effects. The operating conditions, among other parameters, have a strong influence on the efficiency of the separation and can lead to an economical strategy, with optimal specifications for a given mixture [START_REF] Mokhatab | Chapter 7 -Natural Gas Sweetening[END_REF].

Unfortunately, few studies have been reported on membrane contactors operated with physical solvents.

In a pioneering study, Dindore et al [START_REF] Dindore | Membrane-solvent selection for CO2 removal using membrane gas-liquid contactors[END_REF] experimentally investigated the use of propylene carbonate in a hollow fiber membrane contactor for the removal of CO2 at elevated pressure [START_REF] Dindore | CO2 absorption at elevated pressures using a hollow fiber membrane contactor[END_REF]. Their experiments showed that in the case of physical absorption the overall mass transfer is controlled by the liquid-side mass transfer resistance and morphological changes were observed inside the membrane, leading to a wetting of the fiber. More recently, some publications have explored pressurised water absorption (PWA) processes for biogas upgrading; the efficiency of 1D simulations to predict experimental separation performances and the predominant mass transfer resistance of the liquid phase have been shown. In order to circumvent the difficulties of porous materials observed by Dindore (membrane wetting is likely to occur due to the low surface tension of physical solvents and high pressure conditions), dense skin membrane materials have been proposed [START_REF] Dindore | CO2 absorption at elevated pressures using a hollow fiber membrane contactor[END_REF]. Alternatively, ionic liquids are actively investigated for physical absorption processes and some preliminary results with membrane contactors have been reported [START_REF] Albo | Carbon Dioxide Capture from Flue Gases Using a Cross-Flow Membrane Contactor and the Ionic Liquid 1-Ethyl-3-methylimidazolium Ethylsulfate[END_REF]. The quantitative evaluation of the potentialities of membrane contactors in terms of process intensification are however essentially unexplored for the above applications.

This study intends to fill that gap through a systematic parametric study. Four solvents, which are commonly used in the physical absorption of CO2: methanol (Rectisol process), N-methyl-2-pyrrolidone NMP (Purisol process), propylene carbonate (Fluor process), and polyethylene glycol dimethyl ether PGDE (Selexol process) [START_REF] Dindore | Membrane-solvent selection for CO2 removal using membrane gas-liquid contactors[END_REF][START_REF] Luis | Use of monoethanolamine (MEA) for CO2 capture in a global scenario: Consequences and alternatives[END_REF][START_REF] Prevention | Control Reference Document on Best Available Techniques for the Manufacture of Large Volume Inorganic Chemicals-Ammonia[END_REF] have been first selected. Because it is a cheap and green solvent with a high surface tension, water has been added for comparison purposes, despite its low CO2 solubility [START_REF] Dindore | Membrane-solvent selection for CO2 removal using membrane gas-liquid contactors[END_REF]. Interestingly, these five different solvents show different CO2 solubility performances [START_REF] Burr | A COMPARISON OF PHYSICAL SOLVENTS FOR ACID GAS REMOVAL, 87th Annual Gas Processors Association Convention[END_REF] but also different physico-chemical properties (e.g. viscosity, density, surface tension …). The impact of solubility and viscosity on intensification and energy requirement performances will thus be achievable. It can be anticipated that solvent viscosity is of importance for membrane contactor application, because operating conditions with physical solvents require a large specific liquid flowrate (compared to chemical solvents), leading to high pressure drop effects in a compact system. The intensification / energy requirement trade-off should thus to be systematically taken into account when membrane contactors are aimed for applications with physical solvents.

More specifically, the following questions will be addressed:

-What is the maximal intensification factor of membrane contactors for physical absorption processes compared to packed columns ? -What are the best flow configurations (i.e. liquid flowing inside or outside the fibers), module design (packing factor), membrane performances (mass transfer coefficient) ? -What is the interplay between intensification and specific energy requirement ? -How do solvent viscosity and gas solubility impact the intensification performances ?

It will be shown that process intensification is not necessarily achieved with membrane contactors when applied to physical absorption processes ; moreover, the impact of solvent properties drastically changes the level of performances. Nevertheless, impressive volume reduction factors are attainable if specific guidelines for membrane and module design are used.

Overall framework of the study

The aim of the study is to explore the intensification potential of membrane contactors for physical absorption processes. It is expected that the possibilities offered strongly differ from the numerous studies reported up to now for chemical absorption processes. Indeed, given the major differences between physical and chemical absorption processes, summarized in Table 1, the interest of membrane contactors might be reconsidered:

-In terms of membrane performances, a very high membrane mass transfer coefficient (km) is needed for chemical solvents, because of the high liquid mass transfer coefficient. Typically, a 10 -4 m.s -1 or more value is recommended in order to maintain intensification potential. For lower values (such as obtained with wetted membranes), the membrane resistance becomes dominant and the interest of membrane contactors is questionable. For physical solvents, the liquid mass transfer coefficient is much lower (10 -5 m.s -1 or less); lower membrane mass transfer performances are thus required. This opens solutions such as dense selfstanding membranes which can withstand high pressure conditions [START_REF] Li | An Ultrathin Skinned Hollow Fibre Module for Gas Absorption at Elevated Pressures[END_REF][START_REF] Trusov | Gas/liquid membrane contactors based on disubstituted polyacetylene for CO2 absorption liquid regeneration at high pressure and temperature[END_REF].

-Chemical solvents are mostly aqueous solutions showing low viscosities. This leads to limited pressure drop effects on the liquid phase in membrane contactors. Conversely, the larger viscosity and much higher liquid/gas flowrate of physical absorption processes raises the question of excessive pressure drop levels in the liquid phase. -Intensification factors around 4 have been reported for CO2 post-combustion capture by chemical solvents with membranes contactors operated with real flue gases [START_REF] Falk-Pedersen | CO2 Capture with Membrane Contactors[END_REF][START_REF] Klaassen | Membrane contactor applications[END_REF][START_REF] Li | Post-combustion CO2 capture using super-hydrophobic, polyether ether ketone, hollow fiber membrane contactors[END_REF]. The pressure drop impact remains limited and module designs can fit the intensification / energy requirement trade-off [START_REF] Chabanon | Pushing the limits of intensified CO2 post-combustion capture by gas-liquid absorption through a membrane contactor[END_REF]. The corresponding problem is unexplored for physical solvents.

Membrane contactor performances

Intensification factor (I)

Around 4 Unknown

Energy requirement (kWh.Nm -3 )

Acceptable for adequate module design

Unknown Figure 1 presents the synopsis of the study where : a) A membrane contactor unit is studied in order to replace a packed column used for CO2 absorption by a physical solvent. b) The overall volume of the unit (dotted line) and the specific energy requirement will be compared depending on solvent type, membrane performances, module design and operating conditions. The modelling and simulation strategy of the parametric study is detailed in the next section.

Modelling and parametric study

Properties of fluids

The properties of the five solvents at 25°C are listed in Table 2. The values of the CO2 solubility in these solvents are taken from [START_REF] Dindore | Membrane-solvent selection for CO2 removal using membrane gas-liquid contactors[END_REF][START_REF] Burr | A COMPARISON OF PHYSICAL SOLVENTS FOR ACID GAS REMOVAL, 87th Annual Gas Processors Association Convention[END_REF]. In this table, the Henry coefficient is defined as follows: [START_REF] Dindore | Membrane-solvent selection for CO2 removal using membrane gas-liquid contactors[END_REF] The influence of temperature is investigated on propylene carbonate. The density and viscosity of the solvent have been taken from [START_REF] Barthel | Density, Relative Permittivity, and Viscosity of Propylene Carbonate + Dimethoxyethane Mixtures from 25 °C to 125[END_REF]. The Henry coefficient of CO2 in propylene carbonate has been taken from Anouti et al for 333K [START_REF] Anouti | Low pressure carbon dioxide solubility in pure electrolyte solvents for lithium-ion batteries as a function of temperature. Measurement and prediction[END_REF] and from Murrieta-Guevara et al for 373K [START_REF] Murrieta-Guevara | Solubilities of carbon dioxide and hydrogen sulfide in propylene carbonate, N-methylpyrrolidone and sulfolane[END_REF].

𝐻𝑒 𝑖 = 𝐶 𝑖 𝑔𝑎𝑠 𝐶 𝑖 𝑙𝑖𝑞𝑢𝑖𝑑 (1)
The determination of the diffusion coefficient and the diffusivity of gases in solvents were performed according to the methodology proposed by Dindore et al [START_REF] Dindore | CO2 absorption at elevated pressures using a hollow fiber membrane contactor[END_REF] which is described as follows.

The determination of the diffusivity of gases in the solvents at 25°C were calculated by the correlation of Diaz et al [START_REF] Díaz | Correlation for the Estimation of Gas-Liquid Diffusivity[END_REF] as follows :

(𝐷 12 ) 𝑇=25°𝐶 = 6.02 • 10

With D12 the diffusivity coefficient of solute 1 in solvent 2 in cm².s -1 , V2 and V1 the molar volumes at normal boiling point temperature for gas (solute) and liquid (solvent) respectively in cm 3 .mol -1 , µ the viscosity of the solvent in cP.

For other temperature, the diffusivity of gases in solvents has been calculated by the correlation of Wilke et Chang [START_REF] Wilke | Correlation of diffusion coefficients in dilute solutions[END_REF] :

(𝐷 12 ) = 7.4 • 10 -8 • √𝜒•𝑀 • 𝑇 µ𝑉 0.6 (3)
Where D12 is the diffusivity coefficient of solute 1 in solvent 2 in cm².s -1 , M the molecular weight of the solvent, T the temperature in K,V the molar volume of the solute at normal boiling point temperature in cm 3 .mol -1 , µ the viscosity of the solvent in cP. 𝜒 is the association parameter. For ordinary nonassociated solvents, 𝜒 = 1.0. [START_REF] Wilke | Correlation of diffusion coefficients in dilute solutions[END_REF][START_REF] Wanderley | Mapping Diluents for Water-Lean Solvents: A Parametric Study[END_REF] The molar volumes V2 of the solvents at their normal boiling point were predicted by a groups contribution method described by Schotte. [START_REF] Schotte | Prediction of the molar volume at the normal boiling point[END_REF] The diffusion coefficient of the binary mixture {CO2 -CH4} was calculated by the Fuller equation [START_REF] Poling | Properties of Gases and Liquids, Fifth Edition[END_REF]:

𝐷 𝐴𝐵 = 0.00143 𝑇 1.75 𝑃𝑀 𝐴𝐵 1/2 [(∑ ) 𝜐 𝐴 1 3 +(∑ ) 𝜐 𝐵 1 3 ]² ( 4
)
Where DAB is the binary diffusion coefficient in cm².s -1 , P is the pressure in bar, T the temperature in Kelvin, ∑ 𝜐 is the summation of atomic diffusion volumes, MA and MB are the molecular weights of A and B, and

𝑀 𝐴𝐵 = 2 [( 1 𝑀 𝐴 ) + ( 1 𝑀 𝐵 )] -1 .

Properties of membrane contactor

Numerous parameters can have an influence on the modelling of CO2 physical absorption and must be taken into account in the parametric study. Figure 2 depicts a membrane contactor and the different parameters divided into four categories: parameters related to the membrane, to the contactor, to the solvent and the operating conditions. The membrane fiber is characterised through its internal radius ri, thickness e and mass transfer coefficient km (in m.s -1 ). Numerous studies addressed the impact of membrane mass transfer coefficient on membrane contactor performances. Typical values range between 10 -4 to 10 -5 for non wetted porous membranes or dense skin composite membranes. Thin dense selfstanding membranes can show km values close to 10 -5 m.s -1 [START_REF] Ritchie | A novel automated method for the determination of membrane permeability in gas-liquid transfer applications[END_REF]. For wetted porous membranes, the mass transfer coefficient strongly decreases, down to 10 -6 m.s -1 .

The contactor can be operated in several configurations: co-current or counter-current and liquid-in or liquid-out. It is well known that the counter-current flow configuration is more efficient than the cocurrent one. Therefore, only the counter-current will be investigated in this work. However, choosing between the liquid-in or liquid-out configurations to obtain the best performances is always an issue. For this reason, both conformations are investigated in this work. The internal diameter of the contactor is fixed at 0.17 m, corresponding to a significant number of fibers and the length of the contactor is left variable and will be determined through the simulations. Finally, the module packing factor φ, corresponding to the volume fraction occupied by the fiber bundle over the overall membrane contactor volume, is investigated on a very large range, between 0.1 and 0.9.

The parameters related to the choice of the solvent are the solubility of CO2 and the properties of the solvent (viscosity, density). As presented before, these parameters are listed in Table 2.

Finally, the operating temperature and pressure are fixed to 298.15K and 1 Bar respectively. The inlet gas enters the contactor with a 40/60 molar ratio of CO2/CH4 mixture at a volumetric flow of 1.39.10 -2 Nm 3 .s -1 . The interstitial velocity ratio, defined as the liquid velocity over the gas velocity inside and outside the fibers, is investigated between 0.1 and 30. The outlet condition is a CO2 molar fraction below 0.02 in the gas phase, typical of CO2 purification targets (e.g. natural gas, biogas…) [START_REF] Heile | Establishing the suitability of symmetric ultrathin wall polydimethylsiloxane hollow-fibre membrane contactors for enhanced CO2 separation during biogas upgrading[END_REF][START_REF] Li | Removal of carbon dioxide from pressurized landfill gas by physical absorbents using a hollow fiber membrane contactor[END_REF][START_REF] Kim | Practical designs of membrane contactors and their performances in CO2/CH4 separation[END_REF].

All the fixed and variable parameters investigated in this study and their values are listed respectively in Tables 3 and4.

Mass transfer

It is assumed that the membrane contactor is operated at steady state under isothermal conditions and no evaporation of the solvent. Moreover, the membrane mass transfer km is presumed constant. Finally, for both gas and liquid phases, plug flow conditions are settled. This set of condition is classically proposed for simulation purposes [START_REF] Chabanon | Modeling strategies of membrane contactors for postcombustion carbon capture: A critical comparative study[END_REF].

The interfacial area inside ain and outside aout the fiber and medium am, the hydraulic diameter of the fiber dh, the log mean radius rml and the number of fibers are calculated according to:

𝑎 𝑖𝑛 = 2 𝜑 𝑟 𝑖𝑛 𝑟 𝑜𝑢𝑡 2 𝑎 𝑜𝑢𝑡 = 2 𝜑 𝑟 𝑜𝑢𝑡 (5)
𝑟 𝑚𝑙 = 𝑒 log(𝑟 𝑜𝑢𝑡 𝑟 𝑖𝑛 ⁄ ) (6)
𝑎 𝑚 = 2 𝜑 𝑟 𝑚𝑙 𝑟 𝑜𝑢𝑡 2 (7)
𝑑 ℎ = 2 𝑟 𝑜𝑢𝑡 (1-𝜑) 𝜑 (8)
𝑁 𝑓𝑖𝑏𝑒𝑟𝑠 = 𝛺 𝜑 𝜋 𝑟 𝑜𝑢𝑡 2 (9)
Where Ω is the section of the membrane contactor, rin and rout are the internal and external radius of the fibers.

The velocities and the dimensionless quantities Reynolds number Re, Schmidt number Sc and Graetz number Gz are defined for inside and outside the fibers as follows :

𝑢 𝑖𝑛 = 𝑄 𝑖𝑛 𝑁 𝑓𝑖𝑏𝑒𝑟𝑠 𝜋 𝑟 𝑖𝑛 2 𝑢 𝑜𝑢𝑡 = 𝑄 𝑜𝑢𝑡 𝛺 (1-𝜑) (10) 𝑅𝑒 𝑖𝑛 = 𝜌 𝑖𝑛 𝑢 𝑖𝑛 2 𝑟 𝑖𝑛 µ 𝑖𝑛 𝑅𝑒 𝑜𝑢𝑡 = 𝜌 𝑜𝑢𝑡 𝑢 𝑜𝑢𝑡 𝑑 ℎ µ 𝑜𝑢𝑡 (11)
𝑆𝑐 𝑖𝑛 = µ 𝑖𝑛 𝜌 𝑖𝑛 𝐷 𝑖𝑛 𝑆𝑐 𝑜𝑢𝑡 = µ 𝑜𝑢𝑡 𝜌 𝑜𝑢𝑡 𝐷 𝑜𝑢𝑡 (12) 𝐺𝑧 𝑖𝑛 = 𝐷 𝑖𝑛 𝑍 𝑢 𝑖𝑛 (2 𝑟 𝑖𝑛 ) 2 𝐺𝑧 𝑜𝑢𝑡 = 𝐷 𝑜𝑢𝑡 𝑍 𝑢 𝑜𝑢𝑡 𝑑 ℎ 2 (13)
Where Q, ρ, µ, D, u are the molar flow, density, viscosity, diffusion coefficient and velocity of the fluids. rin is the internal radius of the fiber, dh is the hydraulic diameter and Z the length of the contactor. The subscripts in and out refer to the inside or the outside of the fiber.

The determination of the transfer coefficient in the lumen and shell sides were determined using the same equations as used in our previous work [START_REF] Bounaceur | Membrane contactors for intensified post combustion carbon dioxide capture by gas-liquid absorption in MEA: A parametric study[END_REF][START_REF] Belaissaoui | Potentialities of a dense skin hollow fiber membrane contactor for biogas purification by pressurized water absorption[END_REF] and are reminded below.

The determination of the Sherwood number inside the fibers was performed using the Graetz equations:

𝑆ℎ 𝑖𝑛 = { 1.3𝐺𝑧 𝑖𝑛 (-1 3 ⁄ ) 𝑖𝑓 𝐺𝑧 𝑖𝑛 < 0.03 4.36 𝑖𝑓 𝐺𝑧 𝑖𝑛 ≥ 0.03 (14)
On the shell side, the Sherwood number is determined for laminar flow (Re<500) as before, using the Graetz [START_REF] Janssen | Transport Phenomena Data Companion[END_REF] equations and for turbulent flow (Re>500) by the correlation of Kartohardjono and Chen [START_REF] Kartohardjono | Mass Transfer and Fluid Hydrodynamics in Sealed End Hydrophobic Hollow Fiber Membrane Gas-liquid Contactors[END_REF].

𝑆ℎ 𝑜𝑢𝑡 = { 1.3𝐺𝑧 𝑜𝑢𝑡 (-1 3 ⁄ ) 𝑖𝑓 𝑅𝑒 𝑜𝑢𝑡 <
The transfer coefficient inside kin and outside kout the fibers are defined according to:

𝑘 𝑖𝑛 = 𝑆ℎ 𝑖𝑛 𝐷 𝑖𝑛 2 𝑟 𝑖𝑛 𝑘 𝑜𝑢𝑡 = 𝑆ℎ 𝑜𝑢𝑡 𝐷 𝑜𝑢𝑡 𝑑 ℎ (16)
The overall mass transfer coefficient is calculated according to a resistance in series approach [START_REF] Gabelman | Hollow fiber membrane contactors[END_REF] as follows:

For liquid out:

1 𝐾 = 𝑎 𝑚 𝑘 𝑖𝑛 𝑎 𝑖𝑛 + 1 𝑘𝑚 + 𝐻𝑒 𝑎 𝑚 𝑘 𝑜𝑢𝑡 𝑎 𝑜𝑢𝑡 (17)
For liquid in:

1 𝐾 = 𝑎 𝑚 𝑘 𝑜𝑢𝑡 𝑎 𝑜𝑢 + 1 𝑘𝑚 + 𝐻𝑒 𝑎 𝑚 𝑘 𝑖𝑛 𝑎 𝑖𝑛 (18)
Finally, the evolution of the molar composition of CO2 in the gas phase can then be determined by the resolution of the following differential equations:

with the following boundary conditions:

{ 𝑦 𝑖 = 𝑦 𝑖,0 𝑧 = 0 𝐶 𝑖 𝑙 = 𝐶 𝑖 𝑙,𝐿 𝑧 = 𝐿 (21)
Where QG and QL are the volume flows of gas and liquid respectively, yi the molar composition of compound i and z the length of the contactor.

The average absorbed CO2 molar flow can be calculated as follows:

𝑁 𝐶𝑂2 = 𝑄 𝐺_0 • 𝑦 𝐶𝑂2_0 -𝑄 𝐺_𝐿 • 𝑦 𝐶𝑂2_𝐿 (22)
Where the subscripts 0 and L correspond to the entrance and the exit of the membrane contactor respectively.

Pressure drop

The pressure drop inside the fibers is given by the Hagen-Poiseuille expression

Δ𝑃 𝑖𝑛 = 𝑓 𝜌 𝑖𝑛 𝑢 𝑖𝑛 2 𝐿 4𝑟 𝑖𝑛 ( 23
)
Where f is the pipe friction coefficient.

According to the type of flow regime, the pipe friction coefficient f can be defined differently.

If the Reynolds number inside the fiber 𝑅𝑒 𝑖𝑛 is lower than 2100 (corresponding to a laminar flow), then :

𝑓 = 64 𝑅𝑒 𝑖𝑛 (24)
Therefore, the pressure drop can be expressed as

Δ𝑃 𝑖𝑛 = 8𝑢 𝑖𝑛 𝜇 𝑖𝑛 𝐿 (𝑟 𝑜𝑢𝑡 -𝑒) 2 (25)
For a turbulent flow (𝑅𝑒 𝑖𝑛 higher than 2100), the pipe friction coefficient f is defined as:

1 √𝑓 = -2 log 10 [ 2.51 𝑅𝑒 𝑖𝑛 1 √𝑓 + 𝜀 3.7(2𝑟 𝑖𝑛 ) ] ( 26
)
Where ε is the roughness factor of fiber.

The latter can be evaluated by setting the value of the ratio 𝜀 (2𝑟 𝑖𝑛𝑡 ) ⁄ , ranging from 10 -6 (smooth pipe) to 0.05 (rough pipe).

Outside the fibers, the pressure drop is determined as follows:

Δ𝑃 𝑜𝑢𝑡 = 4𝐾 𝑜 𝑢 𝑜𝑢𝑡 𝜇 𝑜𝑢𝑡 𝜑 2 𝐿 𝑟 𝑜𝑢𝑡 2 (1-φ) (27)
where 𝐾 𝑜 (-) is the Kozeny constant, that can be evaluated as follows:

𝐾 = 150𝜑 4 -314.44𝜑 3 + 241.67𝜑 2 -83.039𝜑 + 15.97 (28)

Energy efficiency

The efficiency of each solvent and configuration is determined by the transferred CO2 molar flow in comparison with its energy consumption. The transferred CO2 molar flow is defined as the difference of the inlet and outlet CO2 molar flows. The energy consumption is defined as the sum of the pressure drop in each phase, corresponding to the energy consumption of the compressor and the pump. The analytical expressions are given for each configuration.

For liquid-out configuration:

𝑃 𝑐𝑜𝑚𝑝𝑟 = 𝑄 𝑔 𝛾 𝛾-1 ( 𝑃+𝛥𝑃 𝑖𝑛 𝑃 0 ) ( 𝛾-1 𝛾 ) (29)
𝑃 𝑝𝑢𝑚𝑝 = 𝑄 𝑙 𝛥𝑃 𝑜𝑢𝑡 [START_REF] Ritchie | A novel automated method for the determination of membrane permeability in gas-liquid transfer applications[END_REF] For liquid in configuration:

𝑃 𝑐𝑜𝑚𝑝𝑟 = 𝑄 𝑔 𝛾 𝛾-1 ( 𝑃+𝛥𝑃 𝑜𝑢𝑡 𝑃 0 ) ( 𝛾-1 𝛾 ) ( 31
)
𝑃 𝑝𝑢𝑚𝑝 = 𝑄 𝑙 𝛥𝑃 𝑖𝑛 (32)
With 𝛾 =

𝐶 𝑝 𝐶 𝑣 (33)
Where Q is the volumetric flow. The subscripts g and l correspond to the gas and liquid phases respectively. Cp and Cv are the heat capacities of the gas mixture at constant pressure and volume.

Packed column

For the packed column, taken as a baseline process here (Figure 1.a), simulations were performed at room temperature with the same inlet gas composition and flow. The influence of the pressure is investigated between 1 and 10 Bar and the operating point is set at 80% of the flooding area. The diameter of the packing dppack is set at 0.035 m with a specific area a of 140 m 2 .m -3 . Some relevant steps of the strategy are described below.

The column was design with the flooding determination by Eckert [START_REF] Eckert | A new llok at distillation tower packings comparative performance[END_REF]:

𝑌 𝑓𝑙𝑜𝑜𝑑𝑖𝑛𝑔 = 𝑒 (0.1117-4.012𝑋 0.25 ) (34)
𝑋 = 𝑢 𝐿 𝑢 𝐺 √ 𝜌 𝐿 𝜌 𝐺 (35)
𝑢 𝑓𝑙𝑜𝑜𝑑𝑖𝑛𝑔 = 1 𝜌 𝐺 √ 𝑌 𝑓𝑙𝑜𝑜𝑑𝑖𝑛𝑔 𝜌 𝐿 𝜌 𝐺 𝑔 𝐹 ( 36
)
Where F is the packing factor of the column fixed at 187 m 2 .m -3 .

Ω =

𝑄 𝑔 𝑢 𝑓𝑙𝑜𝑜𝑑𝑖𝑛𝑔 0.8 [START_REF] Kartohardjono | Mass Transfer and Fluid Hydrodynamics in Sealed End Hydrophobic Hollow Fiber Membrane Gas-liquid Contactors[END_REF] The effective interfacial area is calculated at loading point according to Billet and Schultes [START_REF] Billet | Prediction of Mass Transfer Columns with Dumped and Arranged Packings: Updated Summary of the Calculation Method of Billet and Schultes[END_REF]:

𝑎 𝑒𝑓𝑓 = 1.5 (𝑎. 𝑑 ℎ ) 0.5 ( 𝜌 𝐿 𝑢 𝐿 𝑑 ℎ µ 𝐿 ) -0.2 ( 𝑢 𝐿 2 𝜌 𝐿 𝑑 ℎ 𝜎 𝐿 ) 0.75 ( 𝑢 𝐿 2 𝑔𝑑 ℎ ) -0.45 (38)
Where 𝜎 𝐿 is the surface tension of the liquid phase.

The determination of the transfer coefficient is performed through the calculation of the dimensionless numbers and the resistance in series approach:

𝑅𝑒 𝐺 =

Results and discussion

Influence of parameters

The evaluation of the solvent's performance is determined by the value of the molar flow of CO2 transferred as a function of the total energy consumption. In our case, the total energy consumption is mostly caused by the pressure drop in the liquid side. A completely different situation is obtained with chemical solvents, where pressure drop on the gas side are usually dominant. Each curve shows the same pattern with an optimal point in term of energy consumption (not always depicted in graphs). A schematic representation is depicted in Figure 3. Each point corresponds to a different velocity ratio. At very low velocity, the transfer is not efficient enough and a huge length of the fiber is required leading to an elevated pressure drop. When the velocity increases, the transfer is favoured, leading to a shorter fiber and a lower pressure drop. After the optimal point, the transfer is still favoured by the rise of the velocity, but this latter generates a rise in the pressure drop.

Influence of solvent properties

Figure 4 shows the results for the five studied solvents at different packing fraction values for a mass coefficient transfer km fixed at 10 -3 m.s -1 and a liquid-in configuration. At first glance, it can be observed that Selexol and water show the lowest performances for the set of specifications fixed for CO2 purification. Indeed, the molar flows of CO2 transferred are the lowest with a very large energy requirement. When looking at low energy values, the performance of the solvents decreases as follows methanol > NMP > PC > Selexol > water. The same trend is observed for the km values of 10 -4 and 10 -5 m.s -1 and for the liquid out configuration also. These results can be explained by the low CO2 solubility in water and by the high viscosity of Selexol. Indeed, an increase of the viscosity will lead to a diminution of the CO2 flux and a high energy consumption. Looking at the viscosity of Rectisol, NMP, PC and Selexol, it can be noticed that the performances of these solvents follow the reverse tendency of the viscosity. Moreover, the CO2 solubility appears to be a critical parameter. Indeed, PC and water seems to have similar physical properties. However, the solubility of CO2 in water is 4 times lower than in PC. The CO2 transferred flux is not in the same order of magnitude: the value is 10 times higher for PC than for water at the same energy consumption. Hence, a low CO2 solubility will lead to higher impact on the energy efficiency than a high viscosity. Etot (kWh.Nm -3 ) e)

Influence of membrane performances

In addition to the liquid side, performances can also be affected by the value of the membrane transfer coefficient km. The value of km was investigated between 10 -3 and 10 -5 m.s -1 for the five solvents. Results in Table 5 are given for the liquid in configuration with a module compacity (packing fraction) of 0.5 and ul/ug ratio of 2. As expected, a higher value of the km coefficient leads to a better CO2 transferred flux and a lower energy consumption for all the solvents. One can observe that the most efficient solvents (methanol and NMP) are the most affected by the decrease of the km value. Moreover, it can be highlighted that a decrease of the km value from 10 -3 to 10 -4 m.s -1 leads to a smaller variation of the energy consumption and transferred flux than a decrease of the km value from 10 -4 to 10 -5 m.s -1 . For example, the ratio of transferred flux over energy consumption for methanol is divided by 4 (transferred flux divided by 2, energy consumption multiplied by 2) when the value of km drops from 10 -3 to 10 -4 m.s -1 . For a change from 10 -4 to 10 -5 m.s -1 , the ratio for methanol goes down to 25 (transferred flux divided by 5, energy consumption multiplied by 5). On the other hand, water is only slightly altered by the modification of km. Indeed, the transfer coefficient value in the liquid side is quite low meaning that the liquid side is more limiting than the membrane. The investigation on the influence of the membrane transfer coefficient km logically shows that the higher the value, the better the absorption performances. A value of km around 10 -3 and 10 -4 m.s -1 seems ideal for physical solvents. A lower value (e.g. 10 -5 m.s -1 ) will lead to a limitation by the membrane. Therefore, for porous membranes, it remains essential to avoid any process such as membrane wetting that could decrease the km value. Alternatively, thin skin composite membranes, which are not impacted by wetting problems, can achieve km values around 10 -4 m.s -1 , while dense selfstanding membranes seem limited to 10 -5 m.s -1 .

Influence of flow configuration (liquid in or liquid out)

The influence of the configurationliquid-in or liquid-outon the overall efficiency for several packing fraction values is depicted with methanol as example in Figure 5. In the liquid-in configuration, the CO2 transferred flow increases and the energy consumption decreases as the packing fraction increases. Since the number of fibers in the module increases, the specific area increases therefore improves the transfer.

In the meantime, the velocity in each fiber decreases, diminishing the pressure drop in the liquid side.

In the liquid-out configuration, a different phenomenon is found. An increase of the packing factor values from 0.1 to 0.5 will increase the transfer due to the increase of the specific area. However, with a value of 0.6 and higher, the efficiency of the transfer decreases drastically with a rise of the energy consumption caused by a higher pressure drop in the liquid side. Similar effects are observed for all solvents but for different values of compacity (module packing fraction). Thus, the compacity shows an optimum value that is strongly depending of the solvent used. To our knowledge, no study reported up to now the occurrence of optimal module packing fractions for membrane contactors as a function of solvent properties. This result is important for future membrane contactor design strategies. b)

The choice of the liquid-in or liquid-out configuration has always been an issue for the use of membrane contactors. In Figure 6, the maximal CO2 absorption fluxes are represented for each solvent and configuration, namely with a km value of 10 -3 m.s -1 and a compacity of 0.9 for liquid-in configuration and the optimum value for liquid-out configuration. It can be noticed that for all solvent, the liquid-out configuration is the most efficient. The difference is the most impressive for the most efficient solvent (methanol and NMP). The fact that liquid out configuration offers the best performances whatever solvent type is noticeable and should be taken into account for future studies. However, it is important to note that the prediction of pressure drop and mass transfer coefficients on the shell side of membrane contactors is complex. Numerous correlations have been already proposed, depending on fiber geometry and packing fraction, among others. Different correlations will necessarily quantitatively modify the pressure drop and mass transfer predictions. A detailed analysis of the impact of different correlations on performances would be of interest but it is outside the scope of this study. We think however that the systematic liquid out option suggested above remains robust whatever the membrane module simulation strategy.

Figure 6: Maximum CO2 flow for each solvent with Km value fixed at 10 -3 m.s -1 . Blue: water; orange: methanol; grey: NMP; yellow: Selexol; green : PC., Continuous lines: liquid in configuration (compacity of 0.9); dotted lines: liquid out configuration (compacity of 0.5)

It should be also stressed that membrane mechanical properties will play a key role, given the pressure drop levels; more specifically, the fibers are subjected to a risk of crushing for the liquid-out configuration or bursting for the liquid-in configuration.

Influence of pressure and temperature

As presented in the introduction of the article, physical solvent processes generally operate at high pressure to favour the solubility of gases in solvent and improve the driving force. On the other hand, the diffusivity coefficient and solvent properties are not significantly impacted by pressure.

Temperature has a very strong impact on several parameters in physical solvent processes. From a mass transfer coefficient point of view, an increased temperature will be interesting because of the lower viscosity of the solvent. This has to be however balanced by the simultaneous decreased gas solubility,

E tot (kWh.Nm -3 ) that is a lower driving force. Because gas-liquid absorption flux depends on the product of driving force and mass transfer coefficient, it is interesting to study the influence of temperature. This is especially true for systems which show a major mass transfer resistance in the liquid, that is for high membrane mass transfer coefficients. Figure 7 shows the results for propylene carbonate for 3 different temperature levels: 298 K, 333K and 373 K. b)

It first can be noticed that membrane contactor with low capacity (φ values below 0.4) can only be operated at low temperature for a reasonable energy consumption (below 10 kWh.Nm -3 ). Moreover, high CO2 absorption fluxes can be achieved for low temperature conditions (Figure 7a). With an increase of temperature up to 333K (Figure 7b), CO2 absorption flux shows an important decrease, but at high temperature (Figure 7c), the values of CO2 flows are surprisingly slightly higher. These results demonstrate that the solubility of gases in solvent is the governing parameter that must be the higher as possible to achieve maximal mass transfer. Thus, it is advised to operate with a low or ambient temperature. Secondly, the temperature must be high enough to have a significant impact on the solvent properties, especially the viscosity, and on the diffusivity coefficient. With only a small increase of temperature, physical solvent process presents a lose/lose situation with low gas solubility, low diffusivity coefficient and high viscosity. Anyways, the physical properties and diffusivity advantages at high temperature cannot compensate the loss due to the solubility. These results are obtained for propylene carbonate solvent but similar phenomena are expected to hold for other solvents. The combination of gain and loss with temperature requires a full study with strategic parameters for a given mixture.

Comparison with packed columns: Intensification possibilities

As previously presented, the membrane contactor presents numerous advantages in comparison with the packed column. One major objective of this study is to compare the two technologies in term of specific volume (i.e. specific CO2 absorption fluxes). Results of the packed column, the baseline case are presented in Table 6. As for the membrane contactor, the same performance tendency is obtained for the five solvents: methanol > NMP > PC > selexol > water. The absorption fluxes logically increase with a rise of the pressure, together with an increased energy requirement. The intensification factor I can then be evaluated, through the volume ratio of the packed column over the membrane contactor. A series of data is reported in Table 7 for a 1 Bar inlet pressure and a maximal energy requirement of 1 kWh/Nm 3 . For all solvents, both liquid-in and liquid-out configurations are equivalent or better than the packed column. Interestingly, the maximal intensification factors are obtained for the maximal module packing fraction (0.9), while a variable optimal packing fraction is required for the liquid out configuration (0.2 to 0.5). Moreover, the highest I values are systematically obtained for the liquid out configuration, and the optimal module packing factor increases with a decrease of the solvent viscosity. The possibility to achieve impressive intensification factors is clear: for a 1 Bar inlet pressure, intensification factors reach values of 13.3 for methanol, 11.9 for NMP, 8.56 for PC, 8.07 for selexol and 7.08 water. Methanol thus offers the highest intensification possibilities, due to a shorter contactor length and a higher velocity ratio. However, methanol is also the only solvent to develop a turbulent regime for this value of energy consumption. The impact of pressure on the maximal intensification factor is also of interest and it has been explored through a parametric study. Simulations have been performed at 1, 5 and 10 Bar gas pressure for the liquid in configuration. Results are shown in Table 8 for the following conditions: ul/ug = 1, φ = 0.9, km = 10 -3 m.s -1 . The driving force being proportional to the pressure, a rise of the pressure implies a rise of the CO2 absorption flux. In the same way, the length of the contactor required to obtain the outlet 2% CO2 composition is shortened. Consequently, the total energy consumption also decreases. Table 8 also presents the calculations of the intensification factor of the CO2 molar transferred flow for the liquid in configuration of the membrane contactor in comparison with the packed column at the same gas pressures (1, 5 and 10 bars). For each solvent, the intensification factor remains globally the same for different pressures. These results imply that for a first approach, the intensification factor I are transposable in pressure.

Table 8: Influence of the pressure in liquid-in configuration (ul/ug = 1, φ = 0.9, km = 10 -3 m.s -1 ). The impact of temperature has also been investigated on the intensification factor I. Simulations have been performed at 298, 333 and 373 K on membrane contactor and packed column are results are presented in Table 9. As previously shown in section 4.6, for the membrane contactor, a rise of temperature implies two opposite phenomena: better transfer and low viscosity of the solvent, but also a lower solubility of the gas. In the packed column, the low solubility of gas with the increasing temperature requires a high velocity ratio ul/ug that completely modify the design of the column. Therefore, the packed column is more efficient at high temperature with a lower height and a higher CO2 transferred flow. Contrary to the pressure, the intensification factor is strongly temperature dependent and the higher I values are achieved at the lower temperature.

Solvent

Table 9: Influence of the temperaturecomparison with packed columns for propylene carbonate. Membrane contactor parameters: liquid in configuration, ul/ug = 1, φ = 0.9, km = 10 -3 m.s -1 .

Membrane contactor Packed column

Conclusion

This study intended to explore the possibilities of process intensification thanks to membrane contactors for absorption processes based on physical solvents. A systematic parametric study has been performed for CO2 absorption in five different solvents showing a broad range of solubility and viscosity.

The main conclusions can be summarized as follows:

-Membrane contactors offer promising intensification possibilities compared to a classical packed column with volume reduction ratios up to 15 -The maximal intensification factor I is almost independent of the inlet pressure -The liquid out configuration systematically shows the highest intensification factor, providing that an optimal module packing fraction is used. The optimal module fraction depends on solvent properties and range between 0.2 for low viscosity solvents and 0.5 for high viscosity solvents -For the liquid in configuration, the optimal module packing fraction is always maximal (i.e. 0.9 in this study)

-The membrane mass transfer performances are of major importance, both for the intensification performances and specific energy requirement. The effective membrane mass transfer coefficient km should ideally be above 10 -5 , or better, 10 -4 m.s -1 in order to achieve maximal intensification performances. Interestingly, numerous studies have reported experimental data that fit this range, be it with porous, thin film composite or dense selfstanding membranes. Nevertheless, the interest of membrane contactors drastically decreases for values below 10 -5 m.s -1 . This data is typical of porous, wetted membrane materials.

-Increasing the specific absorption fluxes (i.e. pushing the intensification factor) systematically increases the energy requirement, through a non linear relationship. The energy requirement is largely dominated by the liquid pressure drop effect, at the difference of membrane contactors for chemical absorption systems (for which a moderate gas side pressure drop is the dominant energy requirement term). Given the much larger energy requirement data obtained for physical solvents, the intensification / energy requirement trade-off is thus of major importance and should be systematically taken into account. A technico-economical analysis is necessary in order to identify the best balance between large intensification factors (i.e. lower CAPEX) and increased energy requirement (i.e. larger OPEX).

-Finally, the solvent properties also play a role on the overall process performances, both for the intensification possibilities and energy requirement impact. A high solubility and low viscosity should logically be favoured; this statement explains why Rectisol (methanol) lead to the best performances among the five solvents investigated in this study.

Beyond the above conclusion, this study suggests several perspectives for future work.

The interest of membrane contactors for other applications or different solvents could be also investigated. The results obtained here show however that solvent viscosity strongly impacts the interest and feasibility of membrane contactors compared to packed columns. To that extend, it is likely that high viscosity solvents such as ionic liquid, glycerol or oils, used for gas drying, gas purification or VOC recovery applications poorly fit the requirements for membrane contactors to be of interest.

Because the solubility / viscosity interplay plays a key role in the maximal intensification performances, a parametric study on operating temperature would be interesting. It is known that a lower temperature generally increases solubility but also viscosity and reduces the diffusivity coefficient. The intensification strongly depends on the solvent temperature. Strategic parameters can be achieved though the study of gain and loss with temperature. Lastly, for a given gas mixture, the determination of specific solvent properties could be obtained with a process-design methodology.

The influence of solvent volatility, which has not been taken into account in this study, should also be considered. Methanol and water indeed show vapour pressure levels which induce solvent losses during the absorption process. Symmetrically, wet feed mixtures can also lead to water absorption effects when hydrophilic solvents, such as ionic liquids, are used. In that case multicomponent fluxes from and to the liquid phase should be considered. Interestingly, dense membranes (thin film composite or selfstanding) could offer attractive possibilities in order to limit the solvent losses. For the Rectisol (methanol) case, a dense polymer showing a large permeability towards CO2 but a low permeability towards methanol would surely be of interest.

It is expected that this exploratory parametric study will stimulate further researches on intensified processes with physical solvents.

Funding

This research did not receive any specific grant from funding agencies in the public, commercial, or notfor-profit sectors.

Figure 1 :

 1 Figure 1: Synopsis of the study.

Figure 2 :

 2 Figure 2: Membrane contactor and its parameters adapted from [2].

Figure 3 :

 3 Figure 3: Decomposition of the different regimes

Figure 4 :

 4 Figure4: Generic overview of simulation results for 5 solvents with increasing packing factor: molar flow of CO2 transferred as a function of the total energy with a km value of 10 -3 m.s -1 for the liquid in configuration. Figurea) methanol (Rectisol), b) NMP (Purisol), c) DEPG (Selexol), d), propylene carbonate (Fluor solvent), e) water. φ values : •: 0.1 ; ■ : 0.2; -: 0.3 ; × : 0.4; ▲ : 0.5 ; ♦ : 0.6 ; ○ : 0.7; □ : 0.8 ; ∆ : 0.9.

Figure 5 :

 5 Figure5: Molar flow of CO2 transferred as a function of the total energy for methanol with a Km value of 10 -3 m.s -1 for a) liquid in configuration; b) liquid out configuration. φ values : •: 0.1 ; ■ : 0.2; -: 0.3 ; × : 0.4; ▲ : 0.5 ; ♦ : 0.6 ; ○ : 0.7; □ : 0.8 ; ∆ : 0.9.

Figure 7 :

 7 Figure 7: Molar flow of CO2 transferred as a function of the total energy for propylene carbonate in liquid in configuration with a Km value of 10 -3 m.s -1 for several temperature a) T = 298K, b) T = 333K, c) T = 373K. φ values : •: 0.1 ; ■ : 0.2; -: 0.3 ; × : 0.4; ▲ : 0.5 ; ♦ : 0.6 ; ○ : 0.7; □ : 0.8 ; ∆ : 0.9.

Table 1 :

 1 Major differences between physical and chemical absorption processes

	Chemical	Physical

Table 2 :

 2 List of the investigated solvents and their properties at 25°C

Solvant Usual name Molar mass Density Viscosity CO2 solubility CH4 solubility g.mol -1 kg.m -3 mPa.s - -

	MeOH	Rectisol	32	785	0.6	0.32 a	6.17
	DEPG	Selexol	280	1030	5.80	0.28 a	4.2
	N-methyl-2-	Purisol	99	1027	1.65	0.28 a	3.9
	pyrrolidone (NMP)
	Propylenecarbonate	Fluor solvent	102	1195	3.00	0.29 a	7.7
	(PC)
	Water	Water	18	1000	1.00	1.22 b	28.8
	a) from [16]
	b) from

Table 3 :

 3 List of fixed parameters and their value

	Symbol	Parameter	Value	Unit
	Qgas,0	Volume flow of gas at the entrance	0.0139	Nm 3 .s -1
	T	Temperature	298	K
	R	Ideal gas constant	8.314	J.mol -1 .K -1
	α	Selectivity	1	/
	Dint	Internal diameter of the module	0,17	m

Table 4 :

 4 List of variable parameters and their corresponding variation range

	Symbol	Parameter	Value	Unit
	φ	Module packing fraction	0.1 -0.9	/
	ul/ug	Interstitial velocity ratio	0.1 -20	/
	km	Membrane effective mass transfer coefficient	10 -3 -10 -5	m.s -1
	P	Pressure	1 -10	bar
	/	Solvents	/	/
	/	Configurations	/	/

Table 5 :

 5 Comparison of the influence of the km value: 10 -3 -10 -5 m.s -1 . Results are given for a compacity of 0.5 and ul/ug ratio of 2.

	km (m.s -1 )		MeOH	NMP	PC	Selexol	Water
	10 -3	kl (m.s -1 )	1.83 10 -5	1.43 10 -5	9.06 10 -6	8.96 10 -6	8.24 10 -6
		Etot (kWh.Nm -3 )	0.896	1.25	3.69	6.83	22.0
		N CO2 (mol.m -3 .s -1 )	8.11 10 -1	7.50 10 -1	4.61 10 -1	4.82 10 -1	4.55 10 -2
	10 -4	kl (m.s -1 )	1.7 10 -5	1.17 10 -5	7.83 10 -6	7.74 10 -6	8.24 10 -6
		Etot (kWh.Nm -3 )	1.68	2.27	5.71	10.6	23,6
		N CO2 (mol.m -3 .s -1 )	4.32 10 -1	4.15 10 -1	2.98 10 -1	3.11 10 -1	4.25 10 -2
	10 -5	kl (m.s -1 )	1.7 10 -5	1.21 10 -5	7.80 10 -6	7.51 10 -6	8.24 10 -6
		Etot (kWh.Nm -3 )	8,12	10.3	20.8	39.2	38,2
		N CO2 (mol.m -3 .s -1 )	8.99 10 -2	9.2 10 -2	8.18 10 -2	8.44 10 -2	2.62 10 -2

Table 6 :

 6 Results for packed column

	Solvent Pressure Gas feed	Liquid	Column	ul/ug	Column	CO2 flow Column
			flow	flow	diameter		height		volume
		bar	Nm 3 .s -1	m 3 .h -1	m	-	m	mol.m -3 .s -1	m 3
		1	0.0139	20.47	0.36	0.37	3.91	0.56	0.41
	Methanol	5	0.0139	4.09	0.18	0.37	3.40	2.76	0.08
		10	0.0139	2.05	0.13	0.37	3.20	5.26	0.04
		1	0.0139	18.25	0.34	0.33	6.68	0.37	0.62
	NMP	5	0.0139	3.65	0.17	0.33	6.02	1.75	0.13
		10	0.0139	1.83	0.12	0.33	5.73	3.32	0.07
		1	0.0139	19.17	0.35	0.35	9.44	0.25	0.91
	PC	5	0.0139	3.83	0.17	0.35	8.70	1.19	0.19
		10	0.0139	1.92	0.12	0.35	8.34	2.26	0.10
		1	0.0139	18.12	0.34	0.33	17.57	0.14	1.62
	Selexol	5	0.0139	3.62	0.17	0.33	16.33	0.65	0.35
		10	0.0139	1.81	0.12	0.33	15.66	1.22	0.19
		1	0.0139	79.47	0.76	1.43	4.35	0.12	1.97
	Water	5	0.0139	15.89	0.32	1.43	4.34	0.66	0.34
		10	0.0139	7.95	0.22	1.43	4.30	1.35	0.17

Table 7 :

 7 Highest values of NCO2 for a total energy consumption Etot around a value of 1 kWh.Nm -3 (Temperature = 298K, Pressure = 1 bar).

	Solvent	Configuration φ value	Z	NCO2	ul/ug	ul	I
			-	m	mol.m -3 .s -1	-	m.s -1
	Methanol	Liquid in	0.9	4.62	2.30	0.5	3.39	4.11
		Liquid out	0.5	1.44	7.43	5.0	8.36	13.3
	NMP	Liquid in	0.9	5.65	1.89	0.4	2.71	5.11
		Liquid out	0.5	2.42	4.41	2.25	3.76	11.9
	PC	Liquid in	0.9	13.72	0.78	0.2	1.35	3.12
		Liquid out	0.4	4.98	2.14	1.3	2.72	8.56
	Selexol	Liquid in	0.9	20.7	0.51	0.1	0.677	3.64
		Liquid out	0.3	9.46	1.13	0.7	1.95	8.07
	Water	Liquid in	-	-	-	-	-	-
		Liquid out	0.2	12.5	0.85	1.4	5.85	7.08


Corine E Delsing

Mark S Gresnigt

Jenneke Leentjens

Frank Preijers

Florence Allantaz Frager

Matthijs Kox

Guillaume Monneret

Fabienne Venet

Chantal P Bleeker-Rovers

Frank Van

Florence Allantaz Frager

Frank L Van De Veerdonk

Peter Pickkers

Alexandre Pachot

Bart Jan Kullberg

Mihai G Netea
email: mihai.netea@radboudumc.nl

Interferon-gamma as adjunctive immunotherapy for invasive fungal infections: a case series

Keywords: Immunotherapy, Interferon-gamma, Candidemia, Aspergillosis

come

Background

The incidence of fungal infections is steadily increasing in the last years due to invasive medical diagnosis and immunosuppressive treatment modalities. Despite development of new classes of antifungal agents [START_REF] Segal | Combination antifungals: an update[END_REF], the invasive fungal infections remain associated with unacceptable high mortality rates and represent a major cause of death worldwide [START_REF] Wisplinghoff | Nosocomial bloodstream infections in US hospitals: analysis of 24,179 cases from a prospective nationwide surveillance study[END_REF][START_REF] Gudlaugsson | Attributable mortality of nosocomial candidemia, revisited[END_REF][START_REF] Horn | Epidemiology and outcomes of candidemia in 2019 patients: data from the prospective antifungal therapy alliance registry[END_REF][START_REF] Zaoutis | The epidemiology and attributable outcomes of candidemia in adults and children hospitalized in the United States: a propensity analysis[END_REF][START_REF] Warnock | Trends in the epidemiology of invasive fungal infections[END_REF][START_REF] Brown | Hidden killers: human fungal infections[END_REF]. The emergence of significant resistance to the currently available antifungal therapies emphasizes the need for novel approaches to treat invasive fungal infections [START_REF] Rodloff | Epidemiology and antifungal resistance in invasive candidiasis[END_REF][START_REF] Howard | Frequency and evolution of Azole resistance in Aspergillus fumigatus associated with treatment failure[END_REF]. Invasive fungal infection are most commonly observed in individuals with immune defects or a compromised immune system, and the number of these patients is steadily increasing [START_REF] Kriengkauykiat | Epidemiology and treatment approaches in management of invasive fungal infections[END_REF]. Therefore, adjunctive immunotherapy to improve host defence is an attractive strategy to improve the outcome of patients with disseminated fungal infections.

In the past decade, major progress in the understanding of anti-fungal host responses has enabled the development of a number of novel molecular and cell-based immunotherapeutic approaches for invasive fungal infections [START_REF] Romani | Immunity to fungal infections[END_REF]. Although invasive candidiasis and aspergillosis are rather different in their pathogenesis, the major protective host response against both fungi is the effective induction of Th1 and IFN-γ responses [START_REF] Cenci | Induction of protective Th1 responses to Candida albicans by antifungal therapy alone or in combination with an interleukin-4 antagonist[END_REF][START_REF] Netea | Differential role of IL-18 and IL-12 in the host defense against disseminated Candida albicans infection[END_REF][START_REF] Chai | Anti-Aspergillus human host defence relies on type 1 T helper (Th1), rather than type 17 T helper (Th17), cellular immunity[END_REF][START_REF] Cenci | Interleukin-4 causes susceptibility to invasive pulmonary aspergillosis through suppression of protective type I responses[END_REF][START_REF] Centeno-Lima | Kinetics of cytokine expression in mice with invasive aspergillosis: lethal infection and protection[END_REF]. The Th1 cytokine response activates effector phagocytic cells that kill the fungus [START_REF] Ito | T cell immunity and vaccines against invasive fungal diseases[END_REF]. Interestingly, Th1 immunity against A. fumigatus was demonstrated to be crossprotective against C. albicans [START_REF] Stuehler | Cross-protective TH1 immunity against Aspergillus fumigatus and Candida albicans[END_REF].

Interferon-gamma (IFN-γ), the prototype Th1 cytokine, promotes Th1 differentiation and skews the immune response towards a protective Th1 phenotype [START_REF] Schroder | Interferon-gamma: an overview of signals, mechanisms and functions[END_REF]. As such, it has been implicated as a treatment option in (invasive) fungal infections [START_REF] Lehrnbecher | Immunotherapy against invasive fungal diseases in stem cell transplant recipients[END_REF][START_REF] Stevens | Interferon-gamma as an antifungal[END_REF]. Moreover, limited evidence suggests that recombinant IFN-γ (rIFN-γ) has a beneficial effect on the outcome of fungal infections in patients with chronic granulomatous disease (CGD) [START_REF] Group | A controlled trial of interferon gamma to prevent infection in chronic granulomatous disease[END_REF], HIV [START_REF] Riddell | A phase III study of recombinant human interferon gamma to prevent opportunistic infections in advanced HIV disease[END_REF][START_REF] Bodasing | Gamma-interferon treatment for resistant oropharyngeal candidiasis in an HIV-positive patient[END_REF][START_REF] Jarvis | Adjunctive interferon-gamma immunotherapy for the treatment of HIV-associated cryptococcal meningitis: a randomized controlled trial[END_REF], leukemia [START_REF] Poynton | Interferon gamma and granulocytemacrophage colony-stimulating factor for the treatment of hepatosplenic candidosis in patients with acute leukemia[END_REF][START_REF] Dignani | Immunomodulation with interferon-gamma and colony-stimulating factors for refractory fungal infections in patients with leukemia[END_REF], and in patients receiving organ transplants [START_REF] Teo | Exogenous interferon-gamma immunotherapy for invasive fungal infections in kidney transplant patients[END_REF]. However, it has not been investigated whether rIFN-γ actually enhances the immune response in these patients to explain these beneficial clinical effects.

In this report we describe a series of patients with invasive Candida and/or Aspergillus infections in whom we investigated the effects of treatment with rIFN-γ on the host innate and adaptive immune responses.

Methods

Patients and treatment

To assess the feasibility and preliminary efficacy of IFN-γ in combination with anidulafungin for the treatment of candidemia, a single-centre, prospective, randomized open-label pilot (Phase IIIb) study was conducted. This study was registered at ClinicalTrials.gov (NCT01270490) and approved by the local ethics committee of the Radboud University Medical Center. Due to slower than anticipated enrollment rates (from August 2010 until March 2013, only 12 patients could be screened, of which 6 were eligible and provided informed consent [Figure 1]), the study was terminated early. However, during this period, several other patients presented with invasive fungal infections which had an insufficient response to standard antifungal therapy. Although these patients did not meet the inclusion criteria (i.e. presenting with one or more positive cultures of blood or normally sterile tissue growing Candida spp.), they were deemed to benefit from adjunctive immunotherapy as "therapy of last resort" as decided by the attending physician. Within the parameters of standard clinical care these patients were treated according to the same protocol as the patients enrolled in the study, and were therefore included in the present case series. All patients with a history of documented epileptic seizures, pre-existent severe renal impairment (creatinine clearance <30/mL/min) or severe liver failure (defined as a spontaneously increased prothrombin time) were excluded. After obtaining informed consent, eight patients (3 study patients, 5 last resort patients) were treated with rIFN-γ (Immukine, Boehringer Ingelheim, 50 μg/m 2 body surface, subcutaneously, three times a week) in addition to standard antifungal therapy as recommended by national and international treatment guidelines [START_REF] Pappas | Infectious Diseases Society of America: Clinical practice guidelines for the management of candidiasis: 2009 update by the Infectious Diseases Society of America[END_REF][START_REF] Lashof | Optimalisation of the antibiotic policy in the Netherlands XIII. Dutch Working Party on Antibiotic Policy (SWAB) guideline for the treatment of invasive fungal infections[END_REF]. Three patients who were included in the Phase IIIb Candida pilot-study were assigned to the control group and did not receive rIFN-γ.

Blood sampling

Plasma, serum and whole blood specimens were collected at baseline (BL) and serially after the start of antifungal therapy (days 1, 2, 3, 7, 14 and 28). Blood cultures were performed as part of routine care.

Leukocyte populations and surface HLA-DR expression

Heparin anticoagulated blood was stored at 4°C immediately after withdrawal and analyzed by flow cytometry. To determine the extent of immune suppression, HLA-DR expression was determined by calculating % HLA-DR-positive cells and HLA-DR mean fluorescence intensity (MFI) within CD14 + cells and various lymphocyte subsets within CD45 + leukocytes + (see Additional file 1 and Additional file 2: Figure S1 for details and a representative flow diagram). Lymphocyte subsets were defined as: T-cells (CD45 + CD3 + ), T-helper cells (Th, CD45 + CD3 + CD4 + ), cytotoxic T-cells (Tc, CD45 + CD3 + CD8 + ), B-cells (CD45 + CD19 + ), and NK-cells (CD45 + CD3 -CD56 + ). Subset counts were calculated by multiplying the percentage of gated cells by the total lymphocyte count. Patients with <50% HLA-DR positive monocytes at baseline were considered to exhibit immune paralysis. This threshold of 50% is well below the lower bound of the 99% confidence interval obtained in healthy volunteers in an earlier study of our group using the same methodology in the same laboratory [START_REF] Leentjens | Reversal of Immunoparalysis in Humans In Vivo: A Double-Blind, Placebo-controlled, Randomized Pilot Study[END_REF]. Therefore mHLA-DR expression levels below 50% are likely to represent immunoparalysis.

Cytokine assays

Venous blood was drawn into 10 mL EDTA tubes, after which peripheral blood mononuclear cells (PBMCs) were isolated as described previously [START_REF] Netea | Aspergillus fumigatus evades immune recognition during germination through loss of toll-like receptor-4-mediated signal transduction[END_REF]. In short, blood was diluted in phosphate buffered saline (PBS) (1:1) and fractions were separated by Ficoll (Ficoll-Paque Plus, GE healthcare, Zeist, The Netherlands) density gradient centrifugation. Cells were washed twice with PBS and resuspended in RPMI-1640+ (RPMI-1640 Dutch modification supplemented with 10 μg/mL gentamicin, 10 mM L-glutamine, and 10 mM pyruvate) (Gibco, Invitrogen, Breda, The Netherlands). The PBMCs were counted using a particle counter (Beckmann Coulter, Woerden, The Netherlands) and were plated in 96 well round-bottom plates (Corning, NY, USA) at a final concentration of 2,5×10 6 /mL, in a total volume of 200 μL. The PBMCs were stimulated for 24 hours, 48 hours, and 7 days with medium alone, or medium containing E. coli lipopolysaccharide (LPS; 10 ng/mL), phytohaemaglutinnin (PHA; 10 μg/ml), heat-inactivated Candida albicans blastoconidia (1×10 6 /ml) or heat-inactivated Candida albicans hyphae (derived from 1×10 6 /m conidia). After stimulation, cell culture supernatant was collected and stored at -20°C. When all samples were collected, cytokines were measured using commercially available ELI-SAs (R&D Systems, MN, USA and Sanquin, Amsterdam, The Netherlands) according to the protocols supplied by the manufacturer. Ex-vivo production of cytokines was assessed at timepoints at which their production has been shown to peak [START_REF] Van De Veerdonk | The macrophage mannose receptor induces IL-17 in response to Candida albicans[END_REF]. Monocyte derived cytokines such as Interleukin (IL)-1β and tumour necrosis factor (TNF)α were measured in culture supernatants of 24 hour cultures, IL-10 was measured in culture supernatants of 48 hour cultures. T-cell derived cytokines IL-17 and IL-22 were measured in culture supernatants of 7 day cultures.

Statistical analysis

In view of the small sample size, normality of distribution was not assumed. Comparisons of baseline with follow up time points were made using Wilcoxons signed rank test (within-group comparisons, 2 groups). A p-value of <0.05 was considered statistically significant. Data are expressed as means and standard error of the mean. Calculations and statistical analyses were performed using GraphPad Prism v 5.0 (GraphPad Software, San Diego, CA, USA).

Results

Patient characteristics

The patients treated with rIFN-γ (5 men, 3 women) had a median age of 49.5 [IQR 28.5-68.8] years. The three female patients in the control group were 36, 51 and 73 years old. Clinical characteristics of the participants are listed in Table 1. Of the 6 patients included in the pilot study, three patients had a positive blood culture for C. albicans, two patients for C. glabrata, and one patient for C. tropicalis. During randomization, the three patients with C. albicans cultures were assigned to rIFN-γ treatment, whereas the two patients with C. glabrata and one with C. tropicalis cultures were assigned to the control group. However, no pathophysiological evidence currently exists to suggest that rIFN-γ therapy would have a different effect on the immune system in case of albicans vs. non-albicans Candida infections. Of the other 5 patients treated with rIFN-γ as therapy of last resort, one patient had proven acute aspergillosis, one had probable acute aspergillosis, and one probable chronic aspergillosis according to the EORTC/MSG criteria [START_REF] De Pauw | Revised definitions of invasive fungal disease from the European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) Consensus Group[END_REF]. One patient had a positive blood culture for C. tropicalis associated with osteomyelitis. This patient developed new suspected lesions on positron emission tomography-computed tomography (PET-CT) while receiving antifungal treatment. In another patient, CTscan revealed progression of suspected hepatic Candida lesions during antifungal treatment. All patients included suffered some degree of immunosuppression: the 6 patients with positive blood cultures for Candida spp. had impaired physical barriers due to the presence of γ γ γ 12 patients were assessed for eligibility 7 patients met one or more exclusion criteria, or did not provide informed consent 6 patients were randomized 3 patients were assigned to receive IFN-3 patients were assigned to receive placebo 3 patients were included in the analysis at day 28 3 patients were included in the analysis at day 28 5 patients who did not meet the inclusion criteria were treated with IFN-as "therapy of last resort" In total, 11 patients were analyzed : 3 study patients and 5 "last resort" patients received IFN-5 patients were included in the analysis at day 28 Figure 1 Screening, randomization, and follow-up of the study patients. The principal investigator was immediately notified when Candida spp. were cultured in blood. With at least one systemic inflammatory response syndrome (SIRS) symptom present in the 24 hours prior to blood culture withdrawal, and administration of systemic antifungal therapy < 72 hours, patients were deemed eligible for the 'IFN-γ as an adjunctive treatment for candidemia' pilot-study. In addition, 5 patients not meeting inclusion criteria but who were also treated with rIFN-γ as a therapy of last resort, were included in analysis.

Clinical outcome

The three patients in the control group and five out of eight patients treated with rIFN-γ recovered uneventfully from the fungal infection (Table 1). Two patients with invasive aspergillosis that were already admitted to the ICU at the time of treatment died due to infectious complications of severe pulmonary aspergillosis, despite rIFN-γ treatment. The patient with a Candida endocarditis, who despite rIFN-γ treatment developed intracerebral mycotic aneurysm, could be discharged from the hospital 93 days after onset of invasive candidiasis. In all patients treated, rIFN-γ was well tolerated. Five patients reported moderate fever upon administration of rIFN-γ, which responded well to acetaminophen. Two patients developed liver enzyme abnormalities for which tuberculostatic antibiotics and voriconazole were temporarily discontinued, resulting in recovery of the liver enzyme abnormalities while rIFN-γ treatment was continued. No other significant adverse events were observed.

Effect of rIFN-γ on ex-vivo IL-1β and TNFα production

To assess the effect of rIFN-γ on the capacity of PBMCs to produce pro-inflammatory cytokines, cells were isolated and stimulated before, during, and after treatment. We monitored the fold change in cytokine production compared with baseline (before start of treatment). IL-1β and TNFα are pro-inflammatory cytokines of the innate immune system crucial in the induction and maintenance of the anti-fungal immune response [START_REF] Eyerich | IL-22 and TNF-alpha represent a key cytokine combination for epidermal integrity during infection with Candida albicans[END_REF][START_REF] Gresnigt | Pattern recognition receptors and their role in invasive aspergillosis[END_REF][START_REF] Sainz | TNFR1 mRNA expression level and TNFR1 gene polymorphisms are predictive markers for susceptibility to develop invasive pulmonary aspergillosis[END_REF][START_REF] Warris | Invasive pulmonary aspergillosis associated with infliximab therapy[END_REF][START_REF] Bellocchio | The contribution of the Toll-like/IL-1 receptor superfamily to innate and adaptive immunity to fungal pathogens in vivo[END_REF][START_REF] Sainz | IL1 gene cluster polymorphisms and its haplotypes may predict the risk to develop invasive pulmonary aspergillosis and modulate C-reactive protein level[END_REF]. Before IFN-γ treatment, inter-patient variability in cytokine production was high (e.g. TNFα median [IQR] concentration after stimulation with LPS was 792 pg/mL ). Nevertheless, in all patients an increase in the capacity to induce different cytokines was observed in the first two days after initiation of IFN-γ treatment, independent of their baseline values (group data shown in Figure 2), at subsequent time points only a trend towards increased could be observed. In contrast, the placebo-treated patients IL-1β and TNFα responses over time remained similar to baseline. The response against hyphae of C. albicans was highly variable between patients. Some rIFN-γ-treated patients demonstrated a profound increase of TNFα production after treatment (up to 70 fold), whereas other patients showed no relevant change in TNFα production. Cytokine production remained similar in patients in the control group.

Effect of rIFN-γ on ex-vivo IL-17 and IL-22 production

Both IL-17 and IL-22 are cytokines that are thought to be protective in the host defence against invasive fungal infections [START_REF] Eyerich | IL-22 and TNF-alpha represent a key cytokine combination for epidermal integrity during infection with Candida albicans[END_REF][START_REF] De Luca | IL-22 defines a novel immune pathway of antifungal resistance[END_REF][START_REF] Gessner | Dectin-1-dependent interleukin-22 contributes to early innate lung defense against Aspergillus fumigatus[END_REF][START_REF] Lin | Th1-Th17 cells mediate protective adaptive immunity against Staphylococcus aureus and Candida albicans infection in mice[END_REF][START_REF] Milner | Impaired T(H)17 cell differentiation in subjects with autosomal dominant hyper-IgE syndrome[END_REF][START_REF] Smeekens | Low interleukin-17A production in response to fungal pathogens in patients with chronic granulomatous disease[END_REF]. PHA-induced IL-17 and IL-22 production was increased 1 day after initiation of rIFN-γ treatment (Figure 3). However, at subsequent time points a trend towards increased IL-17 and IL-22 production was observed, which reverted to baseline levels at day 28. Production of IL-17 and IL-22 upon stimulation with Candida blastoconidia was elevated after rIFN-γ treatment in 6 of 8 patients. Hyphae induced IL-17 and IL-22 production was increased in 4 of 8 and 5 of 8 patients respectively. Patients who received placebo therapy did not display a trend towards increased IL-17 or IL-22 production during the course of treatment (group data shown in Figure 3).

Effect of rIFN-γ on ex-vivo IL-10 production

In addition to pro-inflammatory cytokines, the capacity to produce anti-inflammatory cytokines can also influence disease outcome. In particular the anti-inflammatory cytokine IL-10 has been associated with protection against immunopathology during severe infections. IL-10 production in response to stimulation with LPS, PHA and Candida was highly variable between patients and did not show a distinct pattern following rIFN-γ treatment (Figure 4). No relevant differences compared to the placebo-treated patients were observed.

HLA-DR expression

The numbers of HLA-DR-positive monocytes, a marker of immunosuppression, varied substantially between patients at baseline (39.05% [27.5-61.6] vs. 90. 6 [88.7-92.5] in IFN-γ-treated patients and placebo-treated patients, respectively). Five out of eight IFN-γ treated patients exhibited HLA-DR positive monocyte levels below the "immunoparalysis threshold" of 50% and in these patients, an increase of HLA-DR-positive monocytes after IFN-γ treatment between 10% and 44% was observed which persisted throughout the study period (Figure 5).

Patients with a baseline HLA-DR expression higher than 50% did not show a change in expression. The patient with a HLA-DR-expression <50% who did not show increased levels of HLA-DR positive monocyte numbers at any time point, was one of the two patients who died due to infectious complications. No correlation was found between the level of mHLA-DR expression and TNFα production of LPS-stimulated PBMCs. An inverse correlation of baseline mHLA-DR levels with severity of underlying illness and tissue involvement was found (with higher mHLA-DR levels in patients with only impaired physical barriers, e.g. due to indwelling catheters, compared to patients with impaired immune responses, e.g. due to chemotherapy, immune suppressive agents, bone marrow disease; data not shown because this compromises patients anonymity).

Cell populations

There were no significant changes in the total leukocyte and granulocyte numbers in rIFN-γ-treated patients (Additional file 3: Figure S2A). Monocyte counts significantly increased one week after initiation of rIFN-γ therapy (Additional file 3: Figure S2C) and lymphocyte numbers significantly increased at 2 and 7 days after initiation of rIFN-γ therapy (Additional file 3: Figure S2D), which could be attributed to slight changes in CD4 lymphocytes (Additional file 3: Figure S2E), B-lymphocyte (Additional file 3: Figure S2F) and NK-cell numbers (Additional file 3: Figure S2G) and a significant increase of CD8 lymphocytes (Additional file 3: Figure S2H). No clear changes in leukocyte (subset) counts were observed in placebo-treated patients.

Discussion and conclusions

While several small clinical trials illustrated the beneficial clinical effects of adjuvant treatment with IFN-y, the proposed immunostimulating effect of IFN-γ as the mechanism of action has not been investigated. In this case series we demonstrate for the first time that adjunctive immunotherapy with rIFN-γ improves the leukocyte immune responses in patients with severe invasive fungal infections. This was primarily reflected by increased ex-vivo pro-inflammatory cytokine responses of the innate immune system such as IL-1β or TNFα, as well as an increased production of the T-cell cytokines IL-17 and IL-22, which are known to play an important role in the anti-fungal host defence [START_REF] Eyerich | IL-22 and TNF-alpha represent a key cytokine combination for epidermal integrity during infection with Candida albicans[END_REF][START_REF] De Luca | IL-22 defines a novel immune pathway of antifungal resistance[END_REF][START_REF] Gessner | Dectin-1-dependent interleukin-22 contributes to early innate lung defense against Aspergillus fumigatus[END_REF][START_REF] Lin | Th1-Th17 cells mediate protective adaptive immunity against Staphylococcus aureus and Candida albicans infection in mice[END_REF][START_REF] Milner | Impaired T(H)17 cell differentiation in subjects with autosomal dominant hyper-IgE syndrome[END_REF][START_REF] Smeekens | Low interleukin-17A production in response to fungal pathogens in patients with chronic granulomatous disease[END_REF], and by an increase in HLA-DR expression in mHLA-DR expression in those patients with a low cellular expression as a measure of their immune suppression.

In addition to enhanced ex-vivo responses, subtle changes in the leukocyte differentiation were observed following IFN-γ treatment. Although there were no significant differences in total leukocyte numbers after treatment with rIFN-γ, shifts in leukocyte subpopulations such as increased monocyte and lymphocyte counts were apparent. While lymphocyte numbers increased after rIFN-γ therapy, it could not directly be attributed to a specific subset as all of them showed increased values. The most significant increase was that of CD8 cells one week after initiation of rIFN-γ therapy. Monocytes and lymphocytes are known to be crucial cells in the host defence against fungal infections. However, the increase of monocytes and lymphocytes during rIFN-γ therapy was accompanied by slightly decreased circulating granulocyte numbers. It is not known whether this reduction is due to activation and migration into the infected tissue, or whether a true decrease in granulocyte generation was induced by the treatment. Although the decrease in granulocyte numbers was slight, the fact that granulocytes, and especially neutrophils, are crucial in the antifungal host defence warrant careful monitoring of granulocyte numbers during IFN-γ treatment.

Several clinical studies and case reports have previously demonstrated beneficial effects of rIFN-γ in combination with antifungal therapy on outcome of fungal infections (for example in patients with CGD (n = 130) [START_REF] Group | A controlled trial of interferon gamma to prevent infection in chronic granulomatous disease[END_REF][START_REF] Saulsbury | Successful treatment of aspergillus brain abscess with itraconazole and interferon-gamma in a patient with chronic granulomatous disease[END_REF][START_REF] Pasic | Aspergillus osteomyelitis in chronic granulomatous disease: treatment with recombinant gamma-interferon and itraconazole[END_REF], HIV (n = 173) [START_REF] Riddell | A phase III study of recombinant human interferon gamma to prevent opportunistic infections in advanced HIV disease[END_REF][START_REF] Bodasing | Gamma-interferon treatment for resistant oropharyngeal candidiasis in an HIV-positive patient[END_REF][START_REF] Jarvis | Adjunctive interferon-gamma immunotherapy for the treatment of HIV-associated cryptococcal meningitis: a randomized controlled trial[END_REF], leukaemia (n = 5) [START_REF] Poynton | Interferon gamma and granulocytemacrophage colony-stimulating factor for the treatment of hepatosplenic candidosis in patients with acute leukemia[END_REF][START_REF] Dignani | Immunomodulation with interferon-gamma and colony-stimulating factors for refractory fungal infections in patients with leukemia[END_REF], and transplant patients (n = 7) [START_REF] Teo | Exogenous interferon-gamma immunotherapy for invasive fungal infections in kidney transplant patients[END_REF], in a patient with S. aureus liver abscess and invasive C. albicans infection [START_REF] Malmvall | Successful interferon-gamma therapy in a chronic granulomatous disease (CGD) patient suffering from Staphylococcus aureus hepatic abscess and invasive Candida albicans infection[END_REF], in a patient with intracerebral aspergillosis [START_REF] Ellis | Massive intracerebral aspergillosis responding to combination high dose liposomal amphotericin B and cytokine therapy without surgery[END_REF], in two patients with progressive chronic pulmonary aspergillosis [START_REF] Kelleher | Interferon-gamma therapy in two patients with progressive chronic pulmonary aspergillosis[END_REF], and in two patients with idiopathic CD4 lymphopenia and cryptococcal meningitis [START_REF] Netea | Two patients with cryptococcal meningitis and idiopathic CD4 lymphopenia: defective cytokine production and reversal by recombinant interferon-gamma therapy[END_REF]). However, in contrast to our study, ex-vivo immune responses in these patients were not investigated. Due to the limited number of patients and the very heterogeneous population, we could not assess clinical endpoints, although a mean mortality of 25% in the IFN-γ treated patients lies below the mean 40% estimated in patients with invasive fungal infections [START_REF] Kriengkauykiat | Epidemiology and treatment approaches in management of invasive fungal infections[END_REF][START_REF] Pappas | NIAID Mycoses Study Group: A prospective observational study of candidemia: epidemiology, therapy, and influences on mortality in hospitalized adult and pediatric patients[END_REF].

To the best of our knowledge, we are the first to describe mHLA-DR expression, a widely used marker of immunosuppression in (bacterial) sepsis patients [START_REF] Leentjens | Immunotherapy for the adjunctive treatment of sepsis: from immunosuppression to immunostimulation. Time for a paradigm change?[END_REF], in patients with invasive fungal infections. In all IFN-γ treated patients who showed baseline mHLA-DR levels below the immunoparalysis threshold of 50% and survived, IFN-γ mediated upregulation of mHLA-DR expression was observed. In agreement with the data presented in this case series, rIFN-γ has been shown to significantly increase numbers of HLA-DR-positive monocytes both in a human preclinical bacterial sepsis model and in septic patients [START_REF] Leentjens | Reversal of Immunoparalysis in Humans In Vivo: A Double-Blind, Placebo-controlled, Randomized Pilot Study[END_REF][START_REF] Docke | Monocyte deactivation in septic patients: restoration by IFN-gamma treatment[END_REF]. Reduced production of TNFα by leukocytes ex-vivo stimulated with LPS has also been shown to be marker of immunoparalysis in sepsis patients. In contrast to our study, mHLA-DR expression and ex-vivo TNFα production were found to be highly correlated in bacterial sepsis patients [START_REF] Docke | Monocyte deactivation in septic patients: restoration by IFN-gamma treatment[END_REF][START_REF] Carson | Epigenetic regulation of immune cell functions during post-septic immunosuppression[END_REF]. A possible explanation for this discrepancy is that, in contrast with the emerging consensus that immunoparalysis renders patients more vulnerable to opportunistic infections in general [START_REF] Leentjens | Immunotherapy for the adjunctive treatment of sepsis: from immunosuppression to immunostimulation. Time for a paradigm change?[END_REF], different defects in immune defences may be responsible for enhanced susceptibility towards different pathogens.

Based on the apparent inverse correlation of baseline mHLA-DR levels with severity of underlying illness and tissue involvement, mHLA-DR levels seem to reflect disease severity and general immune status, and not specific immune defects per se. Hence, patients with invasive fungal infections and associated impaired antifungal immune responses will probably benefit more from immunostimulatory treatment compared to patients with only impaired physical barriers, e.g. due to indwelling catheters and apparent intact anti-fungal immune responses. Biomarkers reflecting the capacity of specific anti-fungal immune defences are required to identify patients who suffer from invasive fungal infections due to impaired cell-mediated immunity. It is important to identify such patients and attempt a tailored immunotherapeutic approach guided by the actual level and type of immunoparalysis of that specific patient. A blood based assay has been described that demonstrates a failure to induce IFN-γ expression in renal transplant patients and differences in IL-10 and TNFα expression [START_REF] Teo | Renal allograft recipients fail to increase interferongamma during invasive fungal diseases[END_REF], which could be promising biomarkers to identify patients who could benefit from adjunctive immunotherapy.

The intracellular mechanism(s) through which the beneficial effects of IFN-γ are mediated remain to be elucidated. Recently it was proposed that IFN-γ exerts its effects at the transcription level [START_REF] Turrel-Davin | mRNA-based approach to monitor recombinant gamma-interferon restoration of LPS-induced endotoxin tolerance[END_REF], while others have demonstrated that IFN-γ reverses tolerance-associated epigenetic modifications [START_REF] Chen | IFN-gamma abrogates endotoxin tolerance by facilitating Toll-like receptor-induced chromatin remodeling[END_REF]. Another possible mechanism involved in the IFN-γ mediated reversal of immunoparalysis is the downregulation of negative TLR regulators such as IRAK-M, a protein that negatively regulates LPSinduced inflammatory responses and contributes to the development of immunoparalysis [START_REF] Xiong | Induction of endotoxin tolerance in vivo inhibits activation of IRAK4 and increases negative regulators IRAK-M, SHIP-1, and A20[END_REF].

Administration of rIFN-γ was tolerated well. Several patients developed a mild fever upon administration, which responded well to acetaminophen treatment. No other side effects were observed. The most important limitation of the present study is the limited number of patients studied. Because the control group consisted of only three patients, no statistical analysis between the treatment and control groups could be performed. However, despite the small sample size, the increase in HLA-DR expression in patients with mHLA-DR expression levels below 50% and the increased ex-vivo response of several cytokines that are crucial in antifungal host defence is a promising observation that underlines the potential of immunotherapy. The slow enrolment of patients presenting with candidemia was the main factor contributing to the decision to terminate the phase IIIb Candida pilot-study early. With a reported incidence of 2.5-11 per 100,000 persons in Europe [START_REF] Tortorano | Candidaemia in Europe: epidemiology and resistance[END_REF], and based on previous epidemiological data in our hospital this low enrollment was not expected at the time of the initiation of the study. The much lower incidence of candidemia in the last two years in our hospital is most likely due to a new antibiotic stewardship introduced recently in our hospital, which has reduced the incidence of opportunistic infections. The cut-off value of mHLA-DR expression levels of 50% to distinguish between immunoparalyzed and immunocompetent patients is another limitation of this study, as this is an arbitrary value chosen. We chose this value because it is well below the 99% CI of mHLA-DR values in healthy volunteers [START_REF] Leentjens | Reversal of Immunoparalysis in Humans In Vivo: A Double-Blind, Placebo-controlled, Randomized Pilot Study[END_REF]. Therefore, patients with mHLA-DR below 50% do have an impaired antigen presenting capacity of their monocytes which we show to be enhanced by IFN-γ therapy. Whether this cut-off value truly represents immunoparalysis, reflected by enhanced susceptibility to secondary infections or reduced capacity to clear opportunistic infections, remains to be investigated. Furthermore, the use of a standardized analysis technique to quantify mHLA-DR, such as the Quantibrite method, is preferable, because it facilitates an objective comparison of mHLA-DR expression levels between studies and aids in the definitive establishment of a cut-off value to identify immunoparalyzed patients. Larger studies are required to confirm the data obtained here. To do so, multicentre studies should be facilitated in order to fully explore the potential of IFN-γ immunotherapy.

Our data indicate that adjunctive immunotherapy with rIFN-γ in patients with invasive fungal infections partially restores cell-mediated immunity. This suggests that IFN-γ treatment enhances anti-fungal immunity and larger studies are warranted to validate the findings reported here and to assess the impact of IFN-γ treatment on clinical outcome. Biomarkers of impaired anti-fungal immunity should be further investigated in order to identify patients who will benefit most from immunostimulatory therapy.

Figure 2

 2 Figure 2 Effect of rIFN-γ on ex-vivo IL-1β and TNFα production. PBMCs of patients were isolated at baseline and day 1, 2, 7, 14 and 28 after rIFN-γ administration. Isolated PBMCs were stimulated for 24 hours with LPS, PHA, C. albicans blastoconidia, or C. albicans hyphae. IL-1β (A) and TNFα (B) concentrations were measured in culture supernatants. Baseline concentrations were used as control and set at 1; subsequent measurements are plotted as the mean relative fold change ± SEM. Significant change from baseline was determined by subjecting the data to Wilcoxons signed rank test. (* = p < 0.05; ** = p < 0.01).

Figure 3

 3 Figure 3 Effect of rIFN-γ on ex-vivo IL-17 and IL-22 production. PBMCs of patients were isolated at baseline and day 1, 2, 7, 14 and 28 after rIFN-γ administration. Isolated PBMCs were stimulated for 7 days with PHA, C. albicans blastoconidia, or C. albicans hyphae. IL-17 (A) and IL-22 (B) concentrations were measured in culture supernatants. Baseline concentrations were used as control and set at 1; subsequent measurements are plotted as the mean relative fold change ± SEM. Significant change from baseline was determined by subjecting the data to Wilcoxons signed rank test. (* = p < 0.05).

Figure 4

 4 Figure 4 Effect of rIFN-γ on ex-vivo IL-10 production. PBMCs of patients were isolated at baseline and day 1, 2, 7, 14 and 28 after rIFN-γ administration. Isolated PBMCs were stimulated for 48 hours with LPS, PHA, C. albicans blastoconidia, or C. albicans hyphae. IL-10 concentrations were measured in culture supernatants. Baseline concentrations were used as control and set at 1; subsequent measurements are plotted as the mean relative fold change ± SEM.

Figure 5

 5 Figure 5 mHLA-DR expression in rIFN-γ treated patients, divided into immunoparalyzed patients with baseline HLA-DR expression below 50% (solid dots), and without HLA-DR defined immunoparalysis (open dots). Data are expressed as median [IQR].

Table 1

 1 Summary of clinical characteristics of all patients with invasive fungal infections venous catheters (for the need of recurrent blood sampling or total parental nutrition), or an implantable cardioverter-defribillator (ICD) lead. The patient with progression of suspected hepatic Candida lesions on the CT-scan, and one patient with proven acute aspergillosis (diagnosed by isolation of A. fumigatus from lung tissue biopsies) were immunocompromised because of (therapy for) acute myeloid leukaemia. Another patient with acute invasive aspergillosis (A. fumigatus isolated from BAL fluid) received immunosuppressive therapy (prednisone and azathioprine) for sarcoidosis and suffered from a coinfection with Mycobacterium genavense localized in the bone marrow. A third patient developed chronic pulmonary aspergillosis (diagnosed by consistent CT abnormalities with cavitation and culture of A. fumigatus from BAL fluid) after radiotherapy for lung carcinoma. Only two patients, both with acute aspergillosis, were admitted to the Intensive Care Unit to receive organ supportive therapy (mechanical ventilation and hemodynamic support).

	IFN-γ treated patients

indwelling

Acknowledgements

This research was performed within the framework of CTMM, the Center for Translational Molecular Medicine (www.ctmm.nl), project MARS (grant 04I-201). M.G.N. was supported by a Vici grant of the Netherlands Organization for Scientific Research and an ERC Consolidator grant of the European Research Council (nr. 310372). F.vd.V. was supported by a Veni grant of the Netherlands Organization for Scientific Research.

Additional files

Additional file 1: Online Supplement Flow cytometric analysis of mHLA-DR expression and lymphocyte subset counts.

Additional file 2: Figure S1. Representative flow diagram of monocyte HLA-DR measurements. Heparin blood was first analysed on forward-and side scatter to exclude cell debris and erythrocytes (A). Subsequently, CD45 + cells were selected (B) and within the CD45 + fraction was gated for CD14 + cells (C). The CD45 + CD14 + cells (D) were analysed for the percentage of HLA-DR positivity (E).

Additional file 3: Figure S2. Changes in immune cell populations. Total leukocyte numbers (A) and numbers of granulocytes (B), monocytes (C) and lymphocytes (D) measured in peripheral blood. Numbers of CD4 lymphocytes (E), B-lymphocytes (F), CD8 lymphocytes (G) and NK cells (H) within the lymphocyte population were quantified using flowcytytometry.

Competing interests

None of the authors have any competing interests regarding this study. The support for the immunological assessments in this study was provided by an unrestricted grant from BioMérieux.


domain: sdv sde

text:
Sylvain Bart

Joël Amossé

Alexandre R R Pery

Christian Mougin

Céline A Pelosi

à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

A. caliginosa

No avoidance

Number of times the recommended dose (RD)

A worm story by Sylvain BART, But also Joël Amossé, Alexandre Péry, Christian Mougin and Céline Pelosi UMR ECOSYS, INRA, AgroParisTech, Université Paris-Saclay, 78026, Versailles, France However, the intensification of agricultural practices passed through ... and in particular the use of pesticides, which endangered their jobs ...

Impacts of pesticides on an earthworm population

Yes, I am an ecosystem "engineer" (without master's degrees! ). I am involved in key ecological functions

Swing Gold®:

Dimoxystrobin (133 g L -1 ) Epoxiconazole (50 g L -1 ) I would loved to eat a burger with soil and wheat roots ! Oh, some clays.. I feel someone is looking at me oh, a stone… ZZZZzzzz …

Thus, an earthworms species, very common in cultivated fields, volunteered to represent its colleagues.

Cuprafor micro®:

Copper oxychloryde (50%)

IT

And on reproduction?

That's the way they reproduce … And they do it under our feet !!!

A. caliginosa, as earthworms, is hermaphrodite. He produces 1 -2 cocoons per week in our laboratory.

We will assess the impact of copper and Swing Gold® on cocoon production, but also on the cocoons themselves (time and percent of hatching).

??

Let

  C. Laurent, A. R. R. Pery, C. Mougin and C. Pelosi (2017). Differences in sensitivity between earthworms and enchytraeids exposed to two commercial fungicides. Ecotoxicol Environ Saf 140: 177-184.

  's see what's going on Percentage of avoidance of A. caliginosa, exposed to differents Cuprafor micro® concentration (copper, left) and Swing Gold® (right). (Bart et al., 2017) Conceptual design of a population dynamic model (IBM model : Individual Based Model)

Acknowledgements: I would like to thank the funders of my PhD thesis : INRA and the IDEX Paris-Saclay. I also thank ECOSYS and in particular the members of the « pôle écotoxicologie » for their scientific and technical support: Jodie, Jean Pierre, Véronique, Amélie, Sébastien, Ghislaine, Sylvie and many students.

Do they avoid pesticides?

Which impacts on growth?

What to do finally?

Based on experimental data, we will calibrate a Dynamic Energy Model (DEB), which will simulate the life cycle (growth, reproduction) of A. caliginosa.

From the DEB model and avoidance data, we will calibrate an Individual Based Model (IBM) to predict effects of pesticides on population dynamics in field conditions. This model will account for interactions of individuals with their environment (food, temperature, moisture,…) and between each other, as represented by the red arrows.

Switch at the population level

Avoidance

Copper concentration (mg kg -1 )