Biodegradation of trace contaminants – pharmaceuticals and personal care

products

Dr. Abbie PorterMay 3, 2010

Outline

• Discuss abundance of these compounds in the environment

• Overview of bacterial degradation• Specific pathways and how they relate to

model aromatic degradation pathways• Predicting biodegradability

What are PPCPs?

• PPCPs are Pharmaceutical and Personal Care Products.

• Personal care products include products used for personal hygiene

• Classified by use, not structure• Antimicrobials, fragrances,

surfactants

Entry into the environment

• PCPs enter wastewater treatment plants (WWTP) as components of gray water.

• Pharmaceuticals are also components of the waste stream, but may have been modified via conjugation after being ingested.

• While there is removal of these compounds during the wastewater treatment process, it may not be complete, which results in environmental release with the treated water

• Constant use = continued release

PPCP environmental abundance

• Top 5 contaminants included:– DEET (N, N-

diethyltoluamide)– Caffeine– Triclosan– 4-nonylphenol

• Concentrations ranged parts per trillion to parts per billion

Kolpin et al. 2002

Factors related to PPCP environmental influx

• Removal in the WWTP– Abiotic – photolysis/photodegradation– Biotic – microbial (bacterial and fungal) degradation– Sorption to biosolids

Factors related to PPCP environmental influx

WWTP parameters that might have an effect on biodegradation

1. Temperature2. Hydraulic retention time 3. Solids retention time

– Some microbes have a slower growth rate– May require a period of adaptation before degradation– Gene induction– More easily utilized substrates must be removed first

Environmental parameters that might have an effect on biodegradation

1. Temperature2. O2 availability

3. Availability of alternate electron acceptors4. Acclimation of indigenous microbial population

Factors related to PPCP environmental presence

Toxicology concerns: endocrine disruption

• Definition: interference with endocrine system. – Mimic hormones – Block hormones– Cause hormone production at

inappropriate times– Stimulate overproduction of

hormones

Focus today: estrogen mimicking compounds

Estrogenic exposure:

*not always visually obvious, usually expression of certain female biomarkers (proteins associated with egg production)

Environmental estrogens

Effects resulting from estrogen exposure

• Feminization of male trout – intersexual individuals

• Changes in sex ratio to female dominant

• Reduced hatching rates (fish)

• Weak binding to estrogen receptor

• Environmentally relevant concentrations

• Synergistic effects - additive

Estradiol Nonylphenol

Fate of antimicrobials• Compounds: triclosan and triclocarban• Toxicity: possible endocrine disruption activity• Persistence: One study found triclosan in sediment cores dating

back >30 years (Singer et al. 2002)• Triclosan can be degraded aerobically (Hay et al., 2001), but not as

readily anaerobically• Triclocarban can be degraded anaerobically (Miller et al., 2008) but

not as readily aerobically.

OC l

C l H O

C l

TriclosanTriclocarban

Fate of synthetic musks• Trade names Galaxolide (HHCB) and Tonalide (AHTN), HHCB is most

commonly used• Use: fragrance compounds• Toxicity: have shown both estrogenic and estrogen-blocking effects

O

C H3

H3C

H3CC H3

H3CC H3

HHCB

33

C H3H3C

C H3

C HH C

H3C

O

C H3

AHTN

Estrogens

Anoxic

Estrogens

• 17α-ethynylestradiol (EE2) is a component of birth control pills• Some report EE2 as more recalcitrant than E1, E2, or E3, but there

are isolates able to metablize it (strain JCR5)• May be co-metabolized with E1, E2, or E3.

Model of aromatic degradation

Aromatic catabolism

• Common features: mono- or dioxygenation to activate the ring

• Formation of catechol or substituted catechols• Ring cleavage: either ortho or meta

Annu. Rev. Microbiol. 1996. 50:553-590

DEET• Chemical name: N,N-diethyl-m-toluamide• Use: insect repellent• Strain: Pseudomonas putida DTB

DEET 3-methylbenzoate 3-methylcatechol 2-hydroxy-6-oxo-hepta- 2,4-dienoate

diethylamine

Ibuprofen• Ibuprofen is the 3rd most widely used pharmaceutical in the world.• Chemical name: 2-(4-isobutylphenyl)-propionic acid• Use: analgesic, anti-inflammatory frequently found in the

environment, but readily degraded• Strain: Sphingomonas sp. Ibu-2

Ibuprofen Ibuprofen-CoA Isobutylcatechol

Alkylphenol polyethoxylates (APE)

• Nonionic surfactants• Mostly used in agricultural and industrial

processes, but about 15% of the total production goes to household use (cleaners, PCPs)

• Have been banned in the EU

APE degradation - aerobic

OO

OOH

nO

OOH

Di-ethoxylate

OOH

Mono-ethoxylate

OR

Polyethoxylate

OO

OOH

n-1

OO

OOH

n-2

APE degradation - anaerobic

XO

OO

OH

n

OHOO

OOH

n-1

OO

OOH

n-2

OOH

Continued input of APE parent compounds and lack of alkylphenol removal leads to accumulation under anaerobic conditions

Alkylphenols

• Octylphenol (1 isomer) and nonylphenol (>22 isomers)

• Use: metabolites of alkylphenol polyethoxylates• Toxicity: mimic estrogen• Strains: Sphingomonas sp. TTNP3, Sphingobium

xenophagum Bayram, and Sphingomonas sp. PWE1

Hypothesized pathway

LapKLMNOP

OH OH

OH

OH

COO-

CHO

LapB

a. b. c.

LapKLMNOP

OH OH

OH

OH

COO-

CHO

LapB

a. b. c.

O H O H

O H

C O O H C O O H

C H O

C O O H

O H

HOC

C O O H

O

C O O H

C O O H

O

+

+

Ortho Cleavage

Meta Cleavage

Degradation is isomer dependent

• NP isomers with low amounts of branching were co-metabolically transformed

O

OH

OH

OH

O

OH

OH

O

OH

O

OH

OH

a.

f.

e.

d.

c.

b.

g.

O

OH

OH

OH

O

OH

OH

O

OH

O

OH

OH

a.

f.

e.

d.

c.

b.

g.

Degradation via ipso substitution

OP Hydroquinone 1,2,4-benzenetriol

Examples of ipso substitution substrates

OH OH OH

a. b. c.

OH OH OH

a. b. c.

OH OH

d. e.

OH OH

d. e.

OP NP

Bisphenol-A

Kolvenbach et al. 2007

• Chemical name: Bisphenol A• Use: plasticizer• Toxicity: estrogen mimicking compound• Strain: Sphingomonas sp. TTNP3• Mechanism: ipso substitution

BPA Hydroquinone

4-(2-hydroxypropan-2-yl)phenol

Predicting biodegradability• While PPCPs look different at first, there are structural elements that are

frequently found in common, such as the aromatic ring.• Based on the literature, it’s possible to make rational hypotheses as to

how the chemicals could be metabolized without having done any experiments.

• There are programs that have compiled all of the known metabolic mechanisms in the literature and use that information to predict reasonable mechanisms for compounds that have not been published yet.

• This can be very useful. – Keep in mind, experimental data may be more useful that something from an untested

model.– Bacteria continue to surprise us. The obvious pathway may not always be in use (OP

pathway).

Predicting biodegradability

• Database of published pathways• Also a feature to examine the probability that a compound might be degraded through a specific pathway.

Predicting DEET Biodegradation

http://umbbd.msi.umn.edu/predict/index.html

Predicting DEET Biodegradation

Predicting DEET Biodegradation

Predicting OP biodegradationhttp://umbbd.msi.umn.edu/predict/

Predicting OP biodegradation

• This pathway is similar to what had been predicted earlier for OP biodegradation.

• However, this does not appear to be the case for OP biodegradation in the specific Sphingomonas strains studied.

Notes of caution

• The biodegradation prediction function is based on rules generated from pathways that are in the literature.

• There may be multiple pathways for degradation• Not all pathways have been identified and are not in the

database.• While this is useful to provide a starting point for examining

biodegradation, this does not outweigh experimental observations– Example: OP biodegradation pathway

Reasons to study PPCP biodegradation

• Environmental persistence• Possible toxic or endocrine disrupting effects• Widespread use and continual entry into the

environment• Unknown metabolites – need a way to track the fate

of these compounds in the environment