the efficacy of biocatalysts as wax pour point depressants

International Journal of Scientific Research and Engineering Development-– Volume 4 Issue 6, Nov-Dec 2021

Available at www.ijsred.com

ISSN : 2581-7175 ©IJSRED: All Rights are Reserved Page 551

The Efficacy of Biocatalysts as Wax Pour Point Depressants

Ubong U. Isang 1 M.O. Onyekonwu

2 Joseph Ajienka

3 Gideon C. Okpokwasili

4Bella Mmata

5

1(World Bank Africa Centre of Excellence in Oilfield Chemicals Research, University of Port Harcourt,

Nigeria. Email:[email protected]) 2-3

(Department of Petroleum and Gas Engineering, University of Port Harcourt, Nigeria) 4(Department of Microbiology, University of Port Harcourt, Nigeria)

5(Laser Engineering and Resources Consultant, Port Harcourt, Nigeria)

----------------------------------------************************----------------------------------Abstract:

Wax deposition is one of the problems of flow assurance. As a result, operational and remedial costs rise while oil output declines.

One of the mitigation measures is the use of pour point depressants. The study was on the isolation, screening, identification and

characterization of lipase-producing fungi and bacteria for use as depressants. Design, modification, expression, purification, creation and

validation of detailed enzymatic or microbial systems that can catalyze the needed chemical reactions are all part of biocatalyst development.

Saccharomyces cerevisiae, Bacillus cereus and Pseudomonas aeruginosa secreted biocatalysts, were utilised in the study. Inoculations on the

spot for primary screening and Lipase Synthesis in Submerged Fermentation were used during the screening process. The reaction was carried

out at temperatures ranging from 10 to 90 °C to see if temperature had an impact on lipase activity. Most of the lipase was produced by

Pseudomonas aeruginosa, followed by Bacillus cereus, with a small amount of lipase in the shape of a zone around the Saccharomyces

cerevisiae colony. This unique wax mitigation technology combines depressive qualities with a significant reduction in pour point and

improved flow properties.

Keywords: Bacillus cereus, Flow assurance, Lipase, Pseudomonas aeruginosa, Pour point, Saccharomyces cerevisiae.

----------------------------------------************************----------------------------------

I. INTRODUCTION

Wax, commonly known as paraffin, is an organic compound

consisting of numerous forms and combinations of aliphatic

hydrocarbons. The extraction of crude oil from the deposit through

the wellbore to the processing facilities involves temperature and

pressure drops. The main cause of wax problem in the tubing and

pipelines is drop in temperature, which is determined by the crude

oil’s pour point and cloud point also known as wax appearance

temperature. According to research, the pressure reduction enhances

the likelihood of paraffin production due to the related temperature

drop [1].Wax problems have been discovered as a steady-state

problem in which paraffin is progressively deposited along pipe

walls when the wall temperature drops below the Wax Appearance

Temperature WAT for a lengthy period of time. If the WAT is

reached, it can also occur in the reservoir. The network of crystals

continues to develop the strength of an interlocking structure when

crude oil is continuously cooled below its pour point and maintained

at a temperature below the pour point under quiescent conditions [2].

Because the reservoirs are usually isothermal, the temperature profile

of a well does not alter over time.

The carbon number or the percentage of paraffin (C18-C40) in a

crude sample can be used to identify paraffinic or waxy oil [3].

Extensive crude oil sampling and analysis are required at the

appraisal stage of field development in order to formulate a firm field

development plan. This analysis aids in identifying the crude's (WAT)

and Pour Point [4].

The primary concerns related to the wax flow assurance

challenges are the physicochemical qualities and features of a waxy

crude and deposited wax. Wax concerns can have a substantial

impact on well and lease profitability, generating operational issues

and lowering production. When wells and pipelines are shut down,

either intentionally or unintentionally, and the ambient temperature is

below the pour point, such situations can arise. Waxy crude oils

contain high molecular weight components, which may precipitate as

a wax phase at low temperatures. This can lead to pipe clogging and

a variety of other issues. When the temperature of a waxy crude

drops, the heavier fractions are the first to precipitate.

As soon as the cloud point WAT is achieved, crystallization

begins. The waxy crude may gel and solidify depending on the shut-

down time and ambient temperature, among other factors. The waxy

crude oil passes through a phase transition from a liquid state with

Newtonian flow to a gel-like structure with non-Newtonian flow

behavior. Precipitation, deposition and gelation are the three primary

steps in the phase transformation process.

The thermodynamic WAT must be distinguished from the

empirically measured WAT. The solid phase boundary temperature

is defined by the thermodynamic WAT, which is the maximum

RESEARCH ARTICLE OPEN ACCESS




temperature at which the solid and liquid phases are in equilibrium at

a constant pressure. The experimental WAT is the temperature at

which the first crystals are discovered and it is therefore dependent

on the sensitivity of the measurement device and technique.

Researchers have been tackling with how to measure and quantify

the flow behavior of waxy crude oils. The generated waxy-oil gel

structure exhibits a very complicated rheology involving both time-

independent and time-dependent features after cooling under

quiescent conditions. Yield stress, pseudo-plasticity, time-

dependency, fluid compressibility, shrinkage, heat history, shear

history, composition and aging are all factors that affect the flow

behavior of waxy crude oils.

This issue has resulted in a great deal of uncertainty in oilfield

development, resulting in costly through-tubing (TT) workover runs,

unnecessary downtime, income loss and premature well

abandonment in the Niger Delta. Deposition, according to [5], begins

on any surface (called a cold spot) with a temperature lower than the

temperature at which wax begins to emerge. The following are some

of the factors that influence this condition [5]: the waxy crude's

makeup (at least 2% wax content) composition of fluids, the

interaction of crude components may prevent deposition and modify

cloud point. Temperatures drop below the cloud point range as the

time progresses. According to [6], paraffin deposition is affected by

viscosity, flow rate, well maintenance operations such as water

injection, cooling equipment, gas lift and acid/fracture jobs using

cold fluids might change the temperature.

Wax formation and deposition increases pipe roughness and

decreases the effective cross-sectional area of the pipe, resulting in

increased pressure drop, stress, and wear. The deposits also induce

hole, plugging, and malfunction of subsurface/surface equipment.

Wax formation necessitates more frequent and dangerous pigging, as

well as more frequent shutdowns and operating issues. It raises the

fluid's viscosity to the point where the oil forms a gel, making oil

pumping difficult. It can raise energy and chemical consumption

while lowering production throughput and selling value of the

produced fluids when used in excess. Above all, these issues have

substantial financial consequences, which can lead to the removal or

abandonment of an entire oil field production network system.

A. Methods of Handling Wax

To increase the flow properties of waxy crude oil, many strategies

to address wax deposition, formation and precipitation are tentatively

proposed and utilized in practice. The prevention measures, on the

other hand, are far more cost-effective than the remediation ones.

The most common types of preventative strategies are thermal,

mechanical and chemical [7].

[8] shows that their overview of wax-deposition, formation, and

mitigation strategies in the oil and gas industry gives professionals

and the flow-assurance community a general understanding of

diverse wax-deposition-prevention and inhibitory technologies. To

attain the best results in enhancing the flow property of these waxy

crude oils, a combination of chemicals or procedures is usually used.

The diluting with light-ends approach and the heating method are

two of the most extensively used traditional thermal procedures for

improving fluidity.

The mechanical pigging method is well-known for removing

paraffin wax, but it necessitates frequent production shutdowns,

resulting in significant economic losses. As a result, the frequency of

pigging must be considerably reduced in order to generate larger

economic benefits. Chemical techniques like wax inhibitors (WIs)

and pour point depressants (PPDs) are therefore ideal [9].

[10] proposed a method for predicting wax deposition in

hydrocarbon mixtures based on a thermodynamic model that was used to establish the onset condition (s) for wax precipitation.

The goal of the work by [11] was to investigate the process of wax

deposition on the internal surface of oil production pipelines, as well

as the effect of various parameters such as flow rate and pipe wall

temperature on deposit thickness for a light crude oil with high

paraffinic content. The models were then utilized to simulate the

formation of paraffin deposits as a function of flow rate and pipe

temperature, based on the experimental data given by [12]. When

evaluating a constant wall temperature or an axial thermal gradient

with a positive slope, the results showed that greater flow rates

reduce the maximum deposit thickness because it spreads across a

longer distance in the pipe.

[13], on the other hand, demonstrated how to determine and

explain a new and efficient method of minimizing paraffin

depositions using a novel polymer known as Ethylene

TetraFluoroEthylene (ETFE) with the IUPAC designation poly

ethylene cotetrafluoroethylene. Their study analyzes the properties of

three different types of pipes: ETFE internal plastic pipe coating,

rigid PVC plastic pipe coating and steel pipe.

[14] investigated two innovative strategies for preventing wax

formation that were tested on three oil wells in Libya at a surface

temperature of 29°C. During the gas-lift operation, the gas was

injected at a pressure of 83.3 bars and a temperature of 65°C (higher

than the pour point temperature). Wax inhibitors (Trichloroethylene-

xylene (TEX), Ethylene copolymers, and Comb polymers) were

introduced down the casings with the gas in the second approach.

B. Industrial Applications of Biocatalysts

Biocatalysts are natural substances including enzymes derived

from biological sources, dead or alive microorganisms, derivatives or

complete cells. Chemical reactions are accelerated using biocatalysts.

The development of these biocatalysts are basically identical and are

influenced by a variety of factors; including crude oil composition,

biocatalyst structure, alkyl side chain length, average molecular

weight and the carbon number of an alkane chain in the crude oil. To

achieve optimal co-crystallization and surface adsorption for these

wax crystals, the parameters are highly dependent on one another.

Biocatalysts have recently been advocated in the petroleum

industry for a variety of uses. The current biotechnology industry has




discovered that some enzymes can withstand the harsh conditions

found in oil reservoirs.

Biocatalyst could be used to improve oil output recovery, reduce

formation damage and remove sulphur from hydrocarbons. Pre-

treatment of biopolymers with enzymes could advance the handling

of characterizing them. Water shut-off and sand consolidations are

two further applications of enzymes in the oil and gas industry. [15,

16]

Petroleum spills and incomplete combustion of fossil fuels,

according to [17], have resulted in a build up of petroleum

hydrocarbons in the environment. Petroleum and its byproducts have

accumulated to the point where they have become a major

environmental issue. Biocatalysis opens up new avenues for the

advancement of bioremediation methods.

[18] reported on the production of biodiesel from waste frying oil,

methanol, and Rhizopus oryzae PTCC 5174 in both immobilized and

free forms in the current study.

[19] investigated the interaction of non-ionic and anionic

surfactants with enzymes. The study relied on the lipase enzyme,

which is commonly used in industry to analyze the interaction of

non-ionic and anionic surfactants.

To meet the wax remediation criteria, wax inhibitors and pour

point depressants are developed and modified. Wax crystals are

created through nucleation, growth, and agglomeration processes,

with molecular diffusion and shear dispersion aiding in the

deposition of these waxes.

New breakthroughs, such as polymer/inorganic nano-composites

or nano-hybrids for industrial uses, have resulted from advances in

nanotechnology. Because of their unique size, high surface

adsorption effect, and quantum size effect, nanoparticles have the

ability to change polymers.

[20] found that nano-hybrid pour point depressants (PPDs) were

superior to Ethylene Vinyl Acetate copolymers (EVA copolymers)

alone in terms of pour point depression.

[21] showed some acceptable strains were obtained from tainted

samples exposed to hydrocarbon chemicals for a long time in order

to assess the effect of microorganisms on the lowering of the wax

appearance temperature (WAT) of waxy crude oil.

Lipases (EC 3.1.1.3), also known as triacylglycerol ester

hydrolases, are one of the most adaptable biocatalysts, they are

capable of performing a wide range of bioconversion reactions with a

wide range of substrates. Furthermore, lipases have the unique

property of working primarily in organic media at the lipid/water

interface [22]. Food, pharmaceuticals, fine chemicals, oil chemicals,

biodiesel, remediation, oil and gas and industrial detergent industries

are just a few examples where lipases are used [23]. Lipase can

catalyze a variety of substrates, according to [24], microbial enzymes

have some interesting properties that set them apart from animal and

plant lipases, such as specificity, low cost, availability, and ease of

purification, making them a great alternative for potential

commercial exploitation as industrial enzymes [25] and being

broadly classified as bacterial lipases.

[26] showed how to reduce wax precipitation by combining crude

oil with the manufacturer's solvent, which is made up of toluene and

xylene. The purpose was to evaluate biomass-derived solvents for

their ability to slow down the wax deposition process.

[27] used a biosynthetic ingredient from olive oil (Oka Europae)

to lower the pour point and enhance the rheology of a waxy crude

from 43oC to 8oC. The flow behavior (rheology) improved and the

viscosity was reduced by 89.20% adding 1000ppm of PPD at 45oC.

Biosurfactants from bacteria are structurally varied, according to

[28]. They have industrial and environmental applications as well as

the ability to reduce fluid interfacial and surface tension. They

looked at the biochemical properties and uses of various

biosurfactants.

[29] demonstrated that the drag and pour points of waxy crude can

be reduced by using a manufactured biosurfactant derived from the

enzyme.With the addition of the biosurfactant, the pour point was

decreased, and a stable oil-water emulsion was produced. According

to this data, the viscosity was lowered by 70% and the drag was

reduced by 40%.

[30] used polyethylene glycol esters of cashew nut shell liquid to

investigate pour point depression and flow improved performance of

waxy crude oil. Polyethylene glycol was used to esterify natural

cashew nut shell liquid (CNSL) obtained from Anacardium

occidentale waste shells . At 1000 ppm, CNSL derivatives reduced

the pour point of waxy crude oil by 12 °C.

Meanwhile, the adsorption of wax molecules on the surface of the

PPD inhibits the growth of crystals and changes the crystal formation

pattern by creating a micelle core [31]. In petroleum flow assurance

chemistry, bio-based and natural surfactants are also intensively explored as PPDs [32].

Natural surfactants, in contrast to traditional surfactants, have the

following advantages: decreased toxicity, biodegradability, structural

variation, ability to be synthesized from inexpensive renewable

ingredients, and stability across a wide pH range [33]. When used as

PPD on heavy crude oil, [34] discovered that Sapindus mukorossi, a

natural surfactant, dramatically reduced viscosity by 80%.

According to the researchers, another form of bio-surfactant

derived from Halomonas xianhensis bacteria was found to reduce the

PP by up to 24°C [35].

More specifically, the production of polyamine amide from canola

oil, known as PPD, has been demonstrated to be effective in reducing

PP in crude oil. According to [36] at a concentration of only 0.05–

0.10% the temperature of the PP was reduced to 10°C. Flow

improvers are commonly used to boost crude oil mobility [37]. The



©IJSRED: All Rights are Reserved Page 554

effect of the ethylene-vinyl acetate copolymer (EVA) as a flow

improver was investigated in their paper, with different molecular

weight ranges affecting the viscosity and pour point of five Iranian

waxy crude oils.

II. MATERIALS AND METHODS

TABLE 1: LIST OF EQUIPMENT AND MATERIALS

S/NO Equipment/Material Where Applicable

1 Cultures:

Bacillus cereus,Pseudomonas

aeruginosa,Saccharomyces

cerevisiae

Lipase production

2 Potato dextrose agar PDA,

Nutrient agar NA, Nutrient Broth,

Sterilized plate for isolation of

micro-organism

Lipase production

3 NaH2PO4 =12, KH2PO4= 2,

MgSO4·=0.3, CaCl2 =0.25,

(NH4)2SO4= 1 %.

MGM

Lipase production

4

Glucose, Olive oil, Filter paper,

Ethanol(absolute), Cotton wool,

Nutrient broth, Foil paper

Electronic Centrifuge, Autoclave

pot

Lipase production

5 Gas, Generator and Fuel,

Spectrophotometer

Lipase production

6 R1 Buffer, absorbance at 580 nm

Testing kit,R2 substrate,

Tributyrin, Calibrator

Lipase assay

7 Waxy crude sample Water bath,

Pour point measurement apparatus

Pour point

8 Gas Chromatography Fluid composition

9 Brookfield digital viscometer Viscosity

A. The Research Methodology

The biocatalyst development process includes the design,

modification, expression, purification, construction and validation of

precise enzymatic or microbial systems capable of catalyzing the

desired chemical reactions.

A stage-by-stage approach is proposed to achieve the study's

objectives, as depicted in Figure 1.

This entails putting the pour point depressant to the test to see

how causative elements (temperature, concentration and carbon atom

count) affect biochemical performance. Temperature, cooling rate,

concentration, age,and time-dependence effects on the efficiency of

several biochemicals were also investigated. To examine ideal

performance, gradually increase the amount of biochemicals used.

Sample Collection

The soil samples were obtained from the oil-contaminated soils of

Rupkokwu, Aluu, and the environs of the University of Port Harcourt

using normal procedures, and a composite sample was created for

isolation.

Isolation and screening of lipase-producing bacteria and

fungi

In a 250 mL conical flask containing 100 mL of sterile

physiological saline, one gram samples were suspended. Then, for 30

minutes at 120 rpm, frequent and rapid stirring was used to break up

dirt clumps before letting it to settle. The supernatant was decanted

and used to make a 10-fold serial dilution. Serial dilutions were

made, and 0.1 ml of each 10-2 and 10-3 dilution was cultured on

Potato Dextrose Agar (PDA) for fungi and Nutrient Agar for bacteria

and incubated at 30oC for 1 day.

Fig.1: Workflow Biocatalyst efficacy as wax pour point depressants



©IJSRED: All Rights are Reserved 55

On Agar plates, lipase-producing bacteria and fungi were first

screened using tributyrin as a substrate.

Secondary screening was performed in submerged fermentation,

using 500-mL conical flasks containing 180 mL of medium infected

with 20 mL of inoculum and incubated at 30 °C, Centrifuge at 120

rpm for 24 hours, 48 hours, 72 hours, 96 hours and 37 °C,

respectively, for fungi.

Lipase Assay and Characterization

To begin the reaction, 0.4ml of R1 (Buffer) was mixed with 0.1ml

of the sample (lipase), which was then incubated for 3 minutes at pH

of 7 and 37 °C. After that, 2.5ml of R2 (Substrate) was added and

incubated for 1 minute at 37 °C with a pH of 7.

The Effect of Temperature

The reaction was carried out at various temperatures ranging from

10 to 90 °C to assess the effect of temperature on lipase activity.

Before beginning the experiment, the crude enzyme and substrate

were incubated at various reaction temperatures, and the enzyme test

was done as reported previously to find the best incubation

temperature.

Pour Point Measurement using Biocatalysts

With the help of the water bath shown in Figure 2. the waxy oil

sample was poured into a medium-sized test tube and (the test tube

with its content) thermometer was placed in the test tube with the

help of a stopper to position and hold it. Then it was heated to about

37oC, where the enzyme activities are optimal and was placed in a

pour point apparatus. Temperature readings were obtained every two

degrees down until the temperature reached a point where it could

not be poured. The pour point of the oil sample was noticed and

recorded at this temperature. Twenty-two samples of various mixing

ratios of biocatalyst and waxy crude which totalled 50 ml where

prepared.

Three samples containing waxy crude, Bacillus ceresus,

Pseudomonas aeruginosa were selected and extracted in this section

using the solvent extractor outlined in Figure 3.

Waxy crudes without Biocatalyst were recovered by filtration and

then analyzed using gas chromatographs (see Figure 4) and viscosity

studies were also done on the samples.

III. RESULTS AND DISCUSSION

Bacillus cereus and Pseudomonas aeruginosa were grown on

Nutrient Agar (NA), while Saccharomyces cerevisiae and

Aspergillus niger were cultured on Potato Dextrose Agar (PDA).

The clean zone around the colony was seen using Tributyrin Agar

Medium for the primary screening, as reported in Figure 5 to 7.

Fig. 2: HH-S Water Bath

Fig. 3: Extraction of impurity from the waxy crude

Fig. 4: Agilent 7890A Gas Chromatograph



©IJSRED: All Rights are Reserved Page 556

The mycelium was extracted by filtration under vacuum and then

centrifuged at 4000 rpm for 20 minutes during the screening.

Extracellular enzymes were obtained from the cleared supernatant.

Centrifugation at 4000 rpm for 20 minutes separated yeast and

bacterium culture materials.

Temperature is a vital characteristic that must be managed, and it

varies significantly from one organism to the next. From 10°C to

90°C, the optimum temperature for lipase synthesis and bacterial

growth was examined after every 24 hours of culture for 96 hours.

Spectrophotometer readings for the lipase test revealed that 37°C was

the optimum temperature for both lipase production and bacterial

growth (Table 2 to 5). Lambert's rule for determining enzyme

activity was used to convert absorption at 580nm to enzyme units.

Table 2: Spectrophotometer Reading for lipase assay after 24 hours

Enzyme Activity after 24 hours

Tempt(oC) EASC(U/l) EAPA(U/L) EABC (U/L)

10 2 1.8 1.2

20 5 4.5 2.8

30 8.5 8 4.2

35 9.2 8.8 4.46

37 9.31 9.19 4.48

40 9.2 8.7 4.45

45 9 8.2 3.2

50 5 4.8 2.6

60 2 1.9 1.2

70 1.5 1.2 0.6

80 1.1 0.8 0.4

90 0 0 0.2

Table .3: Spectrophotometer Reading for lipase assay after 48 hours

Tempt(oC) EASC(U/l) EAPA(U/L) EAPA(U/L)

10 1.2 0.9 0.2

20 2.2 1.4 0.6

30 2.5 1.9 1.1

35 2.9 2 1.12

37 2.95 2.08 1.14

40 2.92 2.1 1.1

45 2.4 1.7 1

50 2 1.5 0.96

60 1.5 0.9 0.65

70 0.6 0.4 0.6

80 0 0.003 0.4

90 0 0 0.2

The optimum temperature for lipase production from the three lipase

producing microorganism was 37°C.


Fig. 5: Pseudomonas Aeruginosa + NA+ Substrate with clear zone

Fig.6: NA + Substrate+ Bacillus with zone of inhibition after 24 hours

Fig.7: PDA plate+ Saccharomyces cerevisiae+ Substrate after 24hours showing zone of inhibition

Enzyme Activity of Saccharomyces cerevisiae=EASC,

Enzyme Activity of Pseudomonas aeruginosa = EAPA,

Enzyme Activity of Bacillus cereus=EABC





10 4 8 1.2

20 6 10 2.1

30 7.6 14 3.64

35 8.2 16.8 3.7

37 8.4 17.19 3.81

40 8 16.9 3.76

45 6 14 3.5

50 3 8 2.2

60 0.9 2 1.8

70 0.02 1.5 0.9

80 0.1 0.9 0.2

90 0 0.03 0.01



10 0 0 0

20 0.1 0.25 0

30 0.3 0.35 0.3

35 0.5 0.5 0.5

37 0.5 0.5 0.5

40 0.5 0.5 0.4

45 0.4 0.4 0.3

50 0.3 0.003 0.1

60 0.03 0 0

70 0.02 0 0

80 0 0 0

90 0 0 0

Biocatalysts' effect on the pour point of waxy crude oil

The ASTM techniques were used to determine the pour points of

the samples. These studies were critical in determining and

comparing the efficacy of the depressant formulations proposed to

prevent crude oil wax formation. The untreated crude oil solidified at

31.5° C., as shown in Table 6. The viscosity of the liquid is 15.90 c.p.

Meanwhile, crude oil treated with biocatalyst (lipase) from

Pseudomonas aeruginosa, Bacillus cereus, and Saccharomyces

cerevisiae solidified at various temperatures, as indicated in Table 6,

depending on the mixing ratio. This validates the efficacy of these

biocatalyst formulations in lowering the pour point of waxy crude

and improving flow characteristics. This supported the hypothesis

that WAT is related to molar mass and chain structure, with waxes

that are more linear and have greater molar mass crystallizing at a

higher temperature and waxes that are less linear and have lower

molar mass crystallizing at a lower temperature.

The compositions of paraffin waxes have a significant impact on

the efficiency of wax pour point depressants, as shown in Table 6.

We learned from the literature that heavy n-paraffins (n-

C18+paraffins) reduced the effectiveness of a pour point depressant.

Table 6: Pour point measurement

% of Biocatalyst source of biocatalyst Pour Point at instance pour point at 24Hrs PP after 72 Hours

Viscosity MPa.s (c.p)

0 Not available 31.5 31.5 31.5

15.90

5 Bacillus(B) 27.5 25 24

Not available

10 Bacillus(B) 27.5 22 23

Not available

50 Bacillus(B) 22 19 18

8.33

5 Pseudomonas (P) 24.5 21 16

11.33

10 Pseudomonas (P) 29 27.5 28

Not available




50 Pseudomonas (P) 26 26 26

Not available

5 Saccharomyces cerevisiae (S) 27 24 24

Not available


Not available


Not available

5 B+P 26 26 26

Not available

10 B + P 27 26.5 26

Not available

50 B+P 28 28 28

Not available

5 B+P+S 27 27 27

Not available

10 B+P+S 28 27 27

Not available

50 B+P+S 28 28 28

Not available

5 Bacillus(B) 28 27 26

Not available

10 Bacillus(B) 28 27 26

Not available

50 Bacillus(B) 28 27 26

Not available


Not available


Not available


Not available

IV. CONCLUSIONS

The wax issue has significant financial implications, and it can

result in the dismantling or abandonment of an entire oil field

production network system. This causes substantial flow issues that

must be addressed. Concerns about wax can have a significant

influence on well and lease profitability, causing operational

problems and limiting output. To meet the wax remediation criteria,

pour point depressants are designed and improved. Unique and novel

biocatalysts are being developed to replace expensive wax pour point

depressant chemicals that have severe side effects and are harmful to

the environment. The advancement of methods for rational or

evolutionary design of novel or modified enzymes has increased the

demand for quick and accurate approaches for identifying suitable

catalysts. Lipases are versatile biocatalysts with significant biological

and industrial applications. So, from the standpoints of research, the

environment and commerce, study findings on the isolation,

screening, identification, assay and manufacture of the enzyme from

microbial sources are significant. Biocatalyst processing has a

promising future. The best producer of lipase was Pseudomonas

aeruginosa, which was generated by the presence of a low-cost lipid

supply with high enzyme activity (17.19U/L).

.

ACKNOWLEDGMENT

Fig. 10: Chromatograph of Pseudomonas + waxy crude oil




The entire management of the World Bank Africa Centre of

Excellence in Oilfield Chemicals Research, University of Port

Harcourt, Nigeria, is greatly appreciated for providing the facilities

needed for the evaluation, as well as their continuous efforts to

ensure the research is completed effectively.

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