Approaches in Design of Laboratory-Scale UASB Reactors

: Up-ﬂow Anaerobic Sludge Blanket (UASB) reactors are popular tools in wastewater treatment systems due to the ability to work with high feed rates and wastes with high concentration of organic contaminants. While full-scale industrial applications of UASB reactors are developed and described in the available literature, laboratory-scale designs utilized for treatability testing are not well described. The majority of published studies do not describe the laboratory UASB construction details or do use reactors that already had developed a trophic network in microbial consortia under laboratory environment and therefore are more stable. The absence of deﬁned guidelines for geometry design, selection of materials, construction, operation rules, and, especially, the start-up conditions, signiﬁcantly hamper researchers who desire to conduct treatability testing using UASB reactors in laboratory scale. In this article, we compiled and analyzed the information available in the refereed literature concerning UASB reactors used in laboratory environment, where information on geometry and / or operational conditions were provided in detail. We utilized the information available in the literature and the experience gained in our laboratory (Sustainable Waste to Bioproducts Engineering Center) to suggest a uniﬁed operation ﬂowchart and for design, construction, operation, and monitoring for a laboratory-scale UASB reactors.


Introduction
Up-flow Anaerobic Sludge Blanket (UASB) reactor is an anaerobic digester for wastewater treatment, and its operational concept can be described as a vertical up-flow pumping of liquid substrate, including wastewater or growth media, through a layer of anaerobic sludge [1][2][3][4][5][6]. Microbial consortia inside the sludge layer consume digestible components as substrate and decompose them into smaller chemical compounds [7]. Within the scope of a wastewater treatment, the goal of anaerobic digestion is a complete mineralization of organic compounds combined with the production of biogas for the purpose of energy recovery.
A distinguishing feature of UASB reactors is the formation of microbial conglomerates, where the metabolic product of one microbial group is a consumable substrate for another microbial group [8]. Such microbial conglomerates grow into spherical or bean-shaped granules over time [9][10][11][12][13]. The sizes of granules may vary, but typically are reported in the range 0.5 to 6 mm, where longer operation leads to larger sizes [14][15][16]. Granulation of sludge is promoted by the presence of microorganisms that are able to produce and secrete Exocellular Polymeric Substances (EPS) [17]. The term "EPS" includes multiple types of compounds, which serve as a glue to agglomerate microorganisms together and to add some mechanical strength to a granule [18]. The construction concept of the GLSS is shown in Figure 1a, where it's implemented via narrowing the outlet of the reaction tube with baffles. Such baffles are typically referred to as "deflectors" or "collar". The side effect of narrowing the reaction tube outlet is a creation of local velocity gradient (velocity shear), which slightly enhances the formation of granulated particles, their separation from liquid and settling back to the bottom of the reactor. Above the baffles, the GLSS contains the gas collecting structure, where the cross-section looks like a flipped upside-down funnel. In some studies, this funnel is replaced by a tubular structure with diameter larger than the distance between baffles [27,28]. The liquid is forced to flow through the space in between the lower edge of the gas collector and the baffles, to go around the funnel and leave the reactor at the effluent port.
Other existing modifications of GLSS in laboratory-scale reactors can improve the higher solids retention time, such as installing a high rate settler in headspace [29] or modification of three-phase separators [30].
In addition to the operational concept of the UASB reactor shown in Figure 1a, the same authors [11] also describe UASB reactor with a modified gas collector, which is demonstrated in Figure 1b. However, some studies [31] call such a modification of the Up-flow Anaerobic Sludge Baffled Reactor (UASBR). It may also contain the inner mechanical agitation device to prevent foam formation in the gas collecting area [32]. Recently, the Y-shaped variation of UASB reactor also became popular and is depictured in Figure 1c. In the case of the Y-shaped reactor, the GLSS is split into two individual separators: one separator is used to separate gas from the liquid and collect it directly at the top of a main tube, whereas a second collector is a sidearm tube that serves as an inclined settler for separating solids from liquid (similar to a Lamella clarifier). Use of a funnel-shaped gas-collecting element becomes optional in such case, since it serves only the purpose of preventing gas flow to an effluent side-arm.
Considering the concepts described, the optimization goal of a laboratory scale UASB reactor operation is to achieve better performance, where optimization targets for UASB performance include the following: • Higher removal of contaminants; • Higher biogas production rate; • Shortening of adaptation period; and • Resilience (robustness) of sludge.
To achieve some of those optimizations, the classical UASB concept can be combined with other types of reactors, resulting in a range of composite reactors. Some modifications are found in the literature and are presented in Table 1. This table represents options, where another reactor type is incorporated into the UASB itself, but not a sequence of two consecutive reactors.  [40] In a holistic view, the purpose of UASB reactor optimization is to keep the microorganisms in a stage of maximum substrate consumption and active growth. However, from an operational perspective, the optimization of UASB functioning is achievable via adjusting operational parameters, including, but not limited to: Despite the long history since the invention and description of the UASB concept by Lettinga et al. [41] and increasing its application in industry, UASB laboratory scale reactors used for treatability studies are highly variable with regard to terminology, design, construction, and operation processes. This lack of uniformity leads to different results regarding water quality indicators, for example, Chemical Oxygen Demand (COD), as well as bioenergy production, for example for biomethane and biohydrogen. There is a lack of uniformity with regard to the guidelines for operation of laboratory scale conditions, which is highlighted in this manuscript and recommendation are provided for making UASB laboratory studies and results more uniform with results more transferrable among laboratories and more useful for scale up activities. These lack of uniformity with laboratory scale UASB reactors is addressed in this study and guidelines are provided for increasing the uniformity so that results are comparable across different laboratories and are also more meaningful for scale up applications of the UASB reactor process.

Review of Existing Solutions across Various Published Works
Despite a large number of available publications on wastewater treatment involving UASB reactors, a majority of the studies only briefly mentions constructional concepts of the reactor, and dimensional parameters are mentioned even more rarely. We collected available information on physical dimensions among existing studies in Table 2, while Table 3 shows the geometry of either hybrid reactors or where UASB reactors are installed in series with any other reactor. While building those tables we focused on the geometry of the reactor and operational conditions including substrate strength expressed as COD, Biological Oxygen Demand (BOD), etc.; loading rates; volume of reactor; and effluent recycling rates. The type of the substrate used in reported studies is provided for reference purpose only. Where Table 2 does not contain the geometric or operational parameter means that such value was not specified in the reference. Also, Tables 2 and 3 do not provide calculations based on available geometry. All information provided there is information stated in referenced publications, nothing was added. The only modifications were made to units (for COD, BOD, etc.) where they were unified across all publications.
As we can see from Tables 2 and 3, there is no uniformity in parameters of operating the UASB reactor and, and what is in our opinion even more important, information on the start-up of a laboratory UASB reactor. Such inconsistency may complicate the interpretation of results as well as the accuracy and successfulness of an experiment in general. On the larger scale, it also complicates the comparison of results obtained by different laboratories, which creates problems for feasibility studies, if the literature is the primary source of information. To be more precise, in case of a failure, incomplete information does not allow an interpretation of the data and to trace-back the reason for failure, such as unadopted inoculum or its insufficient amount, problem of biomass washout due to the geometry, problematic substrate properties, or wrong OLR or recycle ratio. Inconsistent reporting units (like OLR calculated as per total volume of reactor or per volume of digestion zone) may also harm the attempt to reproduce results of one laboratory in another one, or wrong implementation of a procedure.
Among the inconsistencies found across the studies, we also see terminology problem in the use of terms 'sludge blanket' and 'sludge bed'. The controversy of the terms is in that fact that they are: 'sludge bed' refers to a layer of sludge at the bottom, where it is concentrated and visually seems to be a packed layer, while 'sludge blanket' refers to a part of the reactor where sludge is swimming as flocs above the 'sludge bed' • 'sludge bed' refers to a bottom layer of sludge, and uses 'transition zone' instead of a 'sludge blanket' Below, in Table 4, we attempted to systematize all parameters we were able to identify in the publications reviewed. Information in Table 4 does not intend criticize, but instead the intent is to generalize and categorize information from publications referenced above.              Table 4. Summary of the geometry and operational parameters for existing UASB reactor designs.

Criteria Options/Area of Application/Observations
Height No constraints on height. The smallest found reactor was 30 cm tall, the largest as above 4 m. Perhaps, limited only by the available space in a laboratory. Volume Small volumes are 0.5, 0.75, 1, and 2-2.4 L. Larger volumes of 14 and 55 L were also found. Usually, reactors with volume greater than 1 m 3 are referred as pilot-scale. Height: Diameter (H:D) ratio Since the substrate has an up-flow velocity, the reaction part of UASB reactor in some degree functions as a sedimentation or coagulation column, where ratio H:D should help preventing the biomass washout [19]. This parameter is very rarely reported, and reporting of it can be confusing due to not clear geometry reference. There are reports of H:D ratio as a diameter of a reactor to either a total height of a reactor or to the height of a reactor without GLS. We see reasonable to calculate it as a diameter of reactor to the height without GLSS, since the goal of GLSS is to create a chamber for gas capture above the reaction tube of a reactor. From review studies, such ratio for majority of cases is in range from 8 to 14. However, there are also extreme cases as 3.5 or 23. The particular design varies with the concept of the reactor itself, and options can be split into: Implementation of baffles Gas collection For tubular reactor designs, the deflectors are typically made as an O-rings with a triangular cross-section. For rectangular reactors, a series of inclined baffles are installed to narrow a main liquid flow. For smaller reactors, baffles are sometimes omitted, probably, because it's difficult to implement those in smaller volumes. Another case when deflectors were noticed to be omitted is when GLSS is represented by a separate part (either tube or funnel), wider than the major reaction tube, and a diameter of a gas collector is close to a diameter of a reaction tube. Gas collector is usually represented by a flipped upside-down funnel for smaller reactors. For larger ones, it can be a separate compartment. Y-shaped reactors do not have any specific structure inside. Heating Among the reviewed designs the following heating systems were noticed: Heating pads or tapes Water jacket No heating Inner heaters (helix shaped) Water jacket is the most common option for smaller designs but it complicates the placement/insertion of sensors (like pH, ORP, temperature, etc.) into a reactor. Larger reactors usually use heating pads or a combination of heating pads with thermal insulation material. Temperature ranges Mesophilic: 35 ± 2 • C or 37 ± 1 • C Thermophilic: 55 ± 1 • C Ambient temperature Ambient temperature with thermostat to prevent overcooling Inoculum material No constraints: Granular or flocculated sludge from another UASB Non-granulated anaerobic or active sludge Adjusted inoculum from non-sludge sources, like animal manure Seeding (inoculating) Across the reviewed studies, this was the most inconsistent parameter, which was not even always reported. The process was reported as: (a) filling reactor with raw sludge up to a certain percentage of height; (b) volumetric load of sludge per reactor, sometimes mentioning its VSS and/or TSS equivalent; and (c) final concentration of sludge in reactor as TSS or VSS. Also, few studies suggested to sieve the sludge through 1-3 mm mesh to remove any undigested particulate or residuals before seeding.

Substrate preparation, feeding and pH management
Few studies considered the adjustment of substrate based on ratio COD:N:P. However, the final ratio does not match across publications and varies for COD parameter 300-600:5:1. Surprisingly, no-one mentioned adjusting the C:N ratio, which is recommended for anaerobic treatment in general. Only one publication mentioned the addition of compounds to stimulate granulation. Substrate pH management Researchers use either pH adjustment in substrate directly or pumping pH adjusting solution to the reactor. Used adjusting compounds are either hydroxides or bicarbonates. Interesting fact: addition of 0.5-3 g of NaHCO 3 per 1 L of substrate was sufficient to maintain a stable effluent pH around 7. In some extreme cases 8 g per 1 L of substrate were sufficient to work with OLR 150 kgCOD m 3 ·day . OLR and HRT HRT and OLR are interdependent values and both are optimization points in research. Researchers aim to increase OLR and decrease HRT. These parameters are points of inconsistent reporting: Some sources report OLR and HRT as referred to the total volume of the reactor (both reaction tube and GLSS) Some sources report OLR and HRT as referred to the volume of the reactor without the volume of GLSS Higher limit for OLR is not specified, since it depends on chemical composition of influent wastewater and its strength.

Substrate distribution system
Typically is not reported, but where it is mentioned it's either: a circular tube with evenly distributed outlet holes and an inlet from the side through the wall of reactor an inlet into conical-shaped bottom of reactor a side inlet through the wall into bottom compartment with inclined bottom

Discussion
Studies, involving the UASB trials, are usually purposed for: (a) treatability testing and energy recovery estimation; (b) microbiology studies on changes in microbial consortia during adaptation to new substrates or long-term operation for further modeling of trophic network; or (c) toxicity and granulation process studies. In the scope of this manuscript, we would like to identify the common needs of such research and point out the differences, where it is important. Here, we would like to focus on experimental aspects, which are needed to pay attention to, while designing the reactors and its infrastructure.

Volume of Reactor
The first thing that affects the final volume of a designed reactor is the available amount of sample/substrate. Some samples of substrates are available in very limited quantities due to the policies of supplier companies or may be a subject of special regulations preventing the dumping of effluent to a sewer (Ex. industrial wastewater). Depending on the complexity of substrate and potential inhibitory effects, the reactor start-up period might occupy a substantial period up to 120 days [14,57,59,65,68], thus the volume of a reactor should allow to utilize the available sample volume for both start-up period and experiment duration.

Material of the Reactor
Due to the specifics of laboratory studies, the reactor needs to be constructed with the feature to visually inspect the content. It allows one to: (a) confirm the fact of granulation and (b) inspect the foam or scum formation, etc. This significantly narrows the selection choice of materials, limiting it to (a) polymethyl methacrylate (a.k.a., PMMA, Plexiglas, Perspex, acrylic glass), (b) borosilicate glass, and (c) clear polyvinyl chloride. Each of the mentioned materials can be used, and in our opinion it is more of a question of budget and available stock parts. We compare pros and cons of each material in Table 5. Borosilicate glass is an excellent option if used for studies with sterile cultures, since it can be autoclaved. However, in the author's opinion, the ideal reactor must be manufactured of stainless steel and be featured with an inspection window, a water jacket and multiple sampling ports. Such a design would be chemically resistant under conditions of anaerobic digestion, autoclavable, and meet multiple research needs. However, such construction complicates the customization and should be done for optimized and fully tested design after confirmation of its efficiency. The authors currently use PMMA due to machinability of this material, its transparency, and stability under conditions of anaerobic digestion (AD).

Heating of Reactor
Heating of reactor under laboratory conditions is defined by: (a) actual need for heating and (b) necessity of sampling the content of reactor and location of sampling. If no sampling of reactor content is needed, the water jacket would be the most suitable option. Otherwise, sampling ports complicate the construction of water jacket. Without a water jacket, consider: (a) use of heating tapes or flexible heating pads or (b) preheating of substrate and thermal insulation of reactor to keep the temperature.
Heating tape on the outer surface of PMMA or PVC reactor is not recommended, since it could cause local damage, when the contact point of wall material and heating tape is locally overheated, causing melting or other types of damage. Our laboratory experienced problems with heating tapes even under mesophilic conditions. The reactor that got damaged, was controlled by thermostat with an external submergible temperature sensor. The damage consisted of the tape melting through the wall of the reactor causing leakage. Thus, we moved to a water jacket in our projects.
Perhaps, the use of heating pads would be more secure due to a larger area of contact and, hence, more uniform heating. Extra uniformity may be added by use of heat-transfer pastes, but they will decrease the observability of the process in reactors. However, it is still a viable option when there is a need for the presence of sampling ports on various levels or there is no way to implement a water jacket due to other reasons.

Inoculum: Preparation, Adaptation, and Seeding
While the granulated anaerobic sludge is the desirable inoculum, authors fully realize the probability of a situation when researchers do not have a source for granulated sludge. In such a case, the manure sample of animal origin could be a source of methanogenic microbial consortia, and referenced studies [27,42,43,57,63,65] suggest self-digestion of such sample or mixing it with a substrate and conditioning for up to 3 weeks. The presence of methanogenic microorganisms is required for the generation of methane, but not every manure contains methanogens. The most typical confirmed cases of manure containing methanogens are cattle and swine manures. The presence of methanogenic bacteria could be confirmed by conducting specific methanogenic activity test [69,70], which is very close in technique to a popular Bio-Methane potential (BMP) test [71], but conducted on a nutrient media containing acetate as the only source of carbon [72].
Some studies suggest the sieving of inoculum through a 1-3 mm mesh to remove undigested or large inert material. It is reasonable, if the inoculum originated from manure, since manure samples may contain some animal bedding, or sewage wastewater treatment facility, which may contain hair, etc. However, if the sample originated from an industrial wastewater treating facility, such sieving could be optional, especially if sludge is already granulated and granules are large. Also, the effect of exposing sludge or granules to air during the sieving is not clear. Perhaps, the sieving process should be done in anaerobic chamber.
The seeding of reactor must be calculated and expressed as Volatile Suspended Solids (VSS), introduced with the inoculum, per working volume of reactor according to [73,74] and seeding should be in the range 10 to 20 kg VSS m 3 , (however, it also could be up to 25 kg VSS m 3 ) [10]. Inoculum should be analyzed for Total Solids, (TS), Volatile Solids (VS), Total Suspended Solids (TSS), and Volatile Suspended Solids (VSS) since sludge is also characterized by VS:TS and VSS:TSS ratio, as criteria of alive biomass if condition of sludge is tracked over time [75] and ratio VSS:TSS of sludge in range of 0.7 to 0.85 is likely to cause granulation [59]. The recommended method for solids content analysis is specified in Standard Methods 2540 [76].

Substrate Adjustment
Before any adjustments is done to a substrate, the treatability can be roughly characterized by the ratio BOD 5 :COD, which is referred to as a biodegradability index (BI) [77,78]. For municipal raw wastewater BI is usually in the range 0.4 to 0.8 [79,80] and it is considered to indicate good treatability. Greater index means better bio-treatability and pretreatment can increase the value of BI [81,82]. Estimation of sample degradability based on BOD 5 :COD is inconsistent, but can be generalized as: (a) highly bio-treatable if greater than 0.5; (b) bio-treatable if greater than 0.3; and (c) not bio-treatable when lower than 0.2 [81][82][83][84][85].
One of the primary adjustments for substrate is the C:N ratio [86] by mass, with the optimum in the range 25 to 30 [87][88][89] or 20 to 30 [90] and higher temperature ranges require higher C:N ratio. However, it also could be a substrate-specific optimization parameter [91][92][93]. Some authors also consider C:P ratio for methanogens between 16:1 and 75:1 [94,95] as optimal, while C:N:P ratio is considered to be favorable in the range 400:5:1 to 100:28:6 [95,96]. Some inconsistence to in attempt to meet those ranges may come from measurement techniques, where various authors use either: (a) elemental analyzers [97] or (b) total carbon and Total Kjeldahl Nitrogen (TKN) [90]. Across referenced in this manuscript studies, following compounds were used to correct ratio: KH 2 PO 4 , NaH 2 PO 4 , CO(NH 2 ) 2 (urea or carbamide), and NH 4 H 2 PO 4 .
Individual studies stated the need to account for sulphur [98], and report C:N:P:S ratio as 600:15:5:3 to be the optimal for methanization [99]. Perhaps, such increase of considered elements is reasonable, since elemental composition of anaerobic biomass is reported as C 5 H 7 O 2 NP 0.06 S 0.1 according to [100][101][102], but not yet widely used in anaerobic digestion studies.
The ratio of COD:N:P of 250:5:1 is generally suggested for anaerobic treatment [103][104][105], however, some variation exist between 900:5:1.7 and 150:5:1 [104,106,107] and could be even 300:1:0.1 [108]. Other studies recommend 300:5:1 as start-up conditions specifically for UASB [75,96,109,110]. Important to mention, that "N" in such proportion refers to the total nitrogen [108] pH adjustment for methanogenic bacteria should bring the pH in the optimal range 6.8 to 7.5, while outside of the 5 to 8.5 range, methanogenesis is fully suppressed [111][112][113]. However, for anaerobic digester the range of 6.8 to 7.2 is recommended, due to widely used in wastewater treatment lime as pH adjusting chemical [114]. Across referenced studies we noticed NaOH and NaHCO 3 as widely usable compounds to adjust pH, however, the choice is wider [115].

Granulation Stimulation
If granulation enhancement is needed, the Ca 2+ in concentration 100-200 mg/L of substrate can be added [116], or even 150 . . . 300 mg/L at the start-up [117,118]. The role of calcium in granulation process is not clear, but it is assumed to form precipitates with carbonate and phosphate [21,119]. Use of Mg 2+ is not recommended, since it causes disaggregation of granules [120], even though it is expected to precipitate as MgNH 4 PO 4 [10]. Normally, granulation should be observed within 4-6 weeks after the start of the experiment [73].

Start-up Feeding
The original research of [73] recommends the OLR as 0.05 . . . 0.10 kg COD kg sludge VSS ×day for the start-up period and increasing of OLR after achieving the removal rate of at least 80%, however, the increment values are not specified. The expression of COD load per VSS of sludge per day is called "sludge load", but in studies OLR is usually reported as  [10] in general, but minimal values are not reported and no details were found for the start-up period.

Infrastructure of UASB Reactor
Based on our experience and referenced here studies, we want to suggest a unified operational process flow diagram as in Figure 2, where we would like to emphasize several aspects.

pH Adjusting and Alkalinity
As mentioned above, the composition of pH adjusting solution is a point of choice [115], but, regardless, the solution should be pumped directly into the reactor feeding line (the mixing manifold on schematic of Figure 2. Otherwise there is a potential for growth of competitive microorganisms in the substrate feeding tank, leading to chemical changes of substrate. The concentration of pH adjusting solution should be balanced based on: (1) daily amount of solution needed to pump (pumps may have lower limit of pumping speed) and (2) minimizing substrate dilution. The interim decision could be to use concentrated solution that is pumped and dosed on a timer, if calculated flowrates are below the limits of a pump. High concentrations of pH adjusting solutions can be chemically aggressive. To prevent the contact between the substrate and parts of pumping mechanism, the peristaltic pumps are recommended for use.
Referenced here studies dissolve or add 0.5-3 g of NaHCO 3 per 1 L of substrate and achieve the stable pH in favorable methanogenic range. However, there are more general suggestions to maintain the ratio between alkalinity of substrate (expressed as CaCO 3 ) and its COD as 1.2-1.6 g CaCO 3 g COD [121]. This value can be used as a reference to calculate the dosage portions and intensity for pH adjusting solution, but should be optimized later [122] downwards. Methanogenesis could occur until ratio of 0.8 g CaCO 3 g COD with some extreme cases of 0.3 g CaCO 3 g COD , but lower values should inhibit methanogenesis and stimulate the formation of hydrogen [123,124].
Another reference value for regulation of alkalinity is ratio of Volatile Fatty Acids (VFA) to Total Alkalinity (TA). Industrial guidelines [114] recommends the ratio VFA:TA to be below 0.35 and consider the value below 0.25 as best for anaerobic digesters, and below 0.15 as safe against pH changes in substrate. The VFA should be expressed as equivalent concentration of acetic acid in mg L and TA as equivalent of CaCO 3 in mg L . The determination of alkalinity by titration is described in Standard methods 2320 [76], as well as appropriate methods for VFA with gas chromatography is covered by Standard methods 5560. However, there were attempts to substitute VFA determination by titration [125][126][127], to avoid using of gas chromatograph.

Feeding and Recycling
The feeding of reactor, based on OLR and HRT is the point of optimization targeting the maximum achievable loads, however, the [10] recommends to limit the vertical velocity of liquid depending on the type of sludge and type of waste: After sludge matured and granulated, the flow could be increased by 50%. In the case of insufficient vertical velocity and to prevent clogging, the recycle line can be used to manage it and (a) to dilute substrate with treated wastewater, (b) to reuse of alkalinity [28], or (c) enhance the granulation by increasing the vertical up-flow [128].
Important remark: effluent recycle port must be separate and located below the effluent discharge port. It is made to prevent back-pumping of air from the effluent discharge line. In our laboratory set-up, we used the flow splitters on effluent port to obtain a recycle line, and we noticed some gas bubbles in it.

Manual Injection Port
A manual injection is strongly recommended and is purposed for: • urgent (emergency) injection of solution for managing pH, coagulation/flocculation, or granulation agent problems; • testing an enhancement of inoculant via injection of specific microbial culture(s); and • sampling of substrate which is supplied to a reactor after all mixing procedures.

Biogas Collection and Counting
Notice the installed one-way valves in the gas line in Figure 2. Check valves are important to prevent the back flow of gas and there are several reasons for that particular phenomenon: • drop of the ambient temperature and, consequently, gas compression in gas lines according to the combined gas law; • at the beginning of UASB operation, when substrate gradually fills the reactor and gas tubes have residual air. The oxygen from residual air is consumed and thus the volume shrinks.
Any of those reasons can lead to one of the two undesirable consequences: • ingress of liquid from reactor to a gas line, which potentially grabs the foam and clogs the pipeline. • if water displacement gas counter is used: backflow of liquid from counter back to a reactor.
Important clarification is to use check valves with low cracking pressure. 'Cracking pressure' is a pressure value when check valve starts opening (passing gas through itself) and this pressure (converted into inches of water column) must be taken into consideration when designing the gas separator, specifically, the height and the level difference between gas collecting part and the effluent release port. Usage of valve with high cracking pressure result in need to build tall GLSS, increasing the material needed to build reactor and its dead volume.
We also want to stress that the gas counter working on the water displacement principle is the only option for raw biogas. There are gas counters working on heat transfer principle ( similar to thermal conductivity detectors of gas chromatographs), which seem to be cheaper options, but they should not be used. Those counters can be calibrated for gas flow with constant content only, which is not a case for biogas. However, they can be theoretically applied if biogas was stripped with alkaline solution to remove acidic gasses (carbon dioxide, hydrogen sulfide, etc.) and assumed to be upgraded to bio-methane. We do not recommend the use of that.

Tracking Operational Parameters
The exact set of trackable parameters depend on the purpose of a particular study, but for general cases, we listed those parameters in Table 6.
In addition, the track of biogas composition during the UASB experiments, the total gas yield and methane yield should be logged. Mentioned above parameters for logging and calculations on their basis do provide a basic understanding of ongoing process inside of UASB reactors, while interpretation of calculations result are not the scope of this manuscript to avoid swelling of it. However authors feel also a need to mention, that if some deeper understanding of chemical process or COD balancing is needed, other researchers [59,[131][132][133] suggest calculation of what part of metabolism is presented by certain process according to the equations, collected in Table 8: Table 8. Equations for metabolic ratios estimation. COD mass balance COD in f luent = COD accumulated + COD biogas + COD e f f luent (10) Other parameters, not included here, belong to some partial cases of UASB experiments and are subjects of individual consideration. Examples for a category of such specialized studies could be effects of salinity or metal ions on the process inside of UASB, which would require extra electrical conductivity, ion-selective electrodes, or other quantitative measurements for both influent and effluent [134,135]. If the study is dedicated to toxicity or biodegradation of particular compound, the appropriate assay tests for that compound or its metabolites should be added [136,137], etc.

Conclusions
With this article we would like to draw the researcher's focus towards the need to report in their publications more information on materials and methods, including specifically sketches/operational flowcharts, seeding conditions, inoculum sources and pre-treatments, and all adjustments to the substrate and feeding equipment. The consideration and addition of these details will help to facilitate a strong scientific and engineering community with comparable research results and conditions. Such detailed data and methods reporting will also significantly propel modeling studies that aim to realistically predict bioreactor behavior in various process conditions. Author Contributions: Y.P. conceived the study, collected and analyzed information, wrote the manuscript under supervision and support from R.C.S. and C.D.M. All authors have read and agreed to the published version of the manuscript.

Funding:
The authors acknowledge the financial sponsorship of WesTech-Inc., Salt Lake City (A-43875) and the Huntsman Environmental Research Center (A-17526).

Conflicts of Interest:
The authors declare no conflict of interest.