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Second Supplement to USP 39NF 34 General Information / 1229.13 Sterilization-in-Place 8371

The activities in three unprocessed EIs are 1286, 1000, and 1532 EU/mL
The activities in three processed EIs are 0.634, 0.512, and 0.496 EU/mL
The log reduction is calculated as:
log10 (1000) log10 0.634 = 3 (0.198) = 3.198 log reduction

Historically, a 3-log reduction has been required by regulatory/compliance guidance. However, depending on the process
and historical data, a 3-log reduction may be either excessive or inadequate. For example, for glass vials with a low or non-
measurable endotoxin content upon receipt, the requirement to continually and repeatedly revalidate with an acceptance cri-
terion of a 3-log reduction of the endotoxin spike of >1000 EU is excessive. Alternatively, a fermentation process with an endo-
toxin content of >107 EU/mL in the clarified culture supernatant will require more than a 3-log reduction to achieve safe levels
of endotoxin in the drug substance or drug product. Additionally, given the sensitivity of BET assays, it may not be necessary
to spike with 1000 EU to demonstrate a 3-log reduction. For example, if the assay sensitivity is 0.005 EU/mL, one may choose
to spike with 50 EU and demonstrate an ultimate recovery in the test articles of less than 0.05 EU/mL, which calculates to
greater than a 3-log reduction. In any event, the design of experiments including an appropriate specification for the log re-
duction of processed indicators and required test sensitivity to demonstrate the specified log reduction should be established
and justified in a preapproved protocol for the study. The total reduction, of course, may be achieved over several steps in a
purification process. Thus, the necessary reduction is often achieved additively over the course of multiple purification steps.

REFERENCES

1. Ludwig JD, Avis KE. Dry heat inactivation of endotoxin on the surface of glass. J Parenter Sci Technol. 1990;44(1):412.
2. Bowers K, Tran L. Creation of an in-house naturally occurring endotoxin preparation for use in endotoxin spiking studies
and LAL sample hold time analysis. Am Pharmaceut Rev. 2011;14(6):9297. http://www.americanpharmaceuticalre-
view.com/Featured-Articles/37219-Creation-of-an-In-house-Naturally-Occurring-Endotoxin-Preparation-for-Use-in-Endo-
toxin-Spiking-Studies-and-LAL-Sample-Hold-Time-Analysis/
3. LAL Users Group. Preparation and use of endotoxin indicators for depyrogenation process studies. J Parenter Sci Technol.
1989;43(3):109112.
4. Berzofsky RN, Scheible LS, Williams KL. Validation of endotoxin removal from parenteral vial closures. BioPharm. June
1994:5866.
5. Ross VC, Twohy CW. Endotoxins and medical devices. Prog Clin Biol Res. 1985;189:267281.
6. Twohy CW, Duran AP, Peeler JT. Extraction of bacterial endotoxin from medical devices. J Parenter Sci Technol.
1986;40:287291.
7. Bryans TD, Braithwaite C, Broad J, Cooper JF, Darnell KR, Hitchins VM, et al. Bacterial endotoxin testing: a report on the
methods, background, data, and regulatory history of extraction recovery efficiency. Biomed Instrum Technol. 2004;38(1):
7378.
8. ANSI/AAMI standard ST72:2011. Bacterial endotoxinstest methods, routine monitoring, and alternatives to batch test-
ing. Arlington, VA: Association for the Advancement of Medical Instrumentation; 2011.
2S (USP39)

Add the following:

1229.13 STERILIZATION-IN-PLACE

INTRODUCTION

Sterilization-in-place (SIP) can be defined as the sterilization of a system or piece of process equipment in situ. The purpose
of SIP1 is to eliminate, or greatly reduce, the need for post-sterilization handling, including that necessary to make aseptic con-
nections. Mobile process equipment (e.g., portable tanks, storage vessels, and other equipment), once sterilized in this man-
ner, may be relocated. The SIP process can be carried out by using any of the following physical methods: moist heat, dry
heat, gas, liquid, or vapor (described below) according to the approaches described in Sterilization of Compendial Articles
1229, as adapted for use with the specific equipment or system.

1 AAMI/ISO 13408-5. Aseptic processing of health care productspart 5: sterilization in place; 2008.

Official from December 1, 2016

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8372 1229.13 Sterilization-in-Place / General Information Second Supplement to USP 39NF 34

COMMON ELEMENTS OF SIP PROCESSES

There are a number of considerations appropriate for the design and use of SIP procedures that apply to all of the steriliza-
tion methods:
The use of an SIP method is normally associated with a closed system. Closed systems are almost always sterilized in
situ, and the design elements of the typical closed system are consistent with many SIP process needs.
For large systems, it may be necessary to sterilize in portions. The individual sterilization processes should overlap to en-
sure treatment of all internal surfaces.
The focus of the SIP procedure is sterilization of the product contact surfaces (the interior of the system). Demonstration
of process lethality relies upon physical measurements and biological indicators. This confirmation should extend to the
sterile boundary of the system, including vessel headspace, connections to other vessels/equipment, and other parts of
the system. The interior surfaces of the process equipment, irrespective of their materials of construction, should be ex-
posed to lethal conditions sufficient to sterilize the system and confirmed as lethal with an appropriate biological chal-
lenge.
SIP is accomplished almost exclusively using the overkill approach to sterilization. The components of the physical equip-
ment should be chosen based upon their ability to withstand the sterilizing conditions to be used. Filters in the process
system, whether membrane or high-efficiency particulate air (HEPA), are typically susceptible to damage during SIP, and
care must be taken to preserve their integrity. Filter manufacturers can provide guidance on acceptable sterilization meth-
ods and parameters.
The absence of specifically designed equipment in which the sterilization process is performed places the bulk of responsi-
bility for design onto the user. SIP systems ordinarily cannot be purchased directly in the way one purchases a steam steri-
lizer or a dry heat oven. Instead, the system, which was designed for the operating process, may require modification to
accommodate the SIP process to be used. The user must assume the role of designer for the process, equipment, and
control system.
The system design and operating procedures must provide for an efficient means of introducing and removing the sterili-
zation agent. Sterilization agent removal must consider the potential effects of residual sterilant on the materials to be
processed. Establishment of a reliable process sequence is a critical part of the cycle development exercise. The sterilizing
agent is normally introduced through a filter on the system that may also serve as a process, purge gas, or vent filter.
At the conclusion of the sterilization process sequence and until ready for use, the system should be pressurized with a
purge gas (sterile air or nitrogen are the most common) to prevent the introduction of contaminants to the sterilized sys-
tem.
The critical process parameters for the SIP process should be recorded as the process is executed. The important parame-
ters may include temperature, pressure, concentration, flow rate, humidity, and time, among others.

CLOSED SYSTEMS

In pharmaceutical manufacturing operations, closed systems are used for various applications including maintenance of
large quantities of materials (liquids or powders) in a sterile state; manufacture of biological and synthetic organic active ingre-
dients (especially where microbial absence is essential); and preparation of process equipment for use in sterile drug product
manufacturing and filling. The use of closed systems provides superior separation of sterile materials from the surrounding en-
vironment. Typically, closed systems are maintained under positive pressure at all times. The characteristics of a closed system
that establish its designation as closed include the following:2
It maintains integrity during all operating periods and under all conditions.
It is sterilized-in-place or sterilized while closed before use.
It can be adapted for materials transfer in and/or out while maintaining its sterile state.
It can be connected to other closed systems while maintaining the integrity of all systems.
It is subject to scheduled preventive maintenance.
It uses sterilizing-grade filters for sterilization of liquid and gas process streams.

SIP METHODS

Moist Heat

The use of saturated steam is the most prevalent method for SIP of large systems. The majority of installations use gravity
displacement cycles adapted from those originally used in steam sterilizers (the size and complexity of many systems preclude
the use of prevacuum cycles). Important considerations include the provision for air removal, condensate discharge, and steam

2 Parenteral Drug Association, Technical Report No. 28, Revised. Supplement Volume 60, No. S-2, Process simulation testing for sterile bulk pharmaceutical chemi-

cals. Bethesda, MD: PDA; 2005.

Official from December 1, 2016

Copyright (c) 2017 The United States Pharmacopeial Convention. All rights reserved.
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Second Supplement to USP 39NF 34 General Information / 1231 Water for Pharmaceutical Purposes 8373

removal post-dwell.3,4 This method is commonly used for bioreactors, sterile bulk production, holding tank and delivery lines,
and other large systems.

Superheated Water

Systems used for Water for Injection and Purified Water can be sterilized by using superheated water (water that is heated
above its boiling point and pressurized to maintain it as a liquid) circulating through the system. This method has the ability to
sterilize vessels, filters, and other wetted components at the same time.5 Removal of residual water subsequent to the steriliza-
tion phase is recommended.

Dry Heat

Dry heat has been used for SIP of spray dryers and their associated material collection systems. The air supply for sterilization
in these systems is provided through HEPA filters.

Gas

Gas-phase SIP has been used for non- and low-pressure-rated process equipment, such as freeze dryers, prefreezers, process
vessels, and other equipment.

Liquid

Liquid chemical sterilization is best suited for liquid-handling systems and can be used only for fully wetted surfaces. This
process is similar to those using superheated water except the lethal modality is chemical rather than thermal.

Vapor

Sterilizing vapors have been used for the in situ sterilization of the same types of process equipment as those treated with
sterilizing gases. The precautions associated with vapor sterilization described in Vapor Phase Sterilization 1229.11 are re-
quired.

ROUTINE PROCESS CONTROL

SIP processes are subject to formal controls that maintain a validated state over time. The practices outlined in 1229 in-
clude the general requirements appropriate for all sterilization systems as well as those specific to an individual sterilization
method. Sterilization is accomplished by a number of related practices that are essential for continued use of the process over
an extended period of time. The essential practices to maintain validated status include calibration, physical measurements,
physical integrators or indicators, ongoing process control, change control, preventive maintenance, periodic reassessment,
and training. 2S (USP39)

1231 WATER FOR PHARMACEUTICAL PURPOSES

Change to read:

Table of Contents
1. INTRODUCTION
2. SOURCE WATER CONSIDERATIONS
3. WATERS USED FOR PHARMACEUTICAL MANUFACTURING AND TESTING PURPOSES
3.1 Bulk Monographed Waters and Steam
3.1.1 Purified Water
3.1.2 Water for Injection
3.1.3 Water for Hemodialysis

3 Agalloco J. Steam sterilization-in-place technology and validation. In: Agalloco J, Carleton FJ, editors. Validation of pharmaceutical processes. 3rd ed. New York:

Informa USA; 2007.

4 Parenteral Drug Association, Technical Report No. 61, Steam in place. Bethesda, MD: PDA; 2013.
5 Haggstrom M. Sterilization-in-place using steam or superheated water. In: Proceedings of the PDA Basel conference. Bethesda, MD: PDA; 2002.

Official from December 1, 2016

Copyright (c) 2017 The United States Pharmacopeial Convention. All rights reserved.