Professional Documents
Culture Documents
National
Standard
ANSI/AAMI RD52:2004
Dialysate
for
hemodialysis
ANSI/AAMI RD52:2004
Developed by
Association for the Advancement of Medical Instrumentation
Approved 9 August 2004 by
American National Standards Institute, Inc.
Abstract:
Keywords:
Published by
Association for the Advancement of Medical Instrumentation
1110 N. Glebe Road, Suite 220
Arlington, VA 22201-4795
2004 by the Association for the Advancement of Medical Instrumentation
All Rights Reserved
Publication, reproduction, photocopying, storage, or transmission, electronically or otherwise, of all or any part of this
document without the prior written permission of the Association for the Advancement of Medical Instrumentation is
strictly prohibited by law. It is illegal under federal law (17 U.S.C. 101, et seq.) to make copies of all or any part of
this document (whether internally or externally) without the prior written permission of the Association for the
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damages of $100,000 per offense. For permission regarding the use of all or any part of this document, contact AAMI
at 1110 N. Glebe Road, Suite 220, Arlington, VA 22201-4795. Phone: (703) 525-4890; Fax: (703) 525-1067.
Printed in the United States of America
ISBN 1570202230
Contents
Page
Glossary of equivalent standards .....................................................................................................................v
Committee representation .............................................................................................................................. vii
Foreword ....................................................................................................................................................... viii
Introduction: Need for this AAMI recommended practice ................................................................................ ix
1
Scope........................................................................................................................................................1
1.1
1.2
1.3
General ..........................................................................................................................................1
Inclusions .......................................................................................................................................1
Exclusions ......................................................................................................................................1
Normative references................................................................................................................................1
Definitions .................................................................................................................................................2
Fluid quality...............................................................................................................................................5
4.1
4.2
4.3
Water .............................................................................................................................................5
4.1.1
Maximum level of chemical contaminants in water ..........................................................5
4.1.2
Bacteriology of water .......................................................................................................6
Concentrate....................................................................................................................................7
4.2.1
Maximum level of chemical contaminants in concentrate ................................................7
4.2.2
Bacteriology of concentrate..............................................................................................7
Dialysate ........................................................................................................................................7
4.3.1
Maximum level of chemical contaminants in dialysate .....................................................7
4.3.2
Bacteriology of dialysate ..................................................................................................7
Equipment.................................................................................................................................................8
5.1
5.2
5.3
5.4
5.5
5.6
General ..........................................................................................................................................8
Water purification systems .............................................................................................................8
5.2.1
General ............................................................................................................................8
5.2.2
Sediment filters ................................................................................................................8
5.2.3
Cartridge filters ................................................................................................................8
5.2.4
Softeners .........................................................................................................................9
5.2.5
Carbon adsorption ...........................................................................................................9
5.2.6
Chemical injection systems............................................................................................10
5.2.7
Reverse osmosis ...........................................................................................................10
5.2.8
Deionization ...................................................................................................................10
5.2.9
Ultrafiltration...................................................................................................................11
Water storage and distribution .....................................................................................................11
5.3.1
General ..........................................................................................................................11
5.3.2
Water storage ................................................................................................................11
5.3.3
Water distribution systems.............................................................................................12
5.3.4
Bacterial control devices ................................................................................................12
Concentrate preparation...............................................................................................................13
5.4.1
General ..........................................................................................................................13
5.4.2
Materials compatibility....................................................................................................14
5.4.3
Bulk storage tanks (acid concentrate)............................................................................14
5.4.4
Mixing systems ..............................................................................................................14
5.4.5
Additives ........................................................................................................................15
Concentrate distribution ...............................................................................................................15
5.5.1
Materials compatibility....................................................................................................15
5.5.2
System configurations....................................................................................................16
5.5.3
Acid concentrate distribution systems............................................................................16
5.5.4
Bicarbonate concentrate distribution systems................................................................16
5.5.5
Concentrate outlets........................................................................................................16
Dialysate proportioning.................................................................................................................17
Monitoring ...............................................................................................................................................18
6.1
6.2
6.3
6.4
6.5
6.6
7
General ........................................................................................................................................18
Water purification .........................................................................................................................19
6.2.1
General ..........................................................................................................................19
6.2.2
Sediment filters ..............................................................................................................20
6.2.3
Cartridge filters ..............................................................................................................20
6.2.4
Softeners .......................................................................................................................20
6.2.5
Carbon adsorption .........................................................................................................20
6.2.6
Chemical injection systems............................................................................................21
6.2.7
Reverse osmosis ...........................................................................................................21
6.2.8
Deionization ...................................................................................................................21
6.2.9
Ultrafiltration...................................................................................................................22
Water storage and distribution .....................................................................................................22
6.3.1
General ..........................................................................................................................22
6.3.2
Water storage ................................................................................................................22
6.3.3
Water distribution systems.............................................................................................22
6.3.4
Bacterial control devices ................................................................................................22
Concentrate preparation...............................................................................................................23
6.4.1
Mixing systems ..............................................................................................................23
6.4.2
Additives ........................................................................................................................23
Concentrate distribution ...............................................................................................................23
Dialysate proportioning.................................................................................................................23
General ........................................................................................................................................24
Microbial monitoring methods ......................................................................................................25
7.2.1
General ..........................................................................................................................25
7.2.2
Sample collection...........................................................................................................27
7.2.3
Heterotrophic plate count...............................................................................................27
7.2.4
Bacterial endotoxin test..................................................................................................27
Environment............................................................................................................................................28
Personnel................................................................................................................................................28
Annexes
A
Bibliography ............................................................................................................................................39
Tables
1
Maximum allowable chemical contaminant levels in water used to prepare dialysate and
concentrates from powder at a dialysis facility and to reprocess dialyzers for multiple uses.....................6
Compatibility of common disinfectants with piping materials used in water distribution systems ...........12
Symbols and color coding for different concentrate proportioning ratios .................................................17
Monitoring guidelines for water purification equipment and distribution systems and dialysate .............18
Example of decision tree that can be used to evaluate culture results and initiate corrective action,
if necessary.............................................................................................................................................26
U.S. designation
Equivalency
IEC 60601-1-2:2001
ANSI/AAMI/IEC 60601-1-2:2001
Identical
IEC 60601-2-04:2002
ANSI/AAMI DF80:2003
Identical
IEC 60601-2-24:1998
ANSI/AAMI ID26:1998
IEC TR 60878:2003
ANSI/AAMI/IEC TIR60878:2003
Identical
IEC TR 62296:2003
ANSI/AAMI/IEC TIR62296:2003
Identical
ISO 5840:1996
ANSI/AAMI/ISO 5840:1996
Identical
ISO 7198:1998
ANSI/AAMI/ISO 7198:1998/2001
Identical
ISO 7199:1996
ANSI/AAMI/ISO 7199:1996/(R)2002
Identical
ISO 10993-1:2003
ANSI/AAMI/ISO 10993-1:2003
Identical
ISO 10993-2:1992
ANSI/AAMI/ISO 10993-2:1993/(R)2001
Identical
ISO 10993-3:2003
ANSI/AAMI/ISO 10993-3:2003
Identical
ISO 10993-4:2002
ANSI/AAMI/ISO 10993-4:2002
Identical
ISO 10993-5:1999
ANSI/AAMI/ISO 10993-5:1999
Identical
ISO 10993-6:1994
ANSI/AAMI/ISO 10993-6:1995/(R)2001
Identical
ISO 10993-7:1995
ANSI/AAMI/ISO 10993-7:1995/(R)2001
Identical
ISO 10993-8:2000
ANSI/AAMI/ISO 10993-8:2000
Identical
ISO 10993-9:1999
ANSI/AAMI/ISO 10993-9:1999
Identical
ISO 10993-10:2002
ANSI/AAMI BE78:2002
ISO 10993-11:1993
ANSI/AAMI 10993-11:1993
ISO 10993-12:2002
ANSI/AAMI/ISO 10993-12:2002
Identical
ISO 10993-13:1998
ANSI/AAMI/ISO 10993-13:1999
Identical
ISO 10993-14:2001
ANSI/AAMI/ISO 10993-14:2001
Identical
ISO 10993-15:2000
ANSI/AAMI/ISO 10993-15:2000
Identical
ISO 10993-16:1997
ANSI/AAMI/ISO 10993-16:1997/(R)2003
Identical
ISO 10993-17:2002
ANSI/AAMI/ISO 10993-17:2002
Identical
ISO 11134:1994
ANSI/AAMI/ISO 11134:1993
Identical
ISO 11135:1994
ANSI/AAMI/ISO 11135:1994
Identical
Identical
ISO 11138-1:1994
ANSI/AAMI ST59:1999
International designation
U.S. designation
Equivalency
ISO 11138-2:1994
ANSI/AAMI ST21:1999
ISO 11138-3:1995
ANSI/AAMI ST19:1999
ISO TS 11139:2001
ANSI/AAMI/ISO 11139:2002
Identical
ANSI/AAMI ST60:1996
ISO 11607:2003
ANSI/AAMI/ISO 11607:2000
Identical
ISO 11737-1:1995
ANSI/AAMI/ISO 11737-1:1995
Identical
ISO 11737-2:1998
ANSI/AAMI/ISO 11737-2:1998
Identical
ISO TR 13409:1996
AAMI/ISO TIR13409:1996
Identical
ISO 13485:2003
ANSI/AAMI/ISO 13485:2003
Identical
ISO 13488:1996
ANSI/AAMI/ISO 13488:1996
Identical
ISO 14155-1:2003
ANSI/AAMI/ISO 14155-1:2003
Identical
ISO 14155-2:2003
ANSI/AAMI/ISO 14155-2:2003
Identical
ISO 14160:1998
ANSI/AAMI/ISO 14160:1998
Identical
ANSI/AAMI/ISO 14161:2000
Identical
ANSI/AAMI/ISO 14937:2000
Identical
ISO TR 14969:2004
ANSI/AAMI/ISO TIR14969:2004
Identical
Identical
ISO 15223:2000
ANSI/AAMI/ISO 15223:2000
Identical
ISO 15223/A1:2002
ANSI/AAMI/ISO 15223:2000/A1:2001
Identical
ISO 15223/A2:2004
ANSI/AAMI/ISO 15223:2000/A2:2004
Identical
ISO 15225:2000
ANSI/AAMI/ISO 15225:2000
Identical
ISO 15225/A1:2004
ANSI/AAMI/ISO 15225:2000/A1:2004
Identical
ISO 15674:2001
ANSI/AAMI/ISO 15674:2001
Identical
ISO 15675:2001
ANSI/AAMI/ISO 15675:2001
Identical
ISO TS 15843:2000
ANSI/AAMI/ISO TIR15843:2000
Identical
ISO TR 15844:1998
AAMI/ISO TIR15844:1998
Identical
ISO TR 16142:1999
ANSI/AAMI/ISO TIR16142:2000
Identical
ISO 25539-1:2003
ANSI/AAMI/ISO 25539-1:2003
Identical
ISO 14937:2000
1
vi
Committee representation
Association for the Advancement of Medical Instrumentation
Renal Disease and Detoxification Committee
This recommended practice was developed by the AAMI Renal Disease and Detoxification Committee. Committee
approval of this recommended practice does not necessarily imply that all committee members voted for its approval.
At the time this document was published, the AAMI Renal Disease and Detoxification Committee had the following
members:
Cochairs:
Members:
Alternates:
LeRoy J. Fischbach
Richard A. Ward, PhD
Steven Acres, MD, Carolina Regional Nephrology Associates
Matthew J. Arduino, DrPH, U.S. Centers for Disease Control and Prevention
Robert Berube, Church & Dwight Company, Inc.
D. Michael Blankenship, MD, Texarkana Kidney Disease and Hypertension Center
Danilo B. Concepcion, CHT, CCHT, St. Joseph Hospital Renal Center
R. Barry Deeter, RN, MSN, University of Utah Dialysis Program
Robert Dudek, US Filter
Martin S. Favero, PhD, Johnson & Johnson Advanced Sterilization Products
LeRoy J. Fischbach, Minntech Corporation
Gema Gonzalez, U.S. Food and Drug Administration, Center for Devices and Radiological Health,
Office of Device Evaluations
Bertrand L. Jaber, MD, Tufts University School of Medicine, Caritas St. Elizabeth's Medical Center
Jerome C. James III, PhD, Aksys Limited
James M. Kaar, Baxter Healthcare Corporation
Fei M. Law, Gambro Renal Products, Inc.
Nathan W. Levin, MD, Renal Research Institute LLC
John Lohr, PhD, Associates of Cape Cod, Inc.
Douglas A. Luehmann, DaVita, Inc.
Bruce H. Merriman, Central Florida Kidney Centers
Glenda Payne, RN, MS, CNN, Centers for Medicare & Medicaid Services
John A. Rickert, GE Osmonics
Mark Rolston, Renal Care Group
James D. Stewardson, Brighton, CO
David S. Utterberg, Medisystems Services Corporation
Richard A. Ward, PhD, Kidney Disease Program, University of Louisville
Robert J. Chambers, Gambro BCT Inc.
Christine Colon, Baxter Healthcare Corporation
Conor Curtin, Fresenius Medical Care NA Dialysis Products Division
Russell Dimmitt, AS, Renal Care Group
Mark David Einzinger, BA, MS, Church & Dwight Company Inc.
Barbara I. McCool, U.S. Food and Drug Administration, Center for Devices and Radiological Health,
Office of Device Evaluations
Gregory Montgomery, US Filter
NOTEParticipation by federal agency representatives in the development of this recommended practice does not
constitute endorsement by the federal government or any of its agencies.
Acknowledgment
The AAMI Renal Disease and Detoxification Committee dedicates this recommended practice to the late
Scott N. Walker of Fresenius USA for his outstanding contributions to AAMI dialysis standards work.
vii
Foreword
This recommended practice was developed by the AAMI Renal Disease and Detoxification Committee. The
committees objective is to provide rational guidelines for handling water and concentrates and for the production and
monitoring of dialysate used for hemodialysis. The need for such guidelines is based on the critical role of dialysate
quality in providing safe and effective hemodialysis, and the recognition that day-to-day dialysate quality is under the
control of the health care professionals who deliver dialysis therapy.
This recommended practice reflects the conscientious efforts of health care professionals, patients, medical device
manufacturers, and representatives of federal agencies to develop recommendations for handling water and
concentrates and for the production and monitoring of dialysate for hemodialysis. The document is intended as a
guide for physicians, particularly the directors of dialysis facilities. The recommendations contained in this document
may not be applicable in all circumstances and they are not intended for regulatory application. The term should as
used in this document reflects the committees intent to define goals, not requirements. The term shall as used here
denotes quality recommendations and procedures that are required by applicable standards. The term must is used
only to describe unavoidable situations, including those mandated by government regulation.
The concepts incorporated in this recommended practice should not be considered inflexible or static. The
recommendations presented here should be reviewed periodically in order to assimilate increased understanding of
the role of dialysate purity in patient outcomes and technological developments.
Suggestions for improving this recommended practice are invited and should be sent to: AAMI, Attn: Standards
Department, 1110 N. Glebe Road, Suite 220, Arlington, VA 22201-4795.
NOTEThis foreword does not contain provisions of the American National Standard ANSI/AAMI RD52:2004, Dialysate
for hemodialysis.
viii
ix
ANSI/AAMI RD52:2004
Scope
1.1
General
The intent of this recommended practice is to provide dialysis practitioners with guidance on the preparation of
dialysate for hemodialysis and related therapies, from the point at which municipal water enters their dialysis facility to
the point at which the final dialysate enters the dialyzer. Included in the scope of the recommended practice are: (1)
use, maintenance, and monitoring of equipment used to purify and distribute water used for the preparation of
dialysate and other hemodialysis applications; (2) use, maintenance, and monitoring of equipment used to prepare
concentrate from powder at a dialysis facility; and (3) preparation of the final dialysate from purified water and
concentrate. The equipment used in the various stages of dialysate preparation is generally obtained from specialized
vendors. This recommended practice provides a general description of the system components that these vendors
may provide. These descriptions are intended to provide the user with a basis for understanding why certain
equipment may be required and how it should be configured; they are not intended as detailed design standards.
Dialysis practitioners are generally responsible for maintaining the equipment used to prepare dialysate following its
installation. Therefore, this recommended practice provides guidance on monitoring and maintenance of the
equipment to ensure that dialysate quality is acceptable at all times. At various places throughout this recommended
practice, the user is advised to follow the manufacturers instructions regarding the operation and maintenance of
equipment. In those instances in which the equipment is not obtained from a specialized vendor, it is the responsibility
of the user to validate the performance of the equipment in the hemodialysis setting and to ensure that appropriate
operating and maintenance manuals are available.
The guidance provided by this recommended practice should help protect hemodialysis patients from adverse effects
arising from known chemical and microbial contaminants that may be found in improperly prepared dialysate.
However, the physician in charge of dialysis has the ultimate responsibility for ensuring that the dialysate is correctly
formulated and meets the requirements of all applicable quality standards.
1.2
Inclusions
This recommended practice addresses the users responsibility for the dialysate once equipment has been delivered
and installed. For the purposes of this recommended practice, the dialysate includes water used for the preparation of
dialysate, water used for the preparation of concentrates at the users facility, and water used for the preparation of
ultrapure dialysate, as well as the final dialysate and concentrates. Because it is commonly prepared and distributed
using the same equipment as the water used to prepare dialysate, water used to reprocess dialyzers is also covered
by this recommended practice.
1.3
Exclusions
Excluded from the scope of this recommended practice are sorbent-based dialysate regeneration systems that
regenerate and recirculate small volumes of dialysate, systems for continuous renal replacement therapy that use
prepackaged solutions, and systems and solutions for peritoneal dialysis. This recommended practice excludes home
hemodialysis, although this document may be of use to the home hemodialysis practitioner.
Normative references
The following documents contain provisions that, through reference in this text, constitute provisions of this
recommended practice. At the time of publication, the editions indicated were valid. All standards are subject to
revision. For this reason, it is recommended that the user obtain the most recent editions of the documents indicated
below. The Association for the Advancement of Medical Instrumentation maintains a register of currently valid
AAMI/American National Standards.
2.1
U.S. FOOD AND DRUG ADMINISTRATION. Guidance for the Content of Premarket Notifications for Water
Purification Components and Systems for Hemodialysis. Rockville (MD): U.S. Food and Drug Administration, 1997.
<http://www.fda.gov/cdrh/ode/hemodial.pdf>
2.2
ASSOCIATION FOR THE ADVANCEMENT OF MEDICAL INSTRUMENTATION. Hemodialysis systems
(ANSI/AAMI RD5:2003). Arlington (VA): AAMI, 2003. American National Standard.
2.3
ASSOCIATION FOR THE ADVANCEMENT OF MEDICAL INSTRUMENTATION. Water treatment equipment
for hemodialysis applications (ANSI/AAMI RD62:2001). Arlington (VA): AAMI, 2001. American National Standard.
2.4
ASSOCIATION FOR THE ADVANCEMENT OF MEDICAL INSTRUMENTATION. Concentrates for
hemodialysis (ANSI/AAMI RD61:2000). Arlington (VA): AAMI, 2000. American National Standard.
2.5
UNITED STATES PHARMACOPEIAL CONVENTION, INC. United States PharmacopoeiaNational
Formulary (USP-26-NF21). Rockville (MD): United States Pharmacopeial Convention Inc., 2003.
2.6
U.S. ENVIRONMENTAL PROTECTION AGENCY. Safe Drinking Water Act, 1996 (Public law 104182).
Washington (DC): EPA, 1996.
(See also National Primary and Secondary Drinking Water Regulations. U.S. Environmental Protection Agency, Office
of Ground Water and Drinking Water. <http://www.epa.gov/OGWDW/creg.html>)
2.7
ASSOCIATION FOR THE ADVANCEMENT OF MEDICAL INSTRUMENTATION. Bacterial endotoxinsTest
methodologies, routine monitoring, and alternatives to batch testing (ANSI/AAMI ST72:2002). Arlington (VA): AAMI,
2002. American National Standard.
Definitions
For the purposes of this recommended practice, the following terms and definitions apply.
3.1
acetate concentrate: Concentrated solution of salts that may contain dextrose (sometimes referred to as
glucose), which, when diluted with water, yields dialysate for use in dialysis. Sodium acetate is normally used as the
buffer. This concentrate is used as a single concentrate.
3.2
acetate dialysate: Dialysate without bicarbonate, using acetate as a substitute for the bicarbonate buffer.
NOTEAcetate dialyzing fluid is generally produced from a single concentrate. Acetate is metabolized by the patient to produce
bicarbonate.
3.3
acid concentrate: Acidified concentrated solution of salts that may contain dextrose (sometimes referred to as
glucose), which, when diluted with water and bicarbonate concentrate, yields dialysate for use in dialysis. The term
acid refers to the small amount of acid (usually acetic acid) that is included in the concentrate to establish the buffer
system in the final dialysate by reaction with a small amount of bicarbonate from the bicarbonate concentrate.
3.4
action level: Concentration of a contaminant at which steps should be taken to interrupt the trend toward
higher, unacceptable levels.
3.5
3.6
bacteriology: Area of study within the field of microbiology that deals with the study of bacteria.
3.7
batch system: Apparatus in which the dialysate is prepared in bulk before each dialysis session.
3.8
bicarbonate concentrate: Concentrated solution of sodium bicarbonate that, when diluted with water and acid
concentrate, makes dialysate used for dialysis. Some bicarbonate concentrates also contain sodium chloride.
3.9
NOTEBicarbonate dialysate is generally produced from two concentrates: one containing bicarbonate and the other containing
acid and other electrolytes (see acid concentrate and bicarbonate concentrate).
3.10 biofilm: Coating on surfaces that consists of microcolonies of bacteria embedded in a protective extracellular
matrix. The matrix, a slimy material secreted by the cells, protects the bacteria from antibiotics and chemical
disinfectants.
3.11 bulk delivery: Delivery of large volumes of concentrate in which the product is transferred (pumped) from the
delivery container to a users storage tank. Bulk delivery may include containers such as 55 gallon drums, which are
pumped into a storage tank maintained at the user's facility.
3.12
3.13
NOTEThere is no direct test for measuring combined chlorine, but it can be measured indirectly by measuring both total and free
chlorine and calculating the difference.
3.14
3.15 colony-forming unit (CFU): Organism capable of replicating to form a distinct, visible colony on a culture
plate. In practice, a colony may be formed by a group of organisms.
3.16 concentrate generators: System in which the concentrate is delivered to the consumer as a powder in a
container and then converted on-line into a saturated solution by a dialysis delivery machine. This solution is used by
an individual proportioning system to make the final dialysate delivered to the dialyzer.
3.17 dialysate: Aqueous fluid containing electrolytes and, usually, dextrose, which is intended to exchange solutes
with blood during hemodialysis. The word dialysate is used throughout this document to mean the fluid made from
water and concentrates that is delivered to the dialyzer by the dialysate supply system. Such phrases as dialyzing
fluid or dialysis solution may be used in place of dialysate.
3.18 dialysate supply system: Devices that: (1) prepare dialysate on-line from water and concentrates or that
store and distribute premixed dialysate; (2) circulate the dialysate through the dialyzer; (3) monitor the dialysate for
temperature, conductivity (or equivalent), pressure, flow, and blood leaks; and (4) prevent dialysis during disinfection
or cleaning modes. The term includes reservoirs, conduits, proportioning devices for the dialysate, and monitors and
associated alarms and controls assembled as a system for the characteristics listed above. The dialysate supply
system is often an integral part of single-patient dialysis machines (see 2.2).
3.19 disinfection: Destruction of pathogenic and other kinds of microorganisms by thermal or chemical means.
Disinfection is a less lethal process than sterilization, because it destroys most recognized pathogenic
microorganisms but does not necessarily destroy all microbial forms. This definition of disinfection is equivalent to
low-level disinfection in the Spalding classification.
3.20 electrolyte: Ion capable of transferring or exchanging electrons. In dialysate, the electrolytes are the charged
ions that result from dissociation of salts when they are dissolved in water. These charged ions are responsible for the
conductive property of dialysate.
3.21 empty-bed contact time (EBCT): Measure of how much contact occurs between particles, such as activated
carbon, and water as the water flows through a bed of the particles. EBCT (minutes) is calculated from the following
equation:
EBCT = (7.48 x V)/Q
where V is the volume of particles in the bed (ft3),
Q is the flow rate of water through the bed (gal/min), and
7.48 is the conversion factor for gallons to ft3.
3.22 endotoxin: Major component of the outer cell wall of gram-negative bacteria. Endotoxins are
lipopolysaccharides, which consist of a polysaccharide chain covalently bound to lipid A. Endotoxins can acutely
activate both humoral and cellular host defenses, leading to a syndrome characterized by fever, shaking chills,
hypotension, multiple organ failure, and even death if allowed to enter the circulation in a sufficient dose. (See also
pyrogen.)
3.23 endotoxin units (EU): Units assayed by the Limulus amoebocyte lysate (LAL) method when testing for
endotoxins. Because activity of endotoxins differs on a mass basis, their activity is referred to a standard E. coli
endotoxin. The current standard (EC-6-1) is prepared from E. coli O:113:H10. The relationship between the mass of
an endotoxin and its activity varies with both the lot of LAL and the lot of control standard endotoxin being used. Since
standards for endotoxin were harmonized in 1983 with the introduction of EC-5, the relationship between mass and
activity of endotoxin has been approximately 10 EU/ng.
NOTEIn some countries, endotoxin concentrations are expressed in international units (IU). Since the 1983 harmonization of
endotoxin assays, EU and IU are equivalent.
3.24
feed water: Water supplied to a water treatment system or an individual component of the system.
3.25
3.26 hemodiafiltration: Form of renal replacement therapy in which waste solutes are removed from blood by a
combination of diffusion and convection through a high-flux membrane. Diffusive solute removal is achieved using a
dialysate stream as in hemodialysis. Convective solute removal is achieved by adding ultrafiltration in excess of that
needed to obtain the desired weight loss; fluid balance is maintained by infusing a replacement solution into the blood
either before the dialyzer (predilution hemodiafiltration) or after the dialyzer (postdilution hemodiafiltration).
3.27 hemodialysis: Form of renal replacement therapy in which waste solutes are removed primarily by diffusion
from blood flowing on one side of a membrane into dialysate flowing on the other side. Fluid removal that is sufficient
to obtain the desired weight loss is achieved by establishing a hydrostatic pressure gradient across the membrane.
This fluid removal provides some additional waste solute removal, particularly for higher molecular weight solutes.
3.28 hemofiltration: Form of renal replacement therapy in which waste solutes are removed from blood by
convection. Convective transport is achieved by ultrafiltration through a high-flux membrane. Fluid balance is
maintained by infusing a replacement solution into the blood either before the hemofilter (pre-dilution hemofiltration) or
after the hemofilter (post-dilution hemofiltration).
NOTEThere is no dialysate stream in hemofiltration.
3.29 Limulus amoebocyte lysate (LAL) test: Assay used to detect endotoxin. Exploits the immune response of
the horseshoe crab to endotoxin (Limulus polyphemus).
3.30 manufacturer: Person who designs, manufactures, fabricates, assembles, or processes a finished device.
Manufacturers include, but are not limited to, those who perform the functions of contract sterilization, installation,
relabeling, remanufacturing, repacking, or specification development, and initial distributors of foreign entities
performing these functions.
3.31
microbial: Referring to microscopic organisms, bacteria, fungi, and so forth. (See also bacteriology.)
3.32 microbiological contamination: Contamination with any form of microorganism (e.g., bacteria, yeast, fungi,
and algae) or with the by-products of living or dead organisms such as endotoxins, exotoxins, and microcystin
(derived from blue-green algae).
3.33 microfilter: Filter designed to remove particles in the range 0.1 m to 3 m in diameter. Microfilters have an
absolute size cut-off and are available in both dead-end and cross-flow configurations.
3.34
product water: Water produced by a water treatment system or by an individual component of a system.
3.35
proportioning system: Apparatus that proportions water and hemodialysis concentrate to prepare dialysate.
3.36 pyrogen: Fever-producing substance. Note that pyrogens are most often lipopolysaccharides of gramnegative bacterial origin. (See also endotoxin.)
3.37 spike: Small amount of a single chemical used to increase a constituent or constituents in the concentrate for
a single patients treatment.
NOTEThe spike may be in the form of a dry chemical or may be dissolved in water.
3.38 sterile: Free from all living organisms and viable spores, within the limits of tests for sterility, and maintained in
that state by suitable means.
NOTEFor solutions used in hemodialysis and related therapies, sterile can be used to describe a packaged solution that was
prepared using a terminal sterilization process that has been demonstrated to achieve a six log reduction in appropriate indicator
microorganisms. Alternatively, sterile can be used to describe a solution prepared for immediate use by a continuous process that
has been validated to produce a solution free of microorganisms when challenged with a defined test fluid and that is capable of
meeting this challenge even with a single fault failure.
3.39 storage tank: Large tank at the users facility for storage of purified water or concentrate from bulk deliveries,
or for concentrate prepared in bulk at the user's facility from powder and purified water.
3.40 total dissolved solids (TDS): Sum of all ions in a solution, often approximated by means of electrical
conductivity or resistivity measurements. TDS measurements are commonly used to assess the performance of
reverse osmosis units. TDS values are often expressed in terms of CaCO3 or NaCl equivalents (ppm).
3.41 ultrafilter: Membrane filter with a pore size in the range 0.001 m to 0.05 m. Performance is usually rated in
terms of a nominal molecular weight cut-off (MWCO), which is defined as the smallest molecular weight species for
which the filter membrane has more than 90 % rejection. Depending on their nominal MWCO, ultrafilters may be used
to remove particles and solutes as small as 1,000 dalton. Ultrafilters with a nominal MWCO of 20,000 or less are
generally adequate for endotoxin removal.
NOTEUltrafilters are usually configured in a cross-flow mode. Some ultrafilters also remove endotoxins by adsorption.
3.42 ultrapure dialysate: Highly purified dialysate that can be used in place of conventional dialysate or as feed
solution for further processing to create fluid intended for injection directly into the blood. The definition of ultrapure
dialysate varies, but the recommendation used in the European Renal AssociationEuropean Dialysis and
Transplant Association (ERA/EDTA) Best Practice Guidelines (ERAEDTA, 2002) is < 0.1 CFU/mL and
< 0.03 EU/mL. The endotoxin level may be reduced in the future as more sensitive tests become available.
3.43
United States Pharmacopoeia (USP): The current version of this official compendium (see 2.5).
3.44
user: Physician or physicians representative responsible for the actual production and handling of dialysate.
3.45 water treatment system: Collection of water purification devices and associated piping, pumps, valves,
gauges, etc., that together produce purified water for hemodialysis applications and deliver it to the point of use.
Fluid quality
4.1
Water
The requirements contained in clause 4.1 apply to the purified water as it enters the equipment used to prepare
dialysate or concentrates from powder at a dialysis facility. As such, these requirements apply to the water treatment
system as a whole and not to each of the devices that make up the system. However, collectively the individual
devices shall produce water that, at a minimum, meets the requirements of this clause.
4.1.1 Maximum level of chemical contaminants in water
Product water used to prepare dialysate or concentrates from powder at a dialysis facility, or to process dialyzers for
reuse, shall not contain chemical contaminants at concentrations in excess of those listed in ANSI/AAMI RD62 (see
2.3), which is reproduced in Table 1 below. The manufacturer or supplier of a complete water treatment system
should recommend a system that is capable of meeting the requirements of this clause given the analysis of the feed
water. The system design should reflect possible seasonal variations in feed water quality. The manufacturer or
supplier of a complete water treatment and distribution system shall demonstrate that the complete water treatment,
storage, and distribution system is capable of meeting the requirements of RD62 at the time of installation.
Following installation of a water treatment, storage, and distribution system, the user is responsible for continued
monitoring of the levels of chemical contaminants in the water and for complying with the requirements of this
standard.
Table 1Maximum allowable chemical contaminant levels in water used to prepare dialysate and
concentrates from powder at a dialysis facility and to reprocess dialyzers for multiple uses
(Reproduced from ANSI/AAMI RD62:2001)
Contaminant
Calcium
2 (0.1 mEq/L)
Magnesium
4 (0.3 mEq/L)
Potassium
8 (0.2 mEq/L)
Sodium
70 (3.0 mEq/L)
Antimony
0.006
Arsenic
0.005
Barium
0.10
Beryllium
0.0004
Cadmium
0.001
Chromium
0.014
Lead
0.005
Mercury
0.0002
Selenium
0.09
Silver
0.005
Aluminum
0.01
Chloramines
0.10
Free Chlorine
0.50
Copper
0.10
Fluoride
0.20
Nitrate (as N)
2.0
Sulfate
100
Thallium
0.002
Zinc
0.10
NOTEAmerican National Standards are revised every three to five years. Users
should consult the most recent edition of ANSI/AAMI RD62 to ensure that the levels
listed in this table are still valid.
4.2
Concentrate
Dialysate
installation and confirmed by the user with a regular monitoring and maintenance schedule. The user should follow
the manufacturers instructions for use of the validated system, and the users monitoring and maintenance schedule
should be designed to confirm that the dialysate used to prepare the dialysate for infusion continues to meet the
specifications of the manufacturer of the device or process.
Equipment
5.1
General
This clause provides a brief description of the different components that may be included in a water purification and
distribution system used for hemodialysis applications. Since feed water quality and product water requirements may
vary from facility to facility, not all of the components described in the following clauses will be necessary in every
purification and distribution system.
Routine dialysis requires a well-functioning water purification and distribution system, since dialysis cannot be
performed without an adequate supply of water. In addition, certain components of the water purification and
distribution system are critical to its operation. An example of such a critical component is the circulating pump in an
indirect feed system. A dialysis facility should develop contingency plans to cover the failure of its water purification
and distribution system or a critical component of that system. Such contingency plans should describe how to deal
with events that completely prevent dialysis from being performed, such as failure of the facilitys municipal water
supply or electrical service following a natural disaster or water main break. Other plans should address how to deal
with sudden changes in municipal water quality, as well as with failure of a critical component of the water purification
and distribution system.
5.2
5.2.1 General
Water purification systems consist of three basic sections: a pretreatment section that conditions the water supplied to
the primary purification device, which may be followed by other devices that polish final water quality. The
pre-treatment section commonly includes a sediment filter, cartridge filters capable of retaining particles of various
sizes, a softener, and carbon adsorption beds. The primary purification process most commonly used is reverse
osmosis, which may be followed by deionization and ultrafiltration for polishing the product water from the reverse
osmosis system. Whether a particular device is included in an individual water purification system will be dictated by
local conditions.
5.2.2 Sediment filters
NOTERequirements for sediment filters intended for use in hemodialysis applications can be found in subclause 4.3.8 of
normative reference 2.3.
Permanent, backwashable sediment filters, also known as bed filters, are frequently located at or near the beginning
of hemodialysis water treatment systems and are intended to remove relatively coarse particulate materials from
incoming water. Although a single filtration medium may be used, bed filters known as multimedia filters are more
commonly selected. These units contain multiple layers, each layer retaining progressively smaller particles. In this
way, the bed is used to its fullest extent; the largest particles are removed in the first layer contacted by the water and
the smallest in the final layer.
As the bed accumulates particulate material, open passages begin to clog and resistance to the water flowing through
the filter increases. Ultimately, the increased resistance to flow will lead to a reduction in water supply to downstream
components. To prevent this situation from occurring, bed filters are cleaned by periodic backwashing, which is
accomplished either manually or by using a timer-activated control valve. Bed filters should be fitted with gauges to
measure the hydrostatic pressure at the filters inlet and outlet. These values can be used to determine the dynamic
pressure drop across the filter (delta pressure or P), which serves as an index of resistance to flow and provides a
basis for setting the frequency of backwashing.
5.2.3 Cartridge filters
Cartridge filters consist of a cylindrical cartridge of the filter medium with a central drainage core. The cartridge is
contained within an opaque filter housing with seals to separate the feed and product water streams. Although
cartridge filters may be installed at the inlet to a water system, their usual application is as a final filtration step prior to
reverse osmosis. As the cartridge accumulates particulate material, resistance to flow through the filter increases, as
indicated by an increase in P. When the maximum P recommended by the filter manufacturer is reached, the
cartridge should be replaced according to the manufacturers instructions.
5.2.4 Softeners
NOTERequirements for softeners intended for use in hemodialysis applications can be found in subclause 4.3.10 of normative
reference 2.3.
Water that contains calcium or magnesium can form relatively hard deposits and is called hard water. Water that
has had these elements replaced by sodium ion exchange is called soft water, hence, the term softener. Softeners
also remove other polyvalent cations, most notably iron and manganese, although they are somewhat limited in this
regard. The primary use of softeners in hemodialysis water systems is to prevent hard water deposits from damaging
sensitive reverse osmosis membranes.
A softener is a cylinder or vessel that contains insoluble spheres or beads, called resin, to which sodium ions are
attached. During operation, exchangeable sodium ions in the resin are progressively replaced by calcium and
magnesium ions. When all the sodium ions have been used, the resin bed has reached a condition referred to as
exhaustion. Prior to exhaustion, softeners should be restored; that is, new exchangeable sodium ions are placed on
the resin by a process known as regeneration, which involves exposure of the resin bed to a saturated sodium
chloride solution.
Softeners that automatically regenerate also include a brine tank, from which saturated sodium chloride solution is
drawn during regeneration, and a control valve that regulates regeneration and service cycles.
5.2.5 Carbon adsorption
NOTERequirements for carbon adsorption beds intended for use in hemodialysis applications can be found in subclause 4.3.9 of
normative reference 2.3.
Carbon adsorption systems, often referred to as carbon filters, are the principal means of removing both free chlorine
and chloramine. Removal of free chlorine to a maximum level of 0.5 mg/L and chloramine to a maximum level of 0.1
mg/L is necessary to protect hemodialysis patients from red cell hemolysis. In addition, free chlorine may also
degrade some reverse osmosis membranes, depending on the membrane material.
In addition to removing free chlorine and chloramine, carbon also adsorbs a wide variety of other substances,
including both naturally occurring and synthetic organic compounds. The capacity of carbon to remove free chlorine
and chloramine may be reduced when other substances mask reactive sites on the carbon media. In addition, the
efficiency of free chlorine and chloramine removal is reduced as pH increases or as temperature decreases. The net
effect of those variables is that the finite capacity of carbon beds to remove free chlorine and chloramine cannot be
predicted with any certainty. Therefore, their performance needs to be monitored frequently.
For free chlorine and chloramine removal, carbon adsorption systems and carbon media should be selected and
configured as described in ANSI/AAMI RD62:2001 (see 2.3). That is, two carbon beds shall be installed in series with
a sample port following the first bed. A sample port should also be installed following the second bed for use in the
event of free chlorine or chloramine breaking through the first bed. When samples from the first sampling port are
positive for chlorine or chloramine, operation may be continued for a short time (up to 72 hours) until a replacement
bed is installed, provided that samples from the second sampling port remain negative. The replacement bed should
be placed in the second position, and the existing second bed should be moved to the first position to replace the
exhausted bed. If it is not possible to rotate the position of the beds, both beds should be replaced.
Carbon beds are sometimes arranged as series-connected pairs of beds so that they need not be overly large. The
beds within each pair are of equal size and water flows through them are parallel. In this situation, each pair of beds
should have a minimum empty bed contact time of 5 minutes at the maximum flow rate through the bed. When seriesconnected pairs of beds are used, the piping should be designed to minimize differences in the resistance to flow
from inlet and outlet between each parallel series of beds to ensure that an equal volume of water flows through all
beds.
When granular activated carbon is used as the media, it shall have a minimum iodine number of 900, and each bed
shall have a minimum empty bed contact time of 5 minutes at the maximum flow rate through the bed. Other forms of
carbon should not be used unless there is performance data to demonstrate that each adsorption bed has the
capacity to reduce the chloramine concentration in the feed water to less than 0.1 mg/L when operating at the
maximum anticipated flow rate for the maximum time interval between scheduled testing of the product water for
chloramines. Regenerated carbon shall not be used for hemodialysis applications. Some granular activated carbon
contains aluminum, which can elute from the carbon and add to the burden of aluminum to be removed by reverse
osmosis or ion exchange. The use of acid-washed carbon minimizes this source of aluminum in the water.
In some circumstances, carbon adsorption may not adequately remove chloramines from water. High pH of feed
water, the occurrence of N-chloramines, and the use of orthophosphate or polyphosphate for corrosion control have
been associated with a decrease in the removal of chloramines by carbon adsorption. In those situations, carbon
adsorption may need to be supplemented with other methods of chloramine removal.
Reverse osmosis (RO) systems have become widely used in hemodialysis water purification systems, largely
because these devices remove dissolved inorganic solutes as well as bacteria and bacterial endotoxins.
The RO membrane separation process components are a semipermeable membrane, typically in a spiral-wound
configuration, a pump, and various flow and pressure controls to direct the flow of water through the system. In
operation, feed water is pressurized by the RO pump and is then directed along the surface of the semipermeable
membrane. A portion of the water is forced through the membrane, a process that removes inorganic salts, bacteria,
and bacterial endotoxins. The remainder of the water continues along the membrane surface and is directed to drain.
Water passing through the membrane is referred to as product water or permeate. The water that flows along the
membrane surface and to the drain is known as reject water or concentrate. This flow configuration, known as
cross-flow filtration, prevents a progressive build-up of materials on the membrane surface that would eventually
lead to fouling and membrane failure. In some reverse osmosis systems, a portion of the reject water stream is
recycled to the feed water stream. This recycling allows higher velocities across the membrane surface, which may
help reduce membrane fouling, as well as allowing higher overall use of water. RO systems usually operate in a
single-stage configuration. However, if a higher level of purification is required, a two-stage RO can be used. In a twostage RO, the product water from the first stage acts as the feed water for the second stage.
Depending on membrane configuration and materials of construction, RO systems are sensitive to various feed water
conditions that may lead to diminished performance or premature failure. To avoid such problems, users should
carefully follow the manufacturers instructions for feed water treatment and monitoring to ensure that the RO is
operated within its design parameters.
RO systems should be fitted with a variety of sensors to monitor the systems performance. Conductivity or total
dissolved solids (TDS) sensors in the feed water and product water streams are used to monitor the membranes
ability to remove dissolved inorganic solutes. Flow meters, usually in the product water and reject water streams, are
used to monitor the output of the RO system. RO systems are also fitted with gauges to monitor the pressure at
various points in the system. Although not indicative of treated water quality, monitoring flow rates and pressures can
help ensure that the system is operating within the manufacturers specifications and thus will help ensure RO
reliability.
5.2.8 Deionization
NOTERequirements for deionizers intended for use in hemodialysis applications can be found in subclause 4.3.10 of normative
reference 2.3.
Deionization (DI) is an ion exchange process that removes both anions (negatively charged ions) and cations
(positively charged ions) from water. During the exchange process, hydroxyl ions replace other feed water anions,
and hydrogen ions replace other feed water cations; the hydroxyl and hydrogen ions then combine to form pure water.
Water treated by DI may be very high quality with regard to ionized contaminants, but the process does not remove
nonionized substances, including bacteria and bacterial endotoxins. DI systems may contain anion and cation resin in
separate vessels, known as dual-bed systems, or have may both resin types mixed together in a single vessel,
known as mixed-bed or unibed systems.
10
Systems that include deionizers as a component shall also contain carbon adsorption and ultrafiltration. In such
systems, carbon is placed upstream of the deionizer to avoid formation of carcinogenic nitrosamines, and
ultrafiltration is placed downstream of the deionizer to remove bacteria and endotoxins. The usual application for a
deionizer is as a polisher following reverse osmosis or as a standby process if the reverse osmosis system fails. Use
of deionization as the primary means of purification is not recommended because of the inability of deionization and
ultrafiltration to remove certain low-molecular-weight toxic bacterial products, such as microcystins. In all instances,
deionizers shall be followed by an ultrafilter or other bacteria- and endotoxin-reducing device to remove
microbiological contaminants that may originate in the deionizer resin bed.
DI has a finite capacity that, when exceeded, will cause dangerously high levels of contaminants in the product water.
Fortunately, the quality of product water from DI is easily monitored by resistivity monitors that, when used as
specified in ANSI/AAMI RD62:2001 (see 2.3) (minimum resistivity 1 megohm-cm or greater), can prevent inadvertent
operation of an exhausted deionization system. Specifically, the resistivity monitor following the final deionizer bed
shall be connected to an audible and visible alarm in the dialysis treatment area, and the DI system shall divert
product water to drain or otherwise prevent product water from entering the distribution system should an alarm
condition occur. Under no circumstances shall DI be used when the product water of the final bed has a resistivity
below 1 megohm-cm.
The most common configuration for DI is to have two mixed beds in series, with resistivity monitors being placed
downstream of each bed. Upon exhaustion of the first bed, reliance for water of sufficiently high resistivity shifts to the
second bed, and dialysis operations may be continued for a short time (up to 72 hours) until a replacement bed is
installed.
5.2.9 Ultrafiltration
NOTERequirements for ultrafilters intended for use in hemodialysis applications can be found in subclause 4.3.12 of normative
reference 2.3.
Ultrafilters are membrane-based separation devices that may be used to remove particles as small as 1,000 dalton
and are thus well suited to remove both bacteria and endotoxins. Endotoxin-retentive ultrafilters should be placed in
dialysis water systems at locations downstream of deionization, if deionization is the last process in a water treatment
system (see subclause 4.3.6 of normative reference 2.3) or following ultraviolet irradiation (see subclause 4.3.13 of
normative reference 2.3).
Ultrafiltration membranes used for dialysis applications are typically in either a spiral-wound configuration or in a
hollow-fiber configuration. Spiral-wound ultrafilters are usually operated in a cross-flow mode, with a fraction of the
feed water being forced through the membrane and the remainder being directed along the membrane surface to
drain. As with reverse osmosis, cross-flow filtration is intended to minimize membrane fouling. Hollow-fiber ultrafilters
are typically housed in vessels similar to those used for cartridge sediment filters and these are operated in a deadend (no cross-flow) mode.
Ultrafilters should be fitted with pressure gauges on the inlet and outlet water lines to monitor the pressure drop (P)
across the membrane. Such monitoring will indicate when membrane fouling has progressed to the point at which
membrane replacement or cleaning is needed. Monitoring also ensures that the device is being operated in
accordance with the manufacturers instructions. Ultrafilters should be included in routine disinfection procedures to
prevent uncontrolled proliferation of bacteria in the feed water compartment of the filter. If bacterial proliferation is not
controlled, bacteria may grow through the membrane and contaminate the product water compartment of the filter.
Ultrafilters operated in the cross-flow mode should also be fitted with a flow meter to monitor the flow rate of water
being directed to drain.
5.3
5.3.1 General
The function of the water storage and distribution system is to distribute product water from the purification cascade to
its points of use, including individual hemodialysis machines, hemodialyzer reprocessing equipment, and concentrate
preparation systems. A water storage and distribution system typically contains a large volume of water exposed to a
large surface area of piping and storage tank walls. Because chlorine and chloramines are removed in the purification
process, the water does not contain a bacteriostatic agent. This combination of circumstances predisposes wetted
surfaces to bacterial proliferation and biofilm formation. Therefore, a water storage and distribution system should be
designed specifically to facilitate bacterial control, including measures to prevent bacterial colonization and to allow for
easy and frequent disinfection.
5.3.2 Water storage
When used, storage tanks should have a conical or bowl-shaped base and should drain from the lowest point of the
base. Storage tanks should have a tight-fitting lid and be vented through a hydrophobic 0.2 m air filter. The filter
11
should be changed on a regular schedule according to the manufacturers instructions. A means shall be provided to
effectively disinfect any storage tank installed in a water distribution system. Internal spray mechanisms can facilitate
effective disinfection and rinsing of a storage tank.
5.3.3 Water distribution systems
Two types of water distribution systems are used: direct feed systems and indirect feed systems. In a direct feed
system, water flows directly from the last stage of the purification cascade to the points of use. In an indirect feed
system, water flows from the end of the purification cascade to a storage tank. From there, it is distributed to the
points of use. In general, direct feed systems offer the least favorable environment for bacterial proliferation.
However, with a direct feed system the purification cascade must be sized to provide sufficient water to meet the peak
demand, and the system must have sufficient pressure at the end of the purification cascade to distribute the water to
the points of use. Those two requirements often preclude the use of a direct feed system. Whichever type of system
is used, water distribution systems should be configured as a continuous loop and designed to minimize bacterial
proliferation and biofilm formation (see clause 7). A centrifugal pump made of inert materials is necessary to distribute
the purified water and aid in effective disinfection. A multistage centrifugal pump is preferred for this purpose.
Product water distribution systems shall be constructed of materials that do not contribute chemicals, such as
aluminum, copper, lead, and zinc, or bacterial contaminants to the purified water. The choice of materials used for a
water distribution system will also depend on the proposed method of disinfection. Table 2 provides some guidance
on the compatibility of different materials and disinfection agents. Whatever material is used, care should be taken to
select a product with properties that provide the least favorable environment for bacterial proliferation, such as
smooth internal surfaces.
Bleach
Peracetic acid
Formaldehyde
PVC
CPVC
PVDF
PEX
X
X
SS
PP
PE
ABS
Hot water
Ozone
X
X
PTFE
Glass
PVC = polyvinylchloride, CPVC = chlorinated polyvinylchloride, PVDF = polyvinylidene fluoride, PEX = cross-linked polyethylene,
SS = stainless steel, PP = polypropylene, PE = polyethylene, ABS = acrylonitrile butadiene styrene,
PTFE = polytetrafluoroethylene.
NOTETable 2 is not intended as an exhaustive compilation of all possible compatible combinations of piping material and
disinfectant. Users should verify compatibility between a given germicide and the materials of a piping system with the supplier of
that piping system before using the germicide. Considerations of compatibility should include any joint materials and pipe fittings, as
well as the actual piping material. The concentration of germicide and the duration and frequency of exposure also should be taken
into account.
12
control. UV irradiators also may be used for other applications in a water purification and distribution system (see 5.3.4.2 and
A.5.2.5).
Ultraviolet irradiators (also known as UV lights) may be used to control bacterial proliferation in purified water storage
and distribution systems. UV irradiators contain a low-pressure mercury lamp that emits ultraviolet light at a
wavelength of 254 nm. The lamp is housed in a transparent quartz sleeve that isolates it from direct contact with the
water. If the irradiator is not fitted with a calibrated ultraviolet intensity meter that is filtered to restrict its sensitivity to
the disinfection spectrum and that is installed in the wall of the disinfection chamber at the point of greatest water
depth from the lamp, the dose of radiant energy provided by the lamp shall be at least 30 milliwatt-sec/cm2. If the
irradiator includes a meter as described above, the minimum dose of radiant energy should be at least 16 milliwattsec/cm2.
Ultraviolet irradiation also can be used to control bacteria in the pretreatment section of a water purification system,
such as following carbon adsorption beds to reduce the bacterial burden presented to a reverse osmosis unit.
Because using UV irradiation to kill bacteria increases the level of endotoxins in the water, UV irradiators should be
followed by a means of reducing endotoxin concentrations, such as an ultrafilter in the purified water distribution
system or reverse osmosis in the pretreatment cascade.
To prevent the use of sublethal doses of radiation that may lead to the development of resistant strains of bacteria,
UV irradiators shall be equipped with a calibrated ultraviolet intensity meter, as described above, or with an on-line
monitor of radiant energy output that activates a visible alarm, which indicates that the lamp should be replaced.
Alternatively, the lamp should be replaced on a predetermined schedule according to the manufacturers instructions
to maintain the recommended radiant energy output.
5.3.4.2 Ozone generators
NOTERequirements for ozone generators intended for use in hemodialysis applications can be found in subclause 4.3.15 of
normative reference 2.3.
Ozone may be used to control bacterial proliferation in water storage and distribution systems. Ozone may also
degrade endotoxins. Ozone generators convert oxygen in air to ozone using a corona discharge or ultraviolet
irradiation. The ozonated air is then injected into the water stream. An ozone concentration of 0.2 mg/L to 0.5 mg/L,
combined with a contact time of 10 minutes, is capable of killing bacteria, bacterial spores, and viruses in water.
Destruction of established biofilm may require longer exposure times and/or higher concentrations of ozone.
Ozone may degrade many plastic materials, including PVC and elastomeric O-rings and seals. Therefore, ozone can
be used for bacterial control only in systems constructed from ozone-resistant materials (see 5.3.3 for suitable piping
materials).
5.3.4.3 Hot water disinfection systems
NOTERequirements for hot water disinfection systems intended for use in hemodialysis applications can be found in subclause
4.3.14 of normative reference 2.3.
Hot water ( 80 C) may be used to control bacterial proliferation in water storage and distribution systems. The
manufacturers instructions for using hot water disinfection systems should be followed. If no manufacturers
instructions are available, the effectiveness of the system can be demonstrated by verifying that the system maintains
a specified temperature for a specified time and by performing ongoing surveillance with bacterial cultures and
endotoxin testing. Bacterial kill studies are not required.
Hot water disinfection systems can be used only in systems constructed from heat-resistant materials, such as crosslinked polyethylene, polypropylene, and stainless steel (see 5.3.3).
5.4
Concentrate preparation
5.4.1 General
Dialysate is customarily prepared from two concentrates: the bicarbonate concentrate, which contains sodium
bicarbonate (and sometimes additional sodium chloride), and the acid concentrate, which contains all remaining ions,
acetic acid, and sometimes glucose. Dialysate also can be prepared from a single concentrate that contains acetate.
The buffer is provided to the patient in the form of acetate, which is subsequently metabolized to yield bicarbonate.
Acetate-containing dialysate is now rarely used in clinical practice. In general, acetate-containing concentrate is
handled in a similar manner to that of acid concentrate (see 5.5.3 and 5.5.5), except that acetate-containing
concentrate systems are color-coded white.
Acid concentrate can be supplied by the manufacturer in bulk (usually in 55 gallons drums) or in gallon containers. In
some cases, the manufacturer will pump the acid concentrate from the 55 gallon drums into a holding tank at the
13
dialysis facility. Systems have recently been introduced that allow a user at a dialysis facility to prepare acid
concentrate from packaged powder and purified water using a mixer. Acid concentrates supplied in 55 gallon drums
or gallon containers by the manufacturer are the responsibility of that manufacturer (see 2.4). If the acid concentrate
is pumped into a bulk storage tank at the dialysis facility, the user is responsible for maintaining the concentrate in its
original state and to ensure that the correct formula is used according to the patients prescription. Acid concentrate
prepared at the dialysis facility from powder and water is also the responsibility of the user.
Bicarbonate concentrate can be supplied by the manufacturer in one of three ways: (1) in gallon containers, (2) as
packaged powder that is mixed with purified water at the dialysis facility, and (3) in powder cartridges that are used to
prepare concentrate on-line at the time of dialysis.
5.4.2 Materials compatibility
All components used in concentrate preparation systems (including mixing and storage tanks, pumps, valves, and
piping) shall be fabricated from materials (e.g., plastics or appropriate stainless steel) that do not interact chemically
or physically with the concentrate so as to affect its purity, or with the germicides or germicidal procedure used to
disinfect the equipment. The use of materials that are known to cause toxicity in hemodialysis, such as copper, brass,
galvanized material, and aluminum, are specifically prohibited.
5.4.3 Bulk storage tanks (acid concentrate)
Procedures should be in place to control the transfer of the acid concentrate from the delivery container to the storage
tank to prevent the inadvertent mixing of different concentrate formulations. If possible, the tank and associated
plumbing should form an integral system to prevent contamination of the acid concentrate. The storage tanks and
inlet and outlet connections, if remote from the tank, should be secure and labeled clearly.
5.4.4 Mixing systems
5.4.4.1 General
Concentrate mixing systems require a purified water source (see 2.3), a suitable drain, and a ground fault protected
electrical outlet. Protective measures should be used to ensure a safe work environment. For example, ventilation
and personal protective equipment should be used to handle any residual dust that is introduced into the atmosphere
as powdered concentrates are added to the system and to handle any additional heat produced by the device.
Structural issues, such as the facilitys weight-bearing capacity, should be addressed if systems are to be installed
above ground level. Operators should at all times use appropriate personal protective equipment, such as face
shields, masks, gloves, gowns, and shoe protectors, as recommended by the manufacturer.
If a concentrate mixing system is used, the preparer should follow the manufacturers instructions for mixing the
powder with the correct amount of water. The number of bags or the weight of powder added should be determined
and recorded.
Labeling strategies should permit positive identification by anyone using the contents of mixing tanks, bulk
storage/dispensing tanks, and small containers intended for use with a single hemodialysis machine. Requirements
for such positive identification will vary among facilities, depending on the differences between concentrate
formulations used and on whether single or multiple dialysate proportioning ratios are used. (The use of multiple
dialysate proportioning ratios in a single facility is strongly discouraged.) Although it is the responsibility of facilities to
develop and use labeling to positively identify the contents of mixing tanks, bulk storage/dispensing tanks, and
concentrate jugs, the following guidelines are suggested.
Mixing tanks: Prior to batch preparation, a label should be affixed to the mixing tank that includes the date of
preparation and the chemical composition or formulation of the concentrate being prepared. This labeling should
remain on the mixing tank until the tank has been emptied. Using a photocopy of the concentrate manufacturers
package label provides a convenient and comprehensive means of identifying the chemical composition or
formulation of the concentrate; however, the lot number and expiration date should be marked out because they apply
only to the dry powder.
Bulk storage/dispensing tanks: These tanks should be permanently labeled to identify the chemical composition or
formulation of their contents. As with mixing tanks, bulk storage/dispensing tank labeling can be conveniently
accomplished by affixing a copy of the concentrate manufacturers package label.
Concentrate jugs: Concentrate jugs are typically nondisposable vessels provided by hemodialysis machine
manufacturers and having a capacity sufficient for one or two hemodialysis sessions. The extent of labeling for these
containers depends on the variety of concentrate formulations used and on whether the facility uses dialysis
machines with different proportioning ratios (a practice that is strongly discouraged). At a minimum, concentrate jugs
should be labeled with sufficient information to differentiate the contents from other concentrate formulations used at
the facility. If a chemical spike is added to an individual container to increase the concentration of an electrolyte, the
14
label should show the added electrolyte, the date and time added, and the name of the person making the addition
(see 5.4.5). The additional information may be simple or extensive, but in all cases it should permit users to positively
identify the containers contents.
In addition to container labeling, there should be permanent records of batches produced. These records should
include the concentrate formula produced, the volume of the batch, the lot numbers of powdered concentrate
packages, the manufacturer of the powdered concentrate, the date and time of mixing, any test results, the person
performing the mixing, the person verifying mixing and test results, and the expiration date (if applicable).
Manufacturers recommendations should be followed regarding any preventive maintenance and sanitization
procedures. Records should be maintained indicating the date, time, person performing the procedure, and results
(if applicable).
5.4.4.2 Acid concentrate mixing systems
Acid concentrate mixing tanks should be designed to allow the inside of the tank to be completely emptied and rinsed
according to the manufacturers instructions when concentrate formulas are changed. Use of a tank with a sloping
bottom that drains from the lowest point is one means of facilitating this process. Because concentrate solutions are
highly corrosive, mixing systems should be designed and maintained to prevent corrosion. Acid concentrate mixing
tanks should be emptied completely before mixing another batch of concentrate.
5.4.4.3 Bicarbonate concentrate mixing systems
Bicarbonate concentrate mixing tanks should be designed to drain completely; for example, they should have a
sloping bottom and a drain at the lowest point. High- and low-level alarms can prevent overfilling and air damage to
the pump. Because concentrate solutions are highly corrosive, mixing systems should be designed and maintained to
prevent corrosion. Mixing tanks should have a tight-fitting lid and should be designed to allow all internal surfaces to
be disinfected and rinsed. A translucent tank allows users to see the liquid level; the use of sight tubes is not
recommended because of the potential for microbial growth, such as bacteria, algae, and fungi.
Once mixed, bicarbonate concentrate should be used within the time specified by the manufacturer of the
concentrate. The concentrate shall be shown to routinely produce dialysate meeting the recommendations of 4.3.2.1.
Overagitating or overmixing of bicarbonate concentrate should be avoided, as this can cause CO2 loss and can
increase pH. (Systems designed for mixing dry acid concentrates may use methods that are too vigorous for
dissolving dry bicarbonate.) The mixing tank should be either (1) completely emptied and disinfected according to the
manufacturers instructions, or (2) disinfected using a procedure demonstrated by the facility to be effective in
routinely producing concentrate that allows the recommendations of 4.3.2.1 to be met.
5.4.5 Additives
Manufacturers provide acid concentrates with a wide range of electrolyte compositions for different proportioning
ratios. Most typical dialysate prescriptions can be obtained by using one or more of these commercially available
concentrates. If particular formulations are not available, manufacturers provide additives that can be used to adjust
the level of potassium or calcium in the dialysate. These additives are commonly referred to as spikes.
Concentrate additives should be mixed with liquid acid concentrates according to the manufacturers instructions,
taking care to ensure that the additive is formulated for use in concentrates of the appropriate dilution ratio. When
liquid additives are used, the volume contributed by the additive should be considered when calculating the effect of
dilution on the concentration of the other components in the resulting concentrate. When powder additives are used,
care should be taken to ensure that the additive is completely dissolved and mixed before the concentrate is used.
Containers should be labeled to indicate the final concentration of the added electrolyte, the date and time mixed, and
the person mixing the concentrate. This information should also be recorded in a permanent record. Labels should be
affixed to the containers when the mixing process begins.
5.5
Concentrate distribution
15
16
minimizing the risk that the wrong concentrate will be connected to an outlet. The dispensing outlets should be
labeled with the appropriate symbol (see Table 3) indicating the proportioning ratio for the dialysis machine and
should be color-coded blue for bicarbonate, red for acid.
5.6
Dialysate proportioning
Essentially, all dialysate is produced with three fluid streams: water, acid concentrate, and bicarbonate concentrate.
This three-stream combination produces a highly buffered dialysate with a pH between 6.9 and 7.6. Dialysate can
also be prepared from a single concentrate that contains acetate to provide a dialysate in which buffer is provided to
the patient in the form of acetate, which is subsequently metabolized to yield bicarbonate. However, acetatecontaining dialysate is now rarely used in clinical practice.
Different manufacturers of dialysis machines use different methods of controlling the proportions of the concentrates.
These can be generally grouped into two categories: fixed proportioning and servo control. With both methods, the
operator can select a desired sodium and bicarbonate level, and the machine will make the necessary adjustments to
achieve the selected levels. Both types use a redundant system of controls and monitoring. With fixed proportioning
systems, the pumps are set to established volumes, and the final conductivity is verified. With servo control machines,
the individual concentrates are added until the conductivity achieves the expected value. A final redundant
conductivity monitor verifies the mixture. Some machines may also monitor the pH of the dialysate as an additional
safeguard against gross errors in dialysate formulation. A different type of machine with a batch tank and dedicated
concentrates is also available.
It is important that the acid and bicarbonate concentrates be matched with respect to the proportioning ratio and with
the model and setup configuration of the dialysis machine. Several types of three-stream concentrates are available,
with different ratios of acid concentrate to bicarbonate concentrate to water (see Table 3). The different proportioning
types are not compatible with one another. Generally, bicarbonate is available in one or two forms for each
proportioning type (in liquid, cartridge, or dry powder, and in various sizes). Each proportioning type has numerous
acid concentrate formulations (codes) with different amounts of potassium, calcium, and magnesium ions, plus
dextrose. To help differentiate between concentrates of different proportioning types, AAMI recommends that the
manufacturer include a geometric symbol on the labels along with acid/base color coding.
Table 3Symbols and color coding for different concentrate proportioning ratios
Concentrate
type
Geometric
symbol
Bicarbonate
concentrate
(Blue color coding)
35X
1:34
Square
36.83X
1:35.83
Circle
Dry or liquid
45X
1:44
Triangle
36.1X
1:35.1
Hexagon
Cartridge
Comments
Bicarbonate concentrate
contains some NaCl.
NOTE 1The acid proportioning ratio refers to acid concentrate:water + bicarbonate concentrate.
NOTE 2Acetate-containing concentrate is color-coded white.
Some models of dialysis machines can use concentrates of only one type of proportioning ratio, but others may be set
up or calibrated for use with concentrates of more than one proportioning type. (Note that changing from one
proportioning ratio to another requires recalibration for some models of dialysis machines.) Thus, for those machines,
the type of concentrate should be labeled on the machine or clearly indicated by the machine display. It is strongly
recommended that facilities configure every machine to use only one type of concentrate.
Injuries related to improper dialysate are rare, but they can and do happen when all procedures are not followed.
Frequently, when the error occurs, several patients have been exposed before the facility recognizes the mistake. For
example, because one of the concentrates is quite acidic and the other is basic, connecting the wrong concentrates to
the machine could result in dialysate that could harm the patient. Thus, it is necessary for the operator to follow the
manufacturers instructions regarding dialysate conductivity and to measure approximate pH with an independent
method before starting the treatment of the next patient.
17
Even though a single concentrate is used to prepare acetate dialysate, conductivity and pH should be checked,
because certain mix-ups involving acid concentrate and other chemicals can result in an acceptable conductivity and
very low pH.
Monitoring
6.1
General
Quality control and quality assurance procedures should be established to ensure ongoing conformance to policies
and procedures regarding dialysate quality. This clause defines some of the monitoring activities to be conducted at
the dialysis facility as part of the quality assurance process. The paragraph numbers in 6.2 and 6.3 correspond to the
paragraph numbers in 5.2 and 5.3. The test methods described in 6.2 do not represent the only acceptable methods
available, but are intended to provide examples of acceptable methods. The frequency of monitoring is generally
recommended by the equipment manufacturer. Table 4 can be used as a guideline for setting up a quality assurance
monitoring program in the absence of a manufacturers recommendations or to supplement those recommendations.
Item to
monitor
What to monitor
Special
interval
Normal interval
Specification
Sediment filter
NA
Daily
Sediment filter
backwashing
cycle
NA
Dailybeginning
of the day
Cartridge filter
NA
Daily
Water softener
Product water
softness
NA
Dailyend of the
day
Water softener
brine tank
Level of undissolved
salt in tank
NA
Dailyend of the
day
Water softener
regeneration
cycle
Regeneration cycle
timer setting
NA
Dailybeginning
of the day
Carbon
adsorption
beds
NA
Prior to beginning
each patient shift
Chemical
injection
system
Level of chemical in
the reservoir, injector
function, value of the
controlling parameter
(e.g., pH)
NA
Daily
Reverse
osmosis
Product water
conductivity, total
dissolved solids
(TDS), or resistivity
and calculated
rejection
NA
According to the
manufacturers
recommendations
(continuous
monitors)
Rejection XX%
18
Table 4 (continued)
Item to
monitor
What to monitor
Special
interval
Normal interval
Specification
Reverse
osmosis
NA
Daily (continuous
monitors)
Deionizers
Product water
resistivity
NA
Continuous
Ultrafilters
NA
Daily
Water storage
tanks
Weekly, until a
pattern of
consistent
compliance with
limits can be
demonstrated
NA
Water
distribution
piping system
Weekly, until a
pattern of
consistent
compliance with
limits can be
demonstrated
Monthly
UV light
sources
Energy output
NA
Monthly
Ozone
generators
Concentration in the
water
NA
During each
disinfection
Hot water
disinfection
systems
Temperature and
time of exposure of
the system to hot
water
NA
During each
disinfection
Dialysate
NA
Monthly, rotated
among machines
so that at least
two machines are
tested each month
and so that each
machine is tested
at least once per
year
Dialysate
Conductivity and pH
NA
Each treatment
NOTEIt is not possible to specify universally acceptable operating ranges for each device listed in the table, since some of
these values will be system-specific. In those cases (denoted by Xs in the Specification column of the table), the facility should
define an acceptable operating range based on manufacturers instructions or measurements of system performance.
6.2
Water purification
6.2.1 General
The recommendations below apply.
19
20
Newer RO systems may have a direct reading for percent rejection. Other parameters that should be measured daily
include product and reject stream flow rates and various internal pressures to the extent permitted by RO
instrumentation. Although these parameters are not directly indicative of treated water quality, monitoring them can
help ensure that the system is operating within the manufacturers specifications and thus will aid in maintaining the
performance of the RO membranes. Flow rates can be used to calculate the percent recovery of the RO using the
following formula:
Recovery (%) =
NOTEThe percent recovery is also known as the water conversion factor. The terms are equivalent if none of the reject water
stream is recycled to the feed water stream (see 5.2.7). If some of the reject water stream is recycled, the equation given above
provides a measure of overall water utilization by the reverse osmosis system, rather than the recovery of water during a single pass
through the membrane module.
RO systems are not equipped with instrumentation that permits measurement of product water levels of bacteria,
endotoxins, or the individual contaminants listed in 4.1. Chemical analysis for the contaminants listed in 4.1.1
(Table 1) should be done when the RO system is installed, when membranes are replaced, and at not less than
annual intervals thereafter to ensure that the limits specified in 4.1.1 are met (see Table 1). It is further recommended
that chemical analyses be done when seasonal variations in source water suggest worsening quality or when
rejection rates fall below 90 %.
All results of measurements of RO performance should be recorded daily in an operating log that permits trending
and historical review.
6.2.8 Deionization
Deionizers shall be monitored continuously using resistivity monitors that compensate for temperature and are
equipped with audible and visual alarms. Resistivity monitors shall have a minimum sensitivity of 1.0 megohm-cm.
Patients shall not be dialyzed on deionized water with resistivity less than 1.0 megohm-cm measured at the output of
the deionizer. When deionization is employed as the primary method for removing inorganic contaminants (reverse
osmosis is not employed), or when deionization is necessary to polish RO-treated water, chemical analyses to ensure
that the requirements of 4.1.1 (Table 1) are met should be performed when the system is installed and at annual
intervals thereafter. Resistivity monitor readings should be recorded on a log sheet twice each treatment day.
21
6.2.9 Ultrafiltration
The performance of ultrafilters can be monitored by testing the water that is directly exiting the ultrafilter for bacteria
and endotoxins. Bacteria and endotoxin levels should be measured as specified in ANSI/AAMI RD62:2001 (see 2.3).
The pressure drop across the ultrafilter (P) should be measured using simple inlet and outlet pressure gauges. Such
monitoring will indicate when membrane fouling has progressed to the point that membrane replacement or cleaning
is needed. Monitoring is also necessary to ensure that the device is being operated in accordance with the
manufacturers instructions. Ultrafilters operated in the cross-flow mode should also be monitored in terms of the flow
rate of water being directed to drain (concentrate). Results of pressure measurements and bacteria and endotoxin
levels should be recorded in a log.
6.3
6.3.1 General
The recommendations below apply.
6.3.2 Water storage
Routine monitoring of water storage tanks for bacteria and endotoxin levels is generally accomplished indirectly by
monitoring the water at the first outlet to the distribution loop (see 6.3.3). If direct monitoring of a water storage tank is
performed as part of a troubleshooting process, bacteria and endotoxin levels shall be measured as specified in
ANSI/AAMI RD62:2001 (see 2.3). All bacteria and endotoxin results should be recorded on a log sheet.
6.3.3 Water distribution systems
Water distribution piping systems should be monitored for bacteria and endotoxin levels. Bacteria and endotoxins
shall not exceed the levels specified in 4.1.2. Monitoring should be accomplished by taking samples from the first and
last outlets of the water distribution loop and the outlets supplying reuse equipment and bicarbonate concentrate
mixing tanks. If the results of this testing are unsatisfactory, additional testing (e.g., ultrafilter inlet and outlet, RO
product water, and storage tank outlet) should be undertaken as a troubleshooting strategy to identify the source of
contamination, after which appropriate corrective actions can be taken. Bacteria and endotoxin levels shall be
measured as specified in ANSI/AAMI RD62:2001 (see 2.3). Bacteria and endotoxin testing should be conducted at
least monthly. For a newly-installed water distribution piping system, or when a change has been made to an existing
system, it is recommended that weekly testing be conducted for 1 month to verify that bacteria or endotoxin levels are
consistently within the allowed limits. All bacteria and endotoxin results should be recorded on a log sheet to identify
trends that may indicate the need for corrective action.
6.3.4 Bacterial control devices
6.3.4.1 Ultraviolet irradiators
Ultraviolet irradiators intended for use as a direct means of bacterial control shall be monitored for radiant energy
output. UV irradiators are available equipped with radiant energy intensity sensors. A visual alarm or an output meter
is acceptable for determining if the UV lamp is emitting sufficient radiant energy. UV irradiators should be monitored
at the frequency recommended by the manufacturer. Because the radiant energy decreases with time, annual lamp
replacement is typically required. Periodic cleaning of the quartz sleeve may also be required, depending on the water
quality. A log sheet should be used to indicate that monitoring has been performed.
6.3.4.2 Ozone generators
Ozone generators should be monitored for ozone output at a level specified by the manufacturer. The output of the
ozone generator should be measured by the ozone concentration in the water. A test based on indigo trisulfonate
chemistry, or the equivalent, should be used to measure the ozone concentration. It is recommended that ozone
concentration be measured each time disinfection is performed. An ozone-in-ambient-air test should be conducted on
a periodic basis, as recommended by the manufacturer, to ensure compliance with the OSHA permissible exposure
limit of 0.1 ppm. A log sheet should be used to indicate that monitoring has been performed.
6.3.4.3 Hot water disinfection systems
Hot water disinfection systems should be monitored for temperature and time of exposure to hot water as specified by
the manufacturer. Also, hot water disinfection should be performed at least as often as recommended by the
manufacturer. The temperature of the water should be recorded at a point farthest from the water heaterthat is,
where the lowest water temperature is likely to occur. It is recommended that the water temperature be measured
each time a disinfection cycle is performed. A record that verifies successful completion of the heat disinfection
should be maintained. Successful completion is defined as meeting temperature and time requirements specified by
the equipment manufacturer.
22
6.4
Concentrate preparation
Concentrate distribution
A daily check to ensure that the appropriate acid and bicarbonate concentrate is connected to the corresponding
concentrate delivery line is recommended if the storage tank is not permanently connected to its distribution piping.
Piped bicarbonate concentrate distribution systems should be disinfected either according to the manufacturers
instructions or using a procedure that has been demonstrated by the facility to be effective in routinely producing
concentrate and that allows the recommendations of 4.3.2.1 to be met. It is recommended that the interval between
disinfections not exceed 1 week. If the manufacturer does not supply disinfection procedures, the user should develop
and validate a disinfection protocol. It is recommended that monitoring of concentrate distribution systems be
performed on a routine basis. When reusable concentrate jugs are used to distribute bicarbonate concentrate, they
should be disinfected at least weekly. Bicarbonate concentrate jugs should be rinsed with treated water and stored
inverted at the end of each treatment day. Pick-up tubes should also be rinsed with treated water and allowed to air
dry at the end of each treatment day.
Once a bicarbonate distribution system has been activated, dialysate should be monitored weekly until sufficient data
has been obtained to demonstrate consistent compliance with acceptable levels of contamination. The frequency of
monitoring may then be reduced, but monitoring should be performed at least monthly. If elevated bacteria or
endotoxin levels are found in the dialysate, all systems involved in dialysate preparation, including the bicarbonate
concentrate distribution system should be evaluated and appropriate action, such as disinfection, should be taken.
The frequency of monitoring should then be increased until it can be demonstrated that the problem has been
resolved.
Because acid concentrate distribution systems have been shown not to be subject to bacterial proliferation, it is not
necessary to perform bacteria and endotoxin testing on those systems.
6.6
Dialysate proportioning
Dialysate proportioning should be monitored following the procedures specified by the equipment manufacturer. The
user should maintain a record of critical parameters such as conductivity and approximate pH. When the user has
specific requirements for monitoring dialysate proportioning, such as when dialysis machine settings are changed to
allow the use of concentrates with a different proportioning ratio, the user should develop procedures for routine
monitoring of dialysate electrolyte values.
23
7.1
General
The strategy for controlling the proliferation of microorganisms in hemodialysis systems primarily involves proper
system design and operation, and regular disinfection of water treatment system and hemodialysis machines. A key
concept in ensuring compliance with the requirements of 4.1.2 is that disinfection schedules should be designed to
prevent bacterial proliferation, rather than being designed to eliminate bacteria once they have proliferated to an
unacceptable level (i.e., above the action level). With this strategy, monitoring levels of bacteria and endotoxin serves
to demonstrate that the disinfection program is effective, not to indicate when disinfection should be performed.
Gram-negative water bacteria, their associated lipopolysaccharides (bacterial endotoxins), and nontuberculous
mycobacteria (NTM) most frequently come from the community water supply, and levels of those bacteria can be
amplified depending on the water treatment system, dialysate distribution system, type of dialysis machine, and
method of disinfection.
Two components of hemodialysis water distribution systemspipes and storage tankscan serve as reservoirs of
microbial contamination. Hemodialysis systems frequently use pipes that are of larger diameter and longer than are
needed to handle the required flow. Oversized piping slows the fluid velocity and increases both the total fluid volume
and the wetted surface area of the system. Gram-negative bacteria in fluids remaining in pipes overnight multiply
rapidly and colonize the wet surfaces, thus producing bacterial populations and endotoxin quantities in proportion to
the volume and surface area. Such colonization results in the formation of protective biofilm that is difficult to remove
once formed and that provides a barrier between the bacteria and germicide during disinfection.
Biofilms are communities of microorganisms attached to surfaces. They form just about anywhere a nonsterile fluid
flows over a surface. Biofilm increases the ability of microorganisms to compete for nutrients and other resources.
The complexity of biofilm depends on the degree of water or fluid movement and the availability of nutrients. Thicker
biofilm, and usually a greater diversity of microorganisms, will form in slower moving waters; in faster moving waters,
it is harder for microorganisms to become (and remain) attached to the surface, so biofilm formation takes longer.
Organisms living within biofilm are shielded by an extracellular polymer or glycocalyx. This glycocalyx provides the
bacteria with some protection from the action of disinfectants. Biofilm may exist throughout a hemodialysis distribution
system. Once established in a distribution system or dialysis machine, biofilm can be difficult to eradicate. Bleach and
ozone are generally the most effective agents for biofilm removal, and their use may be more efficacious if the pipes
are treated first with a descaling agent. However, in some cases, complete or partial replacement of the distribution
system may be the only way to eliminate biofilm. Routine low-level disinfection of the pipes should be performed to
control bacterial contamination of the distribution system. The frequency of disinfection will vary with the design of the
system and the extent to which biofilm has already formed in existing systems, but disinfection should be performed
at least monthly.
To minimize biofilm formation, there should always be flow in a piping system. A minimum velocity of 3 ft/sec in the
distal portion of the loop of an indirect feed system and a minimum velocity of 1.5 ft/s in the distal portion of a direct
feed system are recommended when the system is operating under conditions of peak demand. Other measures can
also help protect pipes from contamination. A mechanism should be incorporated in the distribution system to ensure
that disinfectant does not drain from pipes during the disinfection period. Dead-end pipes and unused branches and
taps that can trap fluid should be eliminated because they act as reservoirs of bacteria and are capable of
continuously inoculating the entire volume of the system. Joints between sections of piping and between piping and
fittings should be formed in a manner that minimizes the formation of crevices and other voids that may serve as sites
for bacterial colonization. Pipes should not be cut with a hacksaw. Any burrs should be removed before the joint is
formed. These measures also minimize the possibility that pockets of residual disinfectant could remain in the piping
system after disinfection.
A storage tank in the distribution system greatly increases the volume of fluid and surface area available and can
serve as a niche for water bacteria. Storage tanks are therefore not recommended for use in dialysis systems unless
they are frequently drained and adequately disinfected. It may be necessary for the user to scrub the sides of the tank
to remove bacterial biofilm if the tank design and maintenance are not adequate to prevent bacterial proliferation. An
ultrafilter, distal to the storage tank, or some other form of bacterial control device is recommended.
For most dialysis machines, routine disinfection with hot water or with a chemical germicide connected to a
disinfection port on the machine does not disinfect the line between the outlet from the water distribution system and
the back of the dialysis machine. Users should establish a procedure for regular disinfection of this line. One
approach is to rinse the dialysis machines with water containing germicide or hot water when the water distribution
loop is disinfected. If this procedure is used with a chemical germicide, each dialysis machine should be rinsed and
tested for the absence of residual germicide following disinfection.
Storage times for bicarbonate concentrate should be minimized, as well as the mixing of fresh bicarbonate
concentrate with unused portions of concentrate from a previous batch. The manufacturers instructions should be
followed if they are available. Facilities that reuse concentrate jugs for bicarbonate concentrate should disinfect the
24
jugs at least weekly. Bicarbonate concentrate can support prolific growth of microorganisms. Jugs can be disinfected
with household bleach solutions (300 mg to 600 mg free chlorine, or 30 mL to 60 mL of 6.15 % household bleach per
gallon of water) with a contact time of about 30 minutes or another EPA-registered disinfectant according to the
manufacturers instructions. Following disinfection, jugs should be drained, rinsed, and inverted to dry.
7.2
7.2.1 General
The microbial quality of water should be monitored at least monthly to validate the effectiveness of the disinfection
program. Monitoring can be accomplished by direct plate counts, in conjunction with the measurement of bacterial
endotoxin.
Samples of water should be collected from several places to give an indication of the microbial quality of the water
throughout the water distribution system. In general, samples should be collected in the following areas: from the first
and last outlets of the water distribution loop, where water enters equipment used to reprocess dialyzers, and where
water enters equipment used to prepare bicarbonate concentrate or from the bicarbonate concentrate mixing tank.
Additional testing, such as at the end of the water purification cascade and at the outlet of the storage tank, if one is
used, may be necessary during initial qualification of a system or when troubleshooting the cause of contamination
within the distribution loop.
Dialysate samples should also be collected from at least two machines monthly and from enough machines so that
each machine is tested at least once per year. If testing of any dialysis machine reveals a level of contamination
above the action level, an investigation should be conducted that includes retesting the offending machine, reviewing
compliance with disinfection and sampling procedures, and evaluating microbiological data for the previous 3 months
to look for trends. The medical director also should be notified. An example of a decision tree for this process is given
in Figure 1.
Cultures should be repeated when bacterial counts exceed the allowable levels. If culture growth exceeds permissible
standards, the water system and dialysis machines should be cultured weekly until acceptable results are obtained.
Additional samples should be collected when there is a clinical indication of a pyrogenic reaction or septicemia, and
following a specific request by the clinician or the infection control practitioner.
Samples should always be collected before sanitization/disinfection of the water treatment system and dialysis
machines. If repeat cultures are performed after the system has been disinfected (e.g., with formaldehyde, hydrogen
peroxide, chlorine, or peracetic acid), the system should be flushed completely before collecting samples. Drain and
flush storage tanks and the distribution system until residual disinfectant is no longer detected before collecting
samples. Culture water and dialysis fluid weekly for new systems until a pattern has been established. For established
systems, culture monthly unless a greater frequency is dictated by historical data at a given institution. If bacterial
contamination is suspected, but water cultures are negative, it may be necessary to check for the presence of biofilm
(see 7.2.3).
25
Results < 50
CFU/mL?
Results 50199
CFU/mL?
Results 200
CFU/mL?
Disinfect equipment or
water system if
necessary
No
Disinfect equipment or
water system if
necessary
Redraw sample
Redraw sample
Yes
Results < 50
CFU/mL?
Yes
Results < 50
CFU/mL?
No
Yes
Initiate troubleshooting
protocol
Determine whether
equipment should be
removed from patient
use
Resume/Continue
Routine Monthly Testing
Yes
Results < 50
CFU/mL?
-Evaluate/correct
sample collection
technique
-Evaluate/correct
bicarbonate preparation/
distribution technique
-Evaluate/correct water
system components
-Evaluate/replace
equipment ultrafilters
-Evaluate/implement
biofilm removal protocols
Redraw sample
Figure 1Example of decision tree that can be used to evaluate culture results
and initiate corrective action, if necessary
26
27
The gel-clot LAL assay is not as sensitive as the kinetic assay and provides only a positive or negative result; that is,
it shows if endotoxin is presentor notat a particular concentration. Single tube gel-clot tubes are available from
several commercial sources, and kits with the following sensitivities are available: 0.015 EU, 0.03 EU, 0.06 EU, 0.125
EU, 0.25 EU, and 0.5 EU. At a minimum, two tubes should be run each time the assay is performed. The first tube
contains LAL reagent and the sample to be tested. The second tube contains LAL reagent, a known amount of
endotoxin, and the sample to be tested. The second tube acts as a positive control to confirm the absence of any
interference that might lead to a false negative result. Positive control tubes are available from the suppliers of
commercial LAL assays. More sophisticated testing protocols involving reagent controls may also be used (see 2.7),
but their use is optional in this application.
Environment
The water purification and storage system should be located in a secure area that is readily accessible to authorized
users. The location should be chosen with a view to minimizing the length and complexity of the distribution system.
Access to the purification system should be restricted to those individuals responsible for monitoring and maintenance
of the system.
The layout of the water purification system should provide easy access to all components of the system, including all
meters, gauges, and sampling ports used for monitoring system performance. An area for processing samples and
performing on-site tests is also recommended. Critical alarms, such as those associated with deionizer exhaustion or
low water levels in a storage tank, should be configured to sound in the patient treatment area, as well as in the water
treatment room.
Water systems should include schematic diagrams that identify components, valves, sample ports, and flow direction.
Additionally, piping should be labeled to indicate the contents of the pipe and direction of flow. The use of text labels,
such as RO Water, and color-coded arrow tape provides a convenient means of identifying pipe content and flow
direction.
If water system manufacturers have not done so, users should label major water system components in a manner that
not only identifies a device but also describes its function, how performance is verified, and what actions to take in the
event performance is not within an acceptable range. An example of such labeling for a regenerable softener is given
in Figure 2.
Personnel
Policies and procedures that are understandable and accessible are mandatory, along with a training program that
includes quality testing, the risks and hazards of improperly prepared concentrate, and bacterial issues. Operators
should be trained in the use of the equipment by the manufacturer or should be trained using materials provided by
the manufacturer. The training should be specific to the functions performed (i.e., mixing, disinfection, maintenance,
and repairs). Periodic audits of the operators compliance with procedures should be performed. The user should
establish an ongoing training program designed to maintain the operators knowledge and skills.
28
Annex A
(informative)
Rationale for the development and provisions of this recommended practice
A.1
Scope
It has long been known that chemical and microbiological contamination of dialysate places hemodialysis patients at
risk of acute and chronic adverse events. As a result, the first edition of the American National Standard Hemodialysis
systems (ANSI/AAMI RD5:1981) included chemical and microbiological quality requirements for the water used to
prepare dialysate, as well as microbiological quality requirements for the final dialysate. From the beginning, the AAMI
Renal Disease and Detoxification Committee recognized a problem with including fluid quality requirements in a
standard directed at manufacturers of dialysis equipment. Although the manufacturer is responsible for providing
equipment that, when assembled as a system, provides water that meets the quality requirements of the standard, the
manufacturer has no control over that system once it is installed. Purified water quality may change if the system is
not well maintained or if there is some change in the municipal water feeding the system. The day-to-day
responsibility for maintaining water quality lies with the health care professionals at each dialysis facility under the
leadership of the facilitys medical director. For this reason, ANSI/AAMI RD5 included an appendix (Appendix B) that
provided users with guidelines for monitoring water purity and with criteria for selecting water treatment equipment.
The water and dialysate quality provisions of ANSI/AAMI RD5, as well as the guidelines provided in Annex B, were
subsequently adopted by the Centers for Medicare and Medicaid Services (CMS, formerly the Health Care Financing
Administration (HCFA)) as part of their conditions for coverage of end-stage renal disease services.
In 1996, during a 5-year review of ANSI/AAMI RD5, the AAMI Renal Disease and Detoxification Committee
determined that the hemodialysis community would be better served if ANSI/AAMI RD5 were divided into three parts:
(1) hemodialysis concentrates, (2) water treatment equipment for hemodialysis applications, and (3) hemodialysis
systems. This decision resulted in the publication of ANSI/AAMI RD61:2000, Concentrates for hemodialysis;
ANSI/AAMI RD62:2001, Water treatment equipment for hemodialysis applications; and ANSI/AAMI RD5:2003,
Hemodialysis systems. These standards are addressed to the manufacturers of equipment, although they also
provide users with a basis for understanding the products and processes they cover. In the process of creating the
three new standards, Appendix B of the original ANSI/AAMI RD5 was eliminated.
The critical product resulting from the joint application of the devices addressed by ANSI/AAMI RD61:2000,
ANSI/AAMI RD62:2001, and ANSI/AAMI RD5:2003 is the dialysate, the fluid against which the patients blood is
balanced. Although the proportioning of concentrates and water to produce dialysate is usually accomplished by a
hemodialysis machine, the actual production and handling of dialysate is under the control of health care
professionals who care for these dialysis patients. To fill the void left by the elimination of Appendix B from
ANSI/AAMI RD5, the AAMI Renal Disease and Detoxification Committee undertook the development of this
recommended practice. This recommended practice expands the guidelines originally included in Appendix B of
ANSI/AAMI RD5 to include all aspects of the selection and care of the systems involved in the preparation of
dialysate for hemodialysis. Those systems include water purification and distribution systems, concentrate preparation
and distribution systems, and dialysate proportioning systems. The recommended practice also includes
recommendations for the chemical and microbiological quality of the dialysate. The information provided in this
recommended practice is intended to complement the equipment standards set forth in ANSI/AAMI RD61:2000,
ANSI/AAMI RD62:2001, and ANSI/AAMI RD5:2003.
A.2
Normative references
For the purposes of this recommended practice, the references cited in clause 2 apply.
A.3
Definitions
For the purposes of this recommended practice, the definitions given in clause 3 apply.
A.4
Fluid quality
A.4.1 Water
The water quality recommendations contained in this recommended practice are the same as those set forth in
ANSI/AAMI RD62:2001 (see 2.3).
29
30
31
32
A.5
Equipment
33
The subclauses of 5.2 provide general information. Design and instrumentation of individual purification devices may
vary from these general descriptions. For example, softeners may be configured as a single resin bed that is
regenerated outside the normal operating hours of the dialysis unit, or they may have a dual-bed configuration that
allows one bed to be regenerated while the other is used to provide water for normal dialysis operations.
Depending on the feed water quality and product water requirements, not every component in 5.2 may be required in
a given facility. Likewise, additional components may be required in certain circumstances. For example, carbon
adsorption may not provide adequate chloramine removal if the water contains substances, such as polyphosphates,
that mask the reactive sites on the carbon particles. In those circumstances, other processes, such as infusion of
sodium metabisulfite, may be required to achieve product water that meets the requirements of 4.1.1.
Users are encouraged to obtain detailed descriptions of all purification processes, together with operating manuals
and maintenance procedures, from the manufacturer or the vendor providing the water purification and distribution
system.
A.5.2.5 Carbon adsorption
Although treatment of water by carbon adsorption is the method usually used to meet the requirement of 4.1.1 for
chloramines, the committee recognized that in certain situations carbon adsorption might not adequately remove
chloramines. Inadequate removal of chloramines may occur when the chloramines are in the form of naturally
occurring N-chloramines or when practices such as the use of high pH or the inclusion of orthophosphate or
polyphosphates are used to comply with the EPAs lead and copper rule. In other situations, such as acute dialysis
with portable water treatment systems, it may not be practical to use the volume of carbon required to ensure
adequate chloramine removal. In such circumstances, other strategies for chloramine removal may be needed to
supplement carbon adsorption. The committee is aware that adding sodium metabisulfite prior to the reverse osmosis
system has been successful in eliminating chloramine in hemodialysis applications. Ascorbic acid has also been
added to the acid concentrate used to eliminate chloramine from the final dialysate (Ward, 1996). It should be noted
that some minimum contact time is required for ascorbic acid to neutralize chloramine in water. If ascorbic acid is
being used to neutralize chloramine and if unexplained red cell destruction or anemia occurs, the effectiveness of the
ascorbic acid neutralization of chloramine should be investigated. Other means of removing chloramines, such as
redox alloy media and ultraviolet irradiation at 185 nm, are used in the pharmaceutical and electronics industries.
These processes are currently being evaluated for hemodialysis applications. The final choice of a system for
chloramine removal in hemodialysis settings will depend on local conditions and may need to include more than one
of the processes outlined above.
The committee recognized that it might not be practical to rotate the bed positions in installations that use large,
backwashable carbon beds. However, there was concern that the capacity of the second bed might decrease
unpredictably and no longer provide adequate backup if there was breakthrough of the first bed. For this reason, the
committee recommended replacing both beds if bed rotation was not possible.
A.5.2.6 Chemical injection systems
The committee expressed reservations about the addition of chemicals to the water. However, it recognized that the
addition of chemicals may be necessary in some circumstances if a facility is to meet the maximum contaminant
levels set forth in 4.1.1. For example, if the municipal water contains high levels of N-chloramines or chloramine in the
presence of orthophosphate or polyphosphate, injection of sodium metabisulfite may be one of the few options
available for chloramine removal.
If chemical injection is used in the pretreatment cascade, users should ensure that the addition of the chemical does
not interfere with the operation of subsequent purification processes, including the primary purification process. For
example, the performance of thin-film composite reverse osmosis membranes may be affected by the pH of the feed
water. At pH levels below 7, the rejection of fluoride may be substantially reduced, compared to its rejection at a pH of
8.
A.5.2.8 Deionization
Deionizers are an effective means of removing ionic contaminants from water. However, they do not remove nonionic
species, and they may contribute bacterial contaminants to the water rather than remove them. The inability of
deionizers to remove nonionic contaminants may limit aluminum removal by deionization, since aluminum is an
amphoteric substance that changes from cationic to anionic as the pH varies from acidic to basic (Stumm and
Morgan, 1996). At neutral pH, aluminum is present mostly as colloidal aluminum, which does not carry a charge and
is not removed by deionization (Parkinson, et al., 1981). Furthermore, deionizers have a finite capacity for
contaminant removal. Once the deionizer is depleted of hydrogen and hydroxyl ions, the next least avidly bound ions
will be displaced by more avidly bound ions. For example, once the hydroxyl ions are depleted, anionic contaminants
in the water will displace fluoride ions from the anion exchange resin (Bland, et al., 1996). This phenomenon has led
to high levels of fluoride in the product water, with subsequent patient injury and death (Arnow, et al., 1994; Johnson
34
and Taves, 1974). For the above reasons, use of deionization as the primary means of purification is strongly
discouraged. Deionization may be used to polish product water from a reverse osmosis system or may be used as a
standby if the reverse osmosis system fails. Some members of the committee believe that a two-stage reverse
osmosis system is preferable to a combination of reverse osmosis and deionization.
Deionizers offer a large surface area for bacterial proliferation and deionizers generally contribute to the bioburden in
the water. Therefore, the committee strongly recommends that deionizers be followed by an ultrafilter. The tendency
for deionizers to contribute bacterial contaminants to the water is greater when deionizers are kept as a backup for a
reverse osmosis system, particularly if there is no flow through the deionizers. Some facilities counter this tendency
by connecting the deionizers in parallel to the main water line and by maintaining a low flow through them. An
alternative approach is to contract with a local vendor to provide backup deionizers on demand.
A.5.3 Water storage and distribution
The design of the water storage and distribution system is a critical element in preventing bacterial proliferation that
may otherwise lead to water and dialysate not meeting the quality requirements of clause 4. The water storage and
distribution system should be designed to provide the simplest possible flow path and to contain the smallest volume
of water consistent with the operating needs of the dialysis unit. The simplest system is generally a direct feed
system, in which purified water is piped directly from the last stage of the purification system to the points of use.
However, direct feed systems are frequently impractical. For example, the pressure at the end of the purification
cascade may be insufficient to provide adequate flow and pressure at the points of use without a booster pump. In
this circumstance, an indirect feed system with a storage tank is required. Since storage tanks provide a large surface
area for potential biofilm formation, their volume should be kept to a minimum in order to maximize water turnover in
the tank.
One consequence of the increased attention being paid to bacterial control in the water storage and distribution
system is an interest in alternatives to traditional chemical disinfection. Two approaches have been developed, one
based on ozone and the other on hot water. Both techniques may allow more frequent disinfection of the water
storage and distribution system, because prolonged rinsing is not needed to remove residual disinfectant from the
system before dialysis is recommenced. The use of ozone or hot water is possible only if the systems are constructed
from appropriately resistant materials. This limitation applies not only to the piping and any storage tank that may be
in the system, but also to all pumps, valves, and other fittings, including any O-rings and seals they may contain.
Direct feed water distribution systems typically return unused water to the feed side of the reverse osmosis unit. If the
pressure at the end of the distribution loop decreases to a value below the water pressure at the inlet to the reverse
osmosis pressurizing pump, retrograde flow of nonpurified water into the distribution loop can occur. To minimize this
risk, the committee recommends that dual check valves be used to prevent retrograde flow and that the pressure at
the end of the distribution loop be monitored.
A.5.4 Concentrate preparation
A.5.4.4.3 Bicarbonate concentrate mixing systems
Bicarbonate concentrates have been shown to support bacterial growth and to provide another source of initial
bioburden capable of rapidly increasing after dilution (Ebben, et al., 1987; Bland, et al., 1987). Therefore, additional
precautions should be taken when preparing and handling bicarbonate concentrate to avoid excess growth of
haloduric organisms. Also, prompt use of bicarbonate concentrates prepared in dialysis facilities from powder and
purified water is strongly recommended.
Overagitation or mixing of bicarbonate concentrate may result in loss of CO2 from the solution. Loss of CO2 results in
an increase in pH and favors the formation of carbonate that can lead to precipitation of calcium and magnesium
carbonate in the fluid pathways of the dialysis machine following dialysate proportioning.
A.5.6 Dialysate proportioning
Dialysate is usually prepared by a proportioning system that sequentially adds acid concentrate and bicarbonate
concentrate to purified water. These systems produce a buffered physiologic dialysate with a pH between 6.9 and 7.6.
Current dialysis machines are also capable of supplying acetate dialysate. For acetate dialysate, purified water is
mixed with a single acetate concentrate to produce the dialysate. This system is not buffered, so the pH can vary
depending on the supply water. More recently, systems have been developed that use three concentrates
(bicarbonate, sodium chloride, and an acid concentrate containing the remaining electrolytes) to allow more
sophisticated variation of the dialysate composition during dialysis.
35
A.6
Monitoring
A.7
36
The microbiological purity of packaged liquid concentrates and dry powder cartridges is the responsibility of the
manufacturer (see 2.4). Monitoring of bicarbonate concentrate produced at a dialysis facility from powder and water,
though not required routinely, may be undertaken as part of a troubleshooting investigation. The sodium content of
TSA is sufficient for use in culturing bicarbonate concentrate without supplementation. However, if Reasoners 2A or
TGEA is used to monitor microbial contamination in bicarbonate concentrate, it may be necessary to supplement the
media with 4 % sodium bicarbonate.
A.7.2.4 Bacterial endotoxin test
Bicarbonate concentrate inhibits the LAL assay. Inhibition is caused by the high concentration of solutes in the
concentrate and the pH. In the gel-clot assay, this inhibition results in a failure of the positive control to clot. In kinetic
assays, inhibition results in a failure to recover the spike (product positive control) to within 50 % to +200 % of the
nominal value.
Dilution of the bicarbonate concentrate with water is the usual method for overcoming inhibition. A minimum dilution of
1:16 is necessary; however, higher dilutions are recommended for more sensitive assays: for example, a 1:20 dilution
is recommended when using an assay with a sensitivity of 0.03 EU/mL. In kinetic methods, the sensitivity is the lowest
concentration used to construct the standard curve.
Standard gel-clot tests are incubated at 37 C for 1 hour. At the end of 1 hour, the tubes are inverted to detect the
presence of a clot. A positive test will have a clot, which will remain in the end of the tube as long as it is not shaken
or bumped. A negative test will not have a clot and will tend to flow out of the tube. A clot that is semisolid and flows
slowly is classified as a negative clot. For example, when bicarbonate concentrate is tested using a gel-clot assay with
a sensitivity of 0.03 EU/mL and the concentrate is diluted 1:20 (1.0 mL concentrate plus 19 mL LAL reagent water or
equivalent), the positive control sample should clot, indicating that there is no inhibition of the assay. If the diluted
bicarbonate concentrate sample clots, it indicates that the sample contains at least 0.6 EU/mL of endotoxin. (To test
at the 1.0 EU/mL action level using an assay with a sensitivity of 0.03 EU/mL, the concentrate is diluted 1:32, or 1 mL
concentrate plus 31 mL LAL reagent water or equivalent.) A new version of the gel-clot assay has a matched positive
control, which simplifies the testing and increases the reliability of the results.
Tables A.1 and A.2 show the relationship between assay sensitivity, sample dilution, and the lowest level of endotoxin
that can be detected. Table A.1 shows the lowest level of endotoxin (EU/mL) that can be detected using an assay of a
given sensitivity and a specified dilution of a bicarbonate concentrate sample. Table A.2 shows the maximum sample
dilution that can be used with an assay of a given sensitivity if the objective is to detect endotoxin concentrations as
low as 1 EU/mL or 2 EU/mL.
1:16
1:20
1:32
0.015
0.25
0.3
0.5
0.03
0.5
0.6
1.0
0.06
1.0
1.2
2.0
0.125
2.0
NA
NA
0.25
NA
NA
NA
NA = not applicable. An assay with this sensitivity cannot be used to determine endotoxin
concentrations 2 EU/mL at this dilution.
37
1 EU/mL
2 EU/mL
0.015
1:64
1:128
0.03
1:32
1:64
0.06
1:16
1:32
0.125
1:8*
1:16
0.25
1:4*
1:8*
0.5
1:2*
1:4*
38
Annex B
(informative)
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41