# US 10,309,951 B2 · Bioreactor for the in situ study of microbial biofilms inducing corrosion on metal surfaces

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- Number: US 10,309,951 B2
- Document ID: US10309951B2
- Status: Granted
- Kind: B2
- Application number: 15/241,640
- Filed: 2016-08-19
- Published: 2017-02-23
- Granted: 2019-06-04
- Earliest claimed: 2015-08-20
- Assignees: Instituto Mexicano Del Petroleo
- Applicants: Instituto Mexicano Del Petroleo
- Inventors: Norma Icoquih Zapata Penasco; Vicente Garibay Febles; Florentino Leyte Guerrero; Ubaldo Sadott Pacheco Y Alcala; Jorge Alberto Mendoza Perez; Jesus Emmanuel Juarez Rojo
- Web page: https://patentgrep.com/patent/US10309951B2

## CPC

- G01N 33/20: Metals
- C12M 29/10: {Perfusion}
- C12Q 1/04: Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor {(C12Q1/6897 takes precedence)}
- C12Q 1/045: {Culture media therefor}
- G01N 17/006: {of metals}
- C12M 25/02: {Membranes; Filters (filters or filtration in general B01D24/00-B01D41/00)}
- C12Q 1/06: Quantitative determination
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)
- C12M: APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; {APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS}
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES

## Abstract

A portable, single-phase bioreactor provides the in situ study of mesophilic and thermophilic corrosion inducing microbial biofilms on metal surfaces by oil, chemical, petrochemical, oil refining, food, metallurgical, paper fluids. The bioreactor is a batch-type for turbulent and piston-driven laminar flow, operated by cycles, with continuous circulation of the fluid. A culture medium or industrial or production fluid containing microbiota is introduced into a homogenizing container. The bioreactor includes a support section having removable corrosimetric test coupons that contact the fluid under dynamic conditions corresponding to fluid-carrying pipelines, and promote the formation of the microbial biofilm for analysis of the kinetics of microbial biofilm development. The bioreactor produces turbulence to homogenize the fluid and maintain a temperature of 20 to 80° C. for the microorganism growth. The fluid has a salinity of 2 to 200 ppm (NaCl) and a pH of 2 to 10.

## Claims

### Claim 1

A bioreactor for the in situ study of corrosion inducing mesophilic and thermophilic microbial biofilms on metal surfaces and measuring a rate of corrosion of a surface of a metal test coupon, said bioreactor comprising:
a homogenizing container (1) containing a culture medium, industrial fluid or production fluid, said homogenizing container having an inlet arid an outlet;
a hermetic container having a thermostatic temperature control and pump, an inlet and an outlet;
a first flexible conduit (3) connected to said outlet of said homogenizing container for transporting said fluid from said homogenizing container (1) to said inlet of said hermetic container (4);
a support container for supporting a plurality of test coupons, said support container having substantially tubular shaped wail with an inner perimeter surface forming a longitudinal passage, a fluid inlet at an inlet end of said longitudinal passage, and fluid outlet at an outlet end of said longitudinal passage,
a second flexible conduit (5) connected to said outlet of said hermetic container and to said fluid inlet of said support container to carry fluid pumped from the hermetic container (4) to the support section (6),
a plurality of hermetic holders extending through said wall and removably coupled to said wall of said support container, each hermetic holder supporting a corrosimetric metal test coupons (8) in said longitudinal passage of said support section, said hermetic holders and corrosimetric metal test coupons being longitudinally spaced apart along a longitudinal axis of said longitudinal passage, and angularly spaced apart around a circumference of said inner perimeter surface of said longitudinal passage of said support container; and
a settling container (11) having an inlet with a valve connected to said outlet of said support section for receiving fluid and biomass waste residuals from the support section (6), said settling vessel having an outlet connected to said inlet of said homogenizing container forming a closed loop, said settling vessel having a drain for removing sediment form said settling container.

### Claim 2 (depends on 1)

The bioreactor of claim 1, wherein the bioreactor is a batch portable device providing turbulent and piston-driven laminar flow through said support container, continuous flow in operation cycles, and continuous fluid circulation at temperatures of 20 to 80° C., salinity from 2 to 200 ppm (NaCl) and a pH in the range 2 to 10.

### Claim 3 (depends on 1)

A bioreactor according to claim 1, wherein said culture medium or industrial or production fluid, includes a culture medium and a reference bacterium (monospecific biofilm), a consortium of reference bacteria (multi-specific biofilm), isolated mixed cultures of industrial or production fluids, or an industrial or production fluid from an assessed industry containing microbiota.

### Claim 4 (depends on 1)

A bioreactor according to claim 1, wherein said inlet (2) of the homogenization container (1) is configured for receiving said culture medium or operational or production fluid from an industry to be assessed.

### Claim 5 (depends on 1)

A bioreactor according to claim 1, said homogenization container (1) includes submersible electrodes for the measurement of temperature, salinity, oxygen concentration, pH and conductivity.

### Claim 6 (depends on 1)

A bioreactor according to claim 1, said inlet of said support container (6) is configured for injecting biocides, corrosion inhibitors or scale removers to said support container, and removing a fluid sample from said support section.

### Claim 7 (depends on 1)

A bioreactor according to claim 1, wherein said corrosimetric test coupons (8) are oriented in different spatial orientations in said corrosimetric test coupons support container (6).

### Claim 8 (depends on 1)

A bioreactor according to claim 1, wherein said support container includes seven hermetic holders for a respective corrosimetric test coupons (8), each hermetic holder supporting said respective corrosimetric test coupons at a different spatial orientation and angle in said longitudinal passage of said support container (6).

### Claim 9 (depends on 1)

A bioreactor according to claim 1, wherein said valve of said support container (6) is configured to block the flow of fluid from said support container and operate independently said settling containers (11) and homogenization container (1) when the reactor is off.

### Claim 10 (depends on 1)

A bioreactor according to claim 1, wherein said corrosimetric test coupons support container (6) are independently removable from said support container.

### Claim 11 (depends on 1)

A bioreactor according to claim 1, wherein said valve of the settling container (11) is configured to drain said settling container.

### Claim 12 (depends on 1)

A bioreactor according to claim 1, wherein said homogenization container, hermetic container, support section, and settling container are independently removable from each other for cleaning and sterilization.

### Claim 13 (depends on 1)

A bioreactor according to claim 1, wherein said fluid is oil, chemical fluids, petrochemical fluids, oil refining fluids, food fluids, metallurgical fluids, and paper fluids.

### Claim 14 (depends on 1)

The bioreactor of claim 1, wherein said metal test coupons have a first end coupled to the respective holder and a second end positioned in said longitudinal passage at the inner perimeter surface of said support container, and where said metal test coupons are oriented on said inner perimeter surface to detect a rate of corrosion at the location of said metal test coupons.

### Claim 15 (depends on 1)

The bioreactor of claim 1, wherein said first flexible conduit has a serpentine configuration to control the flow of said fluid to said hermetic container.

### Claim 16 (depends on 1)

A method of measuring corrosion on a metal surface by a biofilm using the bioreactor of claim 1, said method comprising
introducing said fluid containing said culture medium into said homogenization container where said fluid has a temperature of 20 C to 80 C, a salinity of 2 to 200 ppm (NaCl), and a pH of 2 to 10;
carrying said fluid from said homogenizing container through said first flexible conduit to said hermetic container;
carrying said fluid form said hermetic container through said second flexible conduit to said support container;
directing said fluid through said longitudinal passage of said support container into contact with each of said test coupons;
carrying said fluid from said support container to said settling container and continuously carrying said fluid from said settling container to said inlet of said homogenizing container; and
removing said test coupons and analysing formation of biofilm and corrosion of said test coupons.

### Claim 17 (depends on 16)

The method of claim 16, wherein said fluid is a laminar flow through said longitudinal passage of said support section.

### Claim 18 (depends on 15)

The method of claim 15, wherein said fluid is a turbulent flow through said longitudinal passage of said support section.