`ASIAN PACIFIC JOURNAL OF ALLERGY AND IMMUNOLOGY (2010) 28: 53-57
` 
`
`
`The Stability and Sterility of Epinephrine
`Prefilled Syringe
`
`
`
`Saowanee Kerddonfak, Wiparat Manuyakorn, Wasu Kamchaisatian, Cherapat Sasisakulporn,
`Wanlapa Teawsomboonkit and Suwat Benjaponpitak
`
`
`
`
`
`
`
`In anaphylaxis, the vasopressive effect of ep-
`inephrine, along with its effects in preventing and re-
`living laryngeal edema and bronchoconstriction is
`life saving. Epinephrine also generates an inotropic
`effect, which increases in heart rate, contraction of
`the heart and vasoconstriction, thereby increasing the
`blood pressure.
`
`
`The efficacy of epinephrine in suppression
`of mast cells and basophils inflammatory mediators’ 
`release is also considerably importance.4 The fatality
`during witnessed anaphylaxis, most of which occurs
`outside of medical facility, usually results from de-
`
`SUMMARY The commercially available auto-injector epinephrine is considerable expensive. Epinephrine pre-
`filled syringe is an alternative treatment for anaphylaxis patients. The objective of the present study was to evaluate
`the stability and sterility of epinephrine prefilled syringe. Epinephrine prefilled syringe was kept in the pencil box to
`prevent from light exposure. The active ingredients, integrity and level of potency were measured by high-
`performance liquid chromatography (HPLC). The sterility was accessed by aerobic bacteria and fungi culture. The
`epinephrine concentration at 1, 2 and 3 months after the preparation was 101.36, 99.31 and 101.09%, respectively
`(acceptable range 90 - 110%). The pH was 3.17 - 3.23 (acceptable range 2.8 - 3.6). Nor-epinephrine was unde-
`tected. The cultures for bacteria and fungus were both negative. Consequently, epinephrine prefilled syringe was
`stable and sterile at least three month after preparation. Epinephrine prefilled syrine is an alternative low cost
`treatment for anaphylaxis patient. 
` 
` 
` 
` 
`Anaphylaxis is a serious allergic reaction
` 
`that is rapid in onset and may cause death. It is
`commonly triggered by food, insect sting, medica-
` 
`tions or natural rubber latex. Epinephrine (adrena-
`line) is widely advocated as the main drug in the
`
`treatment of anaphylaxis. There are no other medica-
`
`tions with similar efficacy on the body systems that
`
`are potentially involved by anaphylaxis.1
`
`
`
`Epinephrine is an endogenous catecholamine
`produced primarily in the adrenal medulla with a
`wide variety of clinical uses. Epinephrine is poorly
`soluble in water, but readily forms water-soluble
`salts in the presence of acid. Injectable solution is
`buffered to maintain a pH at 2.2 to 5.0. Epinephrine
`solution is colorless. However, oxidation of the cate-
`chol nucleus will impart a pink color then changes to
`a pink-brown color.2, 3
`
`From the Division of Allergy and Immunology, Department of Pe-
`diatrics, Faculty of Medicine Ramathibodi Hospital, Mahidol Uni-
`versity, Bangkok, Thailand 10400
`Correspondence: Suwat Benjaponpitak
`E-mail: rasbj@mahidol.ac.th
`
` 
` 
` 
` 
` 
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`54
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`
`KERDDONFAK, ET AL.
`
`layed administration of epinephrine.5 For out of hos-
`pital emergency treatment, epinephrine auto injector
`such as EpiPen is prescribed. However, self-
`injectable epinephrine is underused when anaphylax-
`is occurrs6. The drawback of epinephrine autoinjector
`includes high cost which limits affordability and
`availability worldwide.7 Moreover, it is impossible to
`give an accurate dosage for infant and many children
`by using currently available autoinjector fixed epi-
`nephrine dose 0.15 or 0.3 mg.8
`
`
`In Thailand, the prefilled syringe with an ap-
`propriate dose of epinephrine is prescribed to pa-
`tients and parents of children with history of anaphy-
`laxis, those currently on immunotherapy treatment
`and patients with history of severe asthma with food
`allergy. Serious concern arises, particularly in hot
`climates, from possible partial solution contamina-
`tions and degradation of the drug.9, 10
`
`
`The aim of this study was to determine the
`physical, chemical stability and sterility of epineph-
`rine prefilled syringe comparing between drawing up
`from ampules into disposable 1-ml syringes under
`laminar flow hood (sterile technique) and open air.
`
`
`M A T E RI A L A ND M E T H O DS
`
`
`Materials
`
`Epineprine 1 mg/ml from the same batch
`were purchased from the Government Pharmaceuti-
`cal Organization (GPO, Bangkok, Thailand). Dis-
`posable plastic 1-ml syringes and 23-guages needles
`were purchased from Nipro corporation (Osaka, Ja-
`pan).
`
`
`Preparation and storage of prefilled epinephrine
`syringes
`
`We evaluated the effect of different mode of
`preparation (under laminar flow hood and open air)
`on chemical, physical stability and sterility of a 1.0
`mg of epinephrine dose loaded in 1 ml disposable
`syringe overtime up to 3 months after preparing and
`storing in the container under room temperature.
`
`
`One hundred and forty syringes were loaded
`with 1 ml of a 1-mg/ml epinephrine solution under
`laminar flow hood or open air. All of the epinephrine
`
`doses were drawn up by the same person and on the
`same day. Air bubbles were removed to reduce oxy-
`gen exposure during storage. Needle were left at-
`tached and recapped.
`
`
`Seventy prefilled epinephrine syringes in
`each group were randomly allocated to 3 time points
`(1 month, 2 months and 3 months). All of the sy-
`ringes were kept in the pencil boxes and left in room
`temperature. Room temperature was recorded every-
`day at 8.00 - 9.00 am.
`
`Physical and chemical stability evaluation
`
`At each time point, 20 prefilled syringes
`were randomly selected for evaluation. Initially, a
`visual inspection was made and any observation was
`noted (dirt, deterioration, cracks, etc.). The pH of
`each sample was determined using a pH meter. Epi-
`nephrine and nor-epinephrine, which is the product
`of epinephrine degradation in solution, concentra-
`tions were measured by high-performance liquid
`chromatography (HPLC). Analysis was performed
`on a reverse-phase Inertsil ODs 3 C18 HPLC column
`(4.6 mm x 150 mm) equilibrated with mobile phase
`at a flow rate of 1.0 ml/minute. Ultraviolet absor-
`bance was monitored at 205 nm. Using a variable
`wavelength ultraviolet detector (Dionex UVD 340 u)
`an injection valve was configured with a 20 l sam-
`ple loop. Epinephrine concentration was determined
`by comparing the peak area ratio (the ratio of the
`area under the epinephrine peak to that of the area
`under the internal standard peak) of sample solution
`to a known standard concentration of epinephrine.
`Standard solution of epinephrine bitartrate (0.02%
`w/v), sample solution (0.012% w/v) and the internal
`standard paracetamol (0.08% w/v) were prepared in
`mobile phase. The mobile phase was prepared by
`combining methanol and 10 mM sodium heptane
`sulfonate in water pH 3.5 containing 0.0002 M Dis-
`odium Edentate (23:77).
`
`Standard solution of nor-epinephrine bitar-
`trate (0.0018% w/v) was prepared in mobile phase.
`Sample solution was the epinephrine injection (non-
`diluted). The peak area of nor-epinephrine obtained
`from the epinephrine injection does not exceed the
`normalized peak area of nor-epinephrine bitartrate
`(0.0018% w/v).11
`
` 
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`STABILITY AND STERILITY OF EPINEPHRINE SYRINGE
` 
`
`55
`
`Bacterial and fungal culture for sterility evalua-
`tion
`
`
`At each time point 2 prefilled syringes were
`randomly selected for evaluation. Each specimen of
`epinephrine (1 ml) was inoculated into a blood agar
`and incubated at 35ºC in air. Each inoculate was ex-
`amined for growth every 24 hour for 3 days. This
`microbiological method supported the growth of
`gram-negative and gram-positive bacteria. For fungal
`cultures, inoculation was made into brain heart blood
`agar at 37 ºC for 3 days and inoculation was into sa-
`bouraud dextrose agar at 37 ºC for 1 month.
`
`Statistical analyses
`
`
`Data obtain from this study, including gen-
`eral appearance, epinephrine and nor-epinephrine
`concentration, pH values and presence or absence of
`microbial growth from each epinephrine syringe, and
`were described as descriptive data.
`
`
`R ESU L TS
`
`A total of 140 of epinephrine prefilled sy-
`ringes were prepared and stored at room temperature
`(26 ± 3 ºC) in the pencil boxes for light protection.
`All samples were clear in appearance at 1, 2 and 3
`months from preparation time, respectively. Epi-
`nephrine concentration was 99.31 - 102.68% (ac-
`ceptable range 90 - 110%).The pH was 3.17 - 3.23
`(acceptable range 2.8 - 3.6) as shown in Table 1. No-
`repinephrine, which is a degradation product of epi-
`nephrine, was not detected in any sample. There
`were no aerobic bacterial or fungal growths in any
`sample after 1, 2 and 3 month from preparation time.
`
`However, we found some brown particles at
`the needle cap in some syringes. These brown par-
`
`
`ticles were sent for bacterial and fungal culture with
`negative result. We hypothesized that the brown par-
`ticles were from the reaction between epinephrine
`and the environment.
`
`
`DISC USSI O N
`
`
`In life-threatening emergencies, it is essen-
`tial to immediately inject epinephrine to the anaphy-
`lactic patient. In many countries, autoinjector epi-
`nephrine is unaffordable or unavailable, anaphylaxis
`patients usually are prescribed with an ampule of ep-
`inephrine and a 1-ml syringe and needle. However,
`when anaphylaxis occurs in the community, patients
`or caregivers of children have to draw up epineph-
`rine from an ampule at a time of crisis. A previous
`study has demonstrated that most parents are unable
`to draw up an infant epinephrine dose rapidly and/or
`accurately. The parents had a significantly longer
`time to draw up an epinephrine dose (142 ± 13
`seconds) compared to emergency department nurses
`(29 ± 0.09 seconds).9 Consequently, it is much bet-
`ter to have syringes already prepared rather than
`drawing up from the ampoules at the emergency
`time.12 Prefilled epinephrine syringes have become
`commonly used in unaffordable or unavailable coun-
`try such as developing country. The reason for the
`widespread use of prefilled epinephrine syringes
`over EpiPen is their much lower price.13 Moreover,
`EpiPen is only available in a fix dosage which may
`cause problem with infant and young children of
`overdose of epinephrine and under dose obese.
`
`
`According to the United States Pharmaco-
`poeia (USP) stability guidelines, a product has chem-
`ical stability if each of the active ingredients retains
`its intrepidity and labeled potency overtime and mi-
`crobiological stability of the product remains sterile
`and resistant to microbial growth. The USP/National
`
`Table 1 The epinephrine concentration and pH of the solution in prefilled syringes left over 1, 2 and 3 months
`
`Time
`Specification
`
`1 month
`
` Laminar flow
`Open air
`hood
`
`2 months
`
` Laminar flow
`Open air
`hood
`
`3 months
`
` Laminar flow
`Open air
`hood
`
`Epinephrine con-
`centration
`pH
`
`90-110%
`
`2.8-3.6
`
`
`
`
`
`101.40
`
`101.36
`
`3.22
`
`3.23
`
`
`
`
`
`102.68
`
`3.22
`
`99.31
`
`3.22
`
`
`
`
`
`101.19
`
`101.09
`
`3.17
`
`3.17
`
`55 
`
`
`
` 
` 
` 
` 
` 
` 
`
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`56
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`KERDDONFAK, ET AL.
`
`Formulary monograph requires the product to con-
`tain 100 ± 10% of label potency. Therefore, the t90%
`of product represents the effective shelf-live of prod-
`uct, otherwise known as the expiry time. The shelf-
`life stability of epinephrine injection stored in US
`hospitals was analyzed as part of the FDA-ASHP vo-
`luntary drug stability program. USP requirements
`are as follow: epinephrine injection is formulated to
`contain 90 - 115% of the labeled amount of epineph-
`rine, the pH of the injections should be in the range
`of 2.5 - 5.0 There are no compound requirements for
`degradation products (such as nor-epinephrine).11
`
`
`The epinephrine ampoule is recommended to
`be stored between 15 to 30 °C by the manufacturer
`in order to keep its stability. In Thailand, however,
`the temperature is between 23.9 °C to 34.9 °C. Kelly,
`et al14 found that epinephrine stability decreased by
`exposure to elevated temperature. Storage at 37 ºC
`for 6 months decreased epinephrine concentration by
`50%. Fry, et al15 also reported that the rate of epi-
`nephrine degradation will increase if the temperature
`increases. Storage at 50 ºC causes a 25% loss within
`2 months. Grant, et al2 determined the biological
`consequence of temperature induced epinephrine de-
`gradation and discovered that the environmental
`temperature variation causes degradation in epineph-
`rine concentration and biological activity. The de-
`gradation of epinephrine ampoule (1:1000) was not
`significant even after 12 weeks of heat exposure. No
`change was noted from control16. It may be con-
`cluded that the epinephrine prefilled syringe should
`not be kept more than 3 months. Up to our know-
`ledge, there are a few studies elucidating the stability
`of prefilled epinephrine syringes. A previous study
`has found that the stability of prefilled epinephrine
`solutions in unsealed syringes was 2 months and 3
`months under 38°C with 15% and 95% humidity, re-
`spectively.17 Nevertheless, there is no earlier study
`has attempted to evlauate the sterility of prefilled ep-
`inephrine syringes before. In this current study, we
`have demonstrated that prefilled epinephrine sy-
`ringe is stable 3 months either preparing in laminar
`flow hood or in open air. However, we have found
`the brown particle at the needle cap of some epi-
`nephrine prefilled syringes when we kept them up to
`3 months. After test with bacterial and fugal culture,
`they all were negative. We hypothesized that the
`brown particle at the tip of the needle cap was caused
`by degradation of adrenaline via oxidation to adre-
`
`nochrome, which turns pink first, then pink-brown
`oxidative product.18 Therefore, we recommended
`changing the needle after drawing up the epinephrine
`and do not push epinephrine back through the needle
`to keep it away from the air. Finally by lengthen time
`of observation, we found that the color of most of the
`4-month prefilled epinephrine syringes was changed
`to pink-brown solution. On the basic of these result,
`we concluded that epinephrine prefilled syringe pre-
`paring in either laminar flow hood or open air was
`stable up to 3 months without loss of chemical stabil-
`ity or sterility.
`
`
`
`A C K N O W L E D G E M E N TS
`
`
`
`This research would not have possible with-
`out the collaboration and effort of the Government
`Pharmaceutical Organization: Pradistha Santikul, the
`Quality Assurance Department, the Government
`Pharmaceutical Organization, Pinya Tanuwong, Mi-
`crobiology Department, Faculty of Medicine Rama-
`thibodi Hospital, Mahidol University, and Achara
`Boonpason, the Quality Assurance Department, the
`Government Pharmaceutical Organization, for de-
`termine property of epinephrine prefilled syringes.
`The authors would also like to thank Piriyaporn
`Chogtrakol, Microbiology Department, Faculty of
`Medicine Ramathibodi Hospital, Mahidol Universi-
`ty, for microbiology examination.
`
`R E F E R E N C ES
`
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`4.
`Art.
`No.:CD006312.
`DOI:10.1002/14651858.CD006312.pub2.
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