Document ID: EPA-HQ-OPP-2007-0566-0014
Agency: epa
Document Type: Supporting & Related Material
Title: 
Posted Date: 2008-08-06T04:00Z

UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

 		     WASHINGTON, D.C. 20460

 ADVANCE \d61 	OFF		ICE OF PREVENTION,

OFFICE OF PESTICIDES AND TOXIC SUBSTANCES        

MEMORANDUM

DATE:		April 7, 2008

SUBJECT:	Non-Target Organism and Endangered Species Assessment in
Support of the Registration Review of Chitin
(Poly-N-Acetyl-D-glucosamine) and Chitosan (Poly-D-Glucosamine). 
Re-review of Existing Non-target Organism Studies

Registration Review Case No.:	6063

Chemical Class: 					Biochemical

PC Codes: 						128991 (Chitin) & 128930 (Chitosan)

			CAS Numbers:			1398-61-4 (Chitin) & 9012-76-4 (Chitosan) 

Tolerance Exemptions: 			40 CFR 180.1089 (Chitin) & 40 CFR 180.1072
(Chitosan)

MRID Numbers: 				None

FROM:		Russell S. Jones, Ph.D., Senior Biologist   /s/  04/07/2008

Biochemical Pesticides Branch

Biopesticides & Pollution Prevention Division (7511P)

TO:			Chris Pfeifer, Regulatory Action Leader

Biochemical Pesticides Branch

Biopesticides & Pollution Prevention Division (7511P) 

BACKGROUND

The Food Quality Protection Act of 1996 mandated the continuous review
of existing pesticides.  All pesticides distributed or sold in the
United States must generally be registered by EPA, based on scientific
data showing that they will not cause unreasonable risks to human
health, workers, or the environment when used as directed on product
labeling.  The Initial Docket for the Registration Review of Chitin and
Chitosan was established for September 2007 and the Final Workplan is
scheduled for January 2008.  In support of the Registration Review, BPPD
has re-evaluated the existing non-target organism studies that have been
submitted in support of the registration of products containing Chitin,
Chitosan, and Chitosan Hydrolysate (a hydrolyzed form of Chitosan). 
These data are re-reviewed in this document to support the Endangered
Species Risk Assessment for the Registration Review of Chitin and
Chitosan.  No new studies were submitted.  

EXECUTIVE SUMMARY

Chitin and its closely related active ingredients, Chitosan and Chitosan
Hydrolysate, are not expected to cause any adverse effects in any
non-target organisms, including threatened and endangered species.  A No
Effects (NE) determination has been made for Chitin and Chitosan
(including Chitosan Hydrolysate).  Chitin is ubiquitous in nature and is
found in many terrestrial and aquatic species (see Effects on Non-Target
Organisms and Endangered Species below).  Chitosan is a partially
deacylated version of Chitin.  Chitosan Hydrolysate is an acid
hydrolyzed version of Chitosan and is comprised of smaller, more
water-soluble chains of Chitosan.  Chitin and its derivatives are
functionally identical and have a non-toxic mode of action (see
Description of the Active Ingredients below).  There is no direct
activity of the active ingredients against the target pest.  

RECOMMENDATIONS AND CONCLUSIONS



Based on the existing data, adverse effects on non-target organisms are
highly unlikely and there will be NO EFFECTS (NE) on threatened or
endangered species resulting from application of products containing
Chitin and Chitosan (including Chitosan Hydrolysate) when products are
used in accordance with approved labeling.  	

NON-TARGET ORGANISMS AND ENDANGERED SPECIES ASSESSMENT

Description of the Active Ingredients

Chitin and Chitosan (which includes Chitosan Hydrolysate) are both
comprised of a naturally-occurring chain of glucose molecules linked via
B-1,4-linkages, that is structurally related to cellulose.  Both
substances are slightly different forms of the same molecule (see Figure
1 below).  

Chitin (Poly- N-Acetyl-D-Glucosamine) is commonly found in nature and is
usually derived from crustacean shells, particularly from crabs and
shrimp.  Historically, it has been used as a food additive and a
fertilizer.  As a pesticide active ingredient, it acts by stimulating
the growth of certain microorganisms in soil, which release substances
that kill pathenogenic nematodes and their eggs.   The compound is also
reputed to play a role as a plant growth regulator by bolstering plant
defenses against disease (Systemic Acquired Response, or SAR inducer)
(Kramer and Muthukrishnan, 1998).  Chitin does not have any direct
toxicity to pests or non-target organisms.    

Chitosan (Poly-D-Glucosamine) is a synthetic compound prepared through
the partial or complete deacetylation of Chitin.  Thus, if deacetylation
is incomplete, a Chitosan chain will contain molecules of Chitin
randomly distributed along its length.  Chitosan also differs from
Chitin in that it contains an amine group, whereas Chitin contains an
amide group.  It is 

somewhat more water-soluble than Chitin.  Chitosan has biomedical
applications.  as a haemostatic agent that is hypoallergenic and,
similar to Chitin, it is known to possess anti-microbial activity. 
These properties also allow for its use as an active ingredient in
anti-microbial pesticides.  As a pesticidal active ingredient, Chitosan
is best known as a plant growth enhancer that boosts the ability of
plants to defend against fungal infections (also an SAR inducer). 
Similar to Chitin, Chitosan has no direct toxicity to pests or
non-target organisms.  

Chitosan Hydrolysate is prepared via the acid hydrolysis of Chitosan and
is comprised of smaller chains of Chitosan (also an SAR inducer). 
Biologically, there is little functional difference between Chitin,
Chitosan, and Chitosan Hydrolysate. 

Figure 1.  The structures of Chitin and Chitosan.  

                                                                        
              

                                                                        
                                                                        
          Chitin                  

 

The actual chemical structure of Chitosan is subject to substantial
variations based on the degree of deacetylation of Chitin; the structure
above depicts a completely deacetylated chain of Chitosan molecules.

Current USEPA-registered Products

There is one USEPA-registered product containing Chitin as its active
ingredient:  Clandosan  618 [EPA Reg. No. 54137-1 (formerly EPA Reg. No.
58200-9)].  There are two USEPA-registered products containing Chitosan
as their active ingredient:  ELEXA-4 (EPA Reg. No. 81045-2) and
ChitoSante (EPA Reg. No. 81446-1).  There are two products containing
Chitosan Hydrolysate as its active ingredient that are currently in
registration review:  Chitosan 

Hydrolysate (EPA File Symbol No. 73512-A) and Keyplex 350 (EPA File
Symbol No. 73512-L).  

Clandosan 618 is registered for non-food use on ornamentals, lawn, and
turf grasses.  Elexa-4 is registered for use on field crops,
ornamentals, turf, home gardens, and nurseries.  Chitosante is
registered for use non-food use as an antimicrobial pesticide on
textiles and surfaces.  Chitosan Hydrolysate (registration pending) is
intended for manufacturing use only.  Keyplex 350 (registration pending)
in intended for use food use on commercial crops, ornamentals, and turf.
 

The amounts of active ingredient in each product are listed in Table 1
below:

Table 1.  Farnesol and Nerolidol Content in USEPA Registered End-Use
Products

Product 

(EPA Reg. No.)	Percentage Active Ingredients 1

	

Chitin	

Chitosan

	Chitosan Hydrolysate

Clandosan 618 (54137-1)	25	-

	Elexa-4 (81045-2)	-	4

	ChitoSante (81446-1) 2	-	6

	Chitosan Hydrolysate (73512-A)3	-

8.33

Keyplex 350 (73512-L) 4	-	-	0.8

1	All active ingredients are closely-related forms of
poly-D-glucosamine.  

2	An antimicrobial product having no outdoor exposure

3	Manufacturing-use only.

4	Also contains 1.5% Salicylic acid as an active ingredient

Effects on Non-Target Organisms and Endangered Species

Chitin and Chitosan (which includes Chitosan Hydrolysate) are both
comprised of a naturally-occurring chain of glucose molecules linked via
B-1,4-linkages, that is structurally related to cellulose.  Both
substances are slightly different forms of the same molecule (see Figure
1 above). 

Chitin is one of the most common polymers found in nature.  It is a
major component of the exoskeletons of aquatic and terrestrial
arthropods (insects, crustaceans), the radula of molluscs, and the beaks
of cephalpods (squid, octopus), and the cell walls of fungi (Campbell,
1996).  In addition, many species possess chitinases (Bishop et al.,
2000; Kramer and Muthukrishnan, 1998), enzymes that metabolize chitin,
particularly those that consume chitin-containing organisms (MacDonald,
2006; Matsumiya et al. 2006).  Chitinases are also found in humans
(Escott and Adams, 1995; Paolettia et al. 2007).



Table 2.  Summary of Non-Target Organism Data/Information for Chitin
(C), Chitosan (CS), and Chitosan Hydrolysate (CSH) End-Use Products (EP)

Study Type/OPPTS Guideline	

LD50/LC50	

Active Ingredient	

Toxicity Category	MRID

Acute Oral Toxicity (rat) OPPTS 870.1100	>5000 mg EP/kg 1

(200 mg a.i./kg)	CSH	Practically non-toxic for EP 1	44931205

	>5000 mg EP/kg 2

(40 mg a.i./kg)	CSH	Practically non-toxic for EP 2	46790003

Freshwater Fish LC50/OPPTS 850.1075 and 

Freshwater Invertebrate/OPPTS 850.1010	See Non-target Aquatic Organism
summary below	

C and CS	Practically non-toxic 3	-

Avian Acute Oral/OPPTS 850.2100 

and

Avian Dietary/OPPTS 850.2200	See Non-target Bird summary below	

C and CS	Practically non-toxic 3	-

Non-target Plants/OPPTS 850.4000 

and 

Non-target Insects/OPPTS 850/3020; 850.3030; and/or 850.3040	See
Non-target Plants and Insects summary below	

C	Practically non-toxic 3	-

1	Elexa-4 (EPA Reg. No. 81045-2) contains 4.0% CSH.

2	Keyplex 350 (EPA File Symbol No. 73512-L) contains 0.8% CSH.  

3	Based on the non-toxic mode of action of the active ingredients, the
presence of chitin as a naturally occurring component of many non-target
organisms, and the presence of chitinases (enzymes that metabolize
chitin) in many non-target organisms that consume chitin-containing
organisms in their normal diets.  

Non-Target Mammals 

An end-use product containing 4% Chitosan hydrolysate has been shown to
be practically non-toxic to rats on acute oral basis (see MRID
44931205).  No adverse effects are expected to mammalian wildlife when
chitin and chitosan-containing products are applied in accordance with
approved labeling.  

Non-Target Aquatic Organisms

Chitin is ubiquitous in nature and is a major component of the
exoskeletons of aquatic arthropods (insects, crustaceans), the radula of
molluscs, and the beaks of cephalpods (squid, octopus), the cell walls
of fungi (Campbell, 1996), and the scales of fish (Uawonggul et al.
2002).  Chitin is a normal component in the diets of fish (e.g.  in the
exoskeletons of aquatic arthropods and in the scales of prey fish) and
fish possess chitinases that metabolize these dietary chitins (Matsumiya
et al. 2006).  No mortalities have been observed in fish fed with diets
supplemented with chitin and chitosan.  No adverse effects are expected
to non-target aquatic organisms when chitin and chitosan-containing
products are applied in accordance with approved labeling.  

Non-Target Birds:  

Chitin is present in the exoskeletons of arthropods and, therefore, is a
regular component of the diets of insectivorous birds.  Chitin and
chitosan have been shown to be relatively indigestible by some birds
(Akaki and Duke, 1999; and Razdan and Pettersson, 1994).  However, many
birds (starlings, raptors, and many seabirds) possess chitinases that
aid in the digestion of chitin, which serves as a source of protein
(MacDonald, 2006).  No mortalities have ever been observed in birds fed
with diets supplemented with chitin and chitosan.  No adverse effects
are expected to birds when chitin and chitosan-containing products are
applied in accordance with approved labeling.  

Non-target Plants and Insects

Chitin is ubiquitous in nature and is a major component of the
exoskeletons of aquatic arthropods (insects, crustaceans). Chitin,
Chitosan, and Chitosan Hydrolysate are intended for use as plant defense
"boosters" to protect plants from fungal pathogens via he induction of
Systemic Acquired Response (SAR).  SAR is a mechanism that stimulates
the internal defense mechanisms of plants to resist pathogen infection. 
In plants, chitin stimulates the production of chitinases which are used
to degrade the chitin-containing cell walls of pathogenic fungi (Kramer
and Muthukrishnan, 1998).  There is no direct activity of Chitin or
Chitosan (including Chitosan Hydrolysate) against the pathogen.  No
adverse effects to non-target insects and plants are anticipated.  

REFERENCES

Bishop, J. A. Dean, and T. Mitchell-Olds.  2000.  PNAS 97(10): 
5322-5327.  

Campbell, N. A. 1996.  Biology (4th edition) Benjamin Cummings, New
York.   HYPERLINK
"http://en.wikipedia.org/wiki/Special:BookSources/0805319573"  ISBN
0-8053-1957-3 

Escott GM, Adams DJ (1995). Infection and Immunity 63(12): 4770-4773. 

Kaspari, M. and A. Joern.  1993.  OIKOS 68:  410-430.  

Kramer, K. and  S. Muthukrishnan.  1998.  Insect Biochemistry and
Biology 27(11):  887-900.

MacDonald, D. 2006.  Protein Requirements of Birds.    HYPERLINK
"http://www.drmacs.com/ProteinRequirementsofBirds.pdf" 
http://www.drmacs.com/ProteinRequirementsofBirds.pdf 

Matsumiya, M., Y. Arikane, A. Haga, S. Muthukrishnan, and K. Kramer. 
2006.  Bioscience, Biotechnology, and Biochemistry 70(4):  971-979.

Paolettia MG, L. Norbertob, R. Daminia, S. Musumeci . 2007.  Annals of
Nutrition and Metabolism 51(3): 244-251.

Razdan, A. and D. Pettersson.  1994.  British Journal of Nutrition. 1994
72:  277-288.

Uawonggul, N., C. Ruksakulpiwat, and S. Chanthai. 2002.  Poster
Abstract.  28th Congress on Science and Technology of Thailand, Queen
Sirikit National Convention Center, Bangkok, Thailand, Oct. 24-26.  

cc:  R. S. Jones, BPPD Subject File/IHAD

	R. S. Jones, FT, OPY:  04/08/2008

 PAGE  7 

Chitin and Chitosan		Reg. Review Case No.:  6069

PC Codes:  128991 & 128930					    	   Initial Docket:  September 2007