CURRENT MEDICAL RESEARCH AND OPINION® 0300-7995

VOL. 21, NO. 8, 2005, 1235–1240 doi:10.1185/030079905X56510

© 2005 LIBRAPHARM LIMITED All rights reserved: reproduction in whole or part not permitted

BRIEF REPORT

Second-degree burns: a
comparative, multicenter,
randomized trial of hyaluronic
acid plus silver sulfadiazine vs.
silver sulfadiazine alone
M. Costagliola a and M. Agrosì b
a
Department of Plastic and Reconstructive Surgery, Centre Hospitalier
Universitaire, Toulouse Rangueil, France
b
Fidia Farmaceutici SpA, Abano Terme (Pd), Italy

Address for correspondence: Dr.ssa Mirella Agrosì, Direzione Ricerca e Marketing Strategico,
Fidia Farmaceutici SpA, Via Ponte della Fabbrica 3/A, 35031 Abano Terme (PD), Italy.
Tel.: +39-049-8232630; Fax: +39-049-810653; email: magrosi@fidiapharma.it
Key words: Burns – Hyaluronic acid – Silver sulfadiazine – Wound healing

ABSTRACT

Aims: This multicenter, multinational, randomized,
double-blind, controlled, parallel-group study,
was designed to assess the efficacy and safety
of a fixed combination topical medicinal product,
containing 0.2% hyaluronic acid and 1% silver
sulfadiazine (HA-SSD) (Connettivina* Plus cream)
versus 1% silver sulfadiazine cream alone (SSD),
in the treatment of second-degree burns.
Methods: 111 adult patients (age 18–75
years) of both sexes, with IIa-degree (superficial)
and IIb-degree (deep dermal) burns, were
randomized to receive HA-SSD or SSD. Treatments
(approximately 5 g/100 cm2) were applied once
a day until the wounds healed, but for no longer
than 4 weeks. Burns had to have occurred within
48 hours from the start of treatment, be caused by
thermal injury, and be confined to the trunk and/or
upper and lower extremities.
Results: Results showed that both the fixed
combination HA-SSD, and SSD alone, were effective
and well tolerated topical agents for the treatment
of second-degree burns. All burns were healed
except in one patient treated with SSD. It was
also observed that the fixed combination HA-SSD
caused a significantly more rapid re-epithelialization
of burns, i.e. a shorter time to healing, than SSD
alone. The difference recorded – 4.5 days – was
statistically significant ( p = 0.0073).
Conclusion: The observed shorter time to
healing caused by the fixed combination is
clinically relevant and further demonstrates the
wound healing activity of HA.

Introduction causes include injury from flame, hot liquids (scald),

Burns cause coagulative necrosis of the epidermis and
underlying tissues, with the depth depending on the
temperature to which the skin is exposed and the
duration of exposure. Burns are classified into five
different causal categories and depths of injury. The
contact with hot or cold objects, chemical exposure, and
conduction of electricity. The first three induce cellular
damage primarily by the transfer of energy, inducing
coagulative necrosis. Chemicals and electricity cause
direct injury to cellular membranes in addition to the
transfer of heat. Burn depth is classified into degree of

* Connettivina is a registered trade name of Fidia Farmaceutici SpA, Abano Terme (PD), Italy

Paper 2970 1235

injury in the epidermis, dermis, subcutaneous fat, and
underlying structures.
First-degree burns are, by definition, injuries
confined to the epidermis. These burns are painful,
erythematous, and blanch to the touch with an intact
epidermal barrier. Second-degree burns are divided
into two types: superficial (IIa) and deep (IIb). All
second-degree burns have some degree of dermal damage,
and the division is based on the depth of injury into this
structure. Superficial dermal burns are erythematous,
painful, blanch to touch, and often blister. Examples
include scald injuries from overheated bathtub water
and flash flame burns from open carburettors. These
wounds spontaneously re-epithelialize from retained
epidermal structures in the rete ridges, hair follicles,
and sweat glands in 7 to 14 days. After healing, these
burns may have some slight skin discoloration over
the long term. Deep dermal burns into the reticular
dermis appear more pale and mottled, do not blanch
to touch, but remain painful to pinprick. These burns
heal in 14 to 35 days by re-epithelialization from hair
follicles and sweat gland keratinocytes, often with severe
scarring as a result of the loss of dermis. The other
degrees of burns are more severe and usually require
hospitalization.
Burn wound healing is a complex process with three
sequential phases: inflammation, proliferation and
tissue formation, and tissue remodelling. The phase
of tissue remodelling extends for weeks and months
after closure of the lesion, and the fibroblast is the
cornerstone of remodelling of the dermis, vessels,
and basement membrane. Usually, in second-degree
burns only a wound colonization is present, defined as
a multiplication of bacteria causing a delay in wound
healing, usually associated with an exacerbation of pain,
but still with no overt host reaction1.
Several topical therapeutic options have been used to
promote healing and to prevent bacterial colonization
to turn into a wound infection, which delays
healing. Among these, hyaluronic acid and silver
sulfadiazine show a well-established pharmacological
profile.
Since its introduction into clinical practice in 1967
by Charles Fox Jr.2, silver sulfadiazine (SSD) has been
the gold standard for topical burn therapy. Sulfadiazine
exerts its bacteriostatic effect by acting on the cell
membranes of micro-organisms, while silver nitrate
exerts its effect on the endocellular structures3. Applied
on exudative wounds, silver sulfadiazine releases its
silver ions, which increase the bacteriostatic effect of the
sulfonamide radical4. Indeed, the topical combination
of silver with absorptive dressings has led to new
therapeutic options for increased bacterial burden in the
surface wound compartment5, yielding a very low rate
1236 Hyaluronic acid plus silver sulfadiazine in second degree burns
of wound colonization and no secondary infection after
local application of a silver sulfadiazine-impregnated
lipidocolloid wound dressing to treat second-degree
burns6.
The addition to SSD of hyaluronic acid (HA), which
forms a substantial part of the human tissue intercellular
matrix, was aimed at overcoming one of very few
disadvantages of SSD, i.e. prolongation of the wound re-
epithelialization process. Recent observations suggested
that the impairment of dermal regeneration and
decreased mechanical strength of dermal tissue resulted
from the cytotoxic effect of SSD on dermal cells7. This
delay in wound healing was reversed by treatment with
epidermal growth factor8.
HA has important mechanical and structural functions
and also plays a key role in wound-healing processes. This
compound triggers macrophagic responses and induces
neoangiogenesis in injured tissues. HA stimulates fibrin
development and the phagocytic mobility and activity
of neutrophil granulocytes and macrophages, and it
increases the release of chemotactic factors for fibroblast
attraction9. Its clinical use for the topical treatment
of slow-recovery ulcerative lesions, such as burns and
bedsores, was well documented10–11.
Since both hyaluronic acid and silver sulfadiazine have
been used in therapy for decades and their efficacy is
well documented, a topical treatment combining these
two agents was formulated. The aim of this multicenter,
randomized, double-blind clinical study was to assess
the efficacy and safety of a fixed combination topical
medicinal product containing 1% silver sulfadiazine and
0.2% hyaluronic acid versus a 1% silver sulfadiazine-
containing cream alone, in the treatment of second
degree burns.
Methods
Study design
This was a multinational, multicenter, randomized,
double-blind, controlled, parallel-group study, comparing
a fixed combination cream containing 0.2% hyaluronic
acid and 1% silver sulfadiazine (HA-SSD) (Connettivina
Plus cream, Fidia Farmaceutici SpA, Abano Terme
(PD), Italy) versus a 1% silver sulfadiazine cream (SSD).
Clinical centers were located in France (two centers,
including the principal investigator), Croatia, Slovenia,
and Germany (one center in each country).
The recruited patients were randomly allocated to one
of the two treatment groups according to a computer-
generated randomization list. To fulfill blindness, the two
cream formulations tested were provided in unmarked
white tubes containing 25 g of cream each. The two
products had the same appearance, colour, and consist-
© 2005 LIBRAPHARM LTD – Curr Med Res Opin 2005; 21(8)
ency. In each patient, after an accurate debridement
and cleaning of the burn wound with appropriate
antibacterial solutions (0.25% sodium hypochlorite
solution or amuchina or chlorexidine), approximately
5 g/100 cm2 of cream was applied topically with a sterile
spatula. Wounds were then covered with sterile line
gauze which was fixed with bandages. Treatments were
applied once a day until the wounds healed, but for no
longer than four weeks. Prior to each new application,
the residue of the previous application was carefully
removed from the wounds using sterile saline solution.
Concomitant treatment with topical and systemic
corticosteroids, immunosuppressants, and drugs known
to interfere with the re-epithelialization process was
not allowed. Analgesics, other than acetylsalicylic acid,
were administered on an as-needed basis and their use
was recorded. Antibiotic therapy was not permitted,
except in the case of specific medical needs and their
use was recorded. Other routine medical treatments
for concomitant diseases were allowed and accurately
recorded.
Ethics
Written informed consent was obtained from each
patient. The study was conducted in accordance
with the Declaration of Helsinki and the general
principles of Good Clinical Practices for trials on
medicinal products in the EEC. The trial protocol and
subsequent amendments were reviewed and approved
by independent ethics committees and/or institutional
review boards, according to local legislation.
Patients
One hundred and eleven male (n = 70) and female
(n = 41) patients took part in this study. The patients,
aged 18 to 75 years, had IIa-degree (superficial) and
IIb-degree (deep dermal) burns not exceeding 5% of the
body surface area, corresponding to about 900 cm2 in an
average adult patient. The burned area was evaluated
by digitalized macrophotography. Burns had to have
occurred within 48 hours from the start of treatment,
be caused by thermal injury, and be confined to the
trunk and/or upper and lower extremities. Physical
examination and medical history were expected to
be normal, as was the laboratory screening, with the
exception of burn-related findings. The following
exclusion criteria were used: presence of cardiovascular,
hematological, endocrinological, gastro intestinal,
hepatic, or renal disease; disease of the CNS; long-term
treatment due to other pathological findings; pregnancy;
hypersensitivity to SDD or HA; or a history of drug and
alcohol abuse.
© 2005 LIBRAPHARM LTD – Curr Med Res Opin 2005; 21(8) Hyaluronic acid plus silver sulfadiazine in second degree burns Costagliola & Agrosì
Assessment of efficacy
Efficacy criteria
Clinical evaluations were performed by suitably trained
and qualified staff surgeons at the participating centers.
After an initial evaluation prior to the start of treatment
(Day 0), clinical signs were assessed weekly for the
entire duration of the treatment period.
The primary efficacy endpoint was evolution of
healing (complete/incomplete) expressed as the total
time in days required for complete healing of the wound.
According to the study protocol, treatment was stopped
when complete healing of the wound had occurred, i.e.
with 100% epithelialization and 0% residual wound area,
and the day upon which this occurred was recorded
as ‘time to healing’. Other parameters included
consumption of analgesics (number of tablets and days
of use), pain [assessed by the investigator by means of a
visual analog scale (Huskisson scale; 0–10 cm)], itching,
impairment to movement, and global comfort (these
latter described by means of frequency distributions
visit by visit).
Adverse events
A clinical examination and screening for adverse events
were both carried out at each visit.
Laboratory tests
Plasma biochemistry tests were carried out at Day 0 and
at the final day of the study for each patient. Hemat-
ological testing included RBC, hemoglobin, haematocrit,
WBC, and platelets. Biochemical testing included
glucose, uric acid, urea, creatinine, total proteins, total
bilirubin, AST, ALD, GGT, and alkaline phosphatase.
Urinalysis was also performed.
Statistical methods
Time to healing was analyzed by means of the incidence
rate of the event, with median time and, if it existed,
inter-quartile range. Statistical analysis for this endpoint
was made by comparing the two treatment groups by
means of survival analysis with the Wilcoxon test.
Consumption of analgesics, other than acetylsalicylic
acid, was evaluated by a qualitative approach (Fisher’s
exact test). Pain was analyzed in a descriptive way
(mean, standard deviation, min, max, Q1, Q3, median)
visit by visit. The change from baseline was also evaluated
giving the 95% confidence intervals. At baseline visit,
the correlation degree between pain of the last 48
hours and pain emerged during the medication, was
computed (Pearson’s correlation coefficient). Changes
1237
from baseline for itching, impairment to movement, and
global comfort were analyzed with Fisher’s exact test.
All statistical tests for efficacy were two-sided, with
alpha level = 0.05.
Results
Patient characteristics
Fifty-six patients were randomly allocated to HA-SSD
and 55 to SSD. One male patient in the SSD group
was lost at follow-up, thus a total of 110 patients
were evaluated. There were no statistically significant
differences between the two groups in subjects’ age
(38.2 ± 12.4 years in the HA-SSD group; 38.5 ± 13.7
years in the SSD group ), sex (33 males and 23 females
in the HA-SSD group; 36 males and 18 females in the
SSD group), and body weight (71.8 ± 10.9 kg in the
HA-SSD group; 74.5 ± 12.8 kg in the SSD group). The
average time elapsed from the thermal injury was similar
in the two treatment groups: 17 hours 8 min for the
HA-SSD group vs. 16 hours 43 min for the SSD group.
The average burn area (SD) at baseline was slightly
greater for the HA-SSD group: 97.4 cm2 (100.7) vs.
91.4 cm2 (55.9) for the SSD group (Table 1). 39.13%
(n = 9) of the patients in the HA-SSD group had a
burn area ≤ 50 cm2, 26.09% (n = 6) between 51 and
100 cm2, 21.74% (n = 5) between 101 and 150 cm2, and
13.04% (n = 3) > 150 cm2, respectively. These figures
were 26.67% (n = 8), 26.67% (n = 8), 30.00% (n = 9),
and 16.67% (n = 5) in the SSD group, respectively. In
91.07% (n = 51) and in 96.30% (n = 52) there was a
single burn lesion in the HA-SSD group and the SSD
group, respectively.
No relevant differences between treatment groups
was observed about the medical/surgical history and
physical examinations. The itching was slightly less
severe in the HA-SSD group with 87.5% of class
Absent–Mild (12.5% in the class Moderate–Severe)
compared to the 81.5% in the SSD group (18.5% in the
class Moderate–Severe).
The same result was observed for the impairment
of movement with 73.2% in the class No–Mild (26.8%
in the class Moderate–Severe) for the HA-SSD group
compared to the 63% in the SSD group (37% in the class
Moderate–Severe). The difference in global comfort
was less evident: 76.8% in the class Excellent–Good
(23.2% in the class Bad–Very bad) for the HA-SSD
group compared to the 75.9% in the SSD group (24.1%
in the class Bad–Very bad).
The maximum allowed period of treatment (28 days)
was needed in only one patient in the HA-SSD group,
while it was required for five patients in the SSD group.
In both treatment groups, 53 patients achieved the
therapeutic effect within the study period and therefore
could discontinue the treatment.

Table 1. Demographic and pre-treatment patient characteristics of treated patient groups

Parameter GROUP HA-SSD GROUP SSD

Sex (%)
males 58.9% (n = 33) 66.7% (n = 36)
females 41.1% (n = 23) 33.3% (n = 18)
Age (years)
mean ± SD 38.2 ± 12.4 (range = 19–62) 38.5 ± 15.1 (range = 18–75)
Body weight (kg)
mean ± SD 71.8 ± 11.0 (range = 51–93) 74.5 ± 12.8 (range = 52–112)
Height (cm)
mean ± SD 172.4 ± 8.0 (range = 159–187) 173.1 ± 8.2 (range = 157–192)
Lifestyle (percentage of patients)
smoker 32.1% (n = 18) 37.0% (n = 20)
coffee/tea drinker 55.4% (n = 31) 42.6% (n = 23)
alcohol drinker 16.7% (n = 9 16.6% (n = 9)
Burn characteristics
mean ± SD
time elapsed since injury (h:min) 17:08 ± 12:35 (range 1–46 hours) 16:43 ± 12:29 (range 1–47 hours)
area (cm2) 97.3 ± 100.7 (range = 6.39–426.26) 91. 4 ± 55.9 (range = 7.97–228.29)
Percentage of patients
location: arms 56.4% (n = 31) 68.5% (n = 37)
location: trunk 16.4% (n = 9) 3.7% (n = 2)
location: legs 27.3% (n = 15) 27.8% (n = 15)
presence of blisters 46.4% (n = 26) 44.4% (n = 24)
presence of exudate 83.9% (n = 47) 90.6% (n = 48)

1238 Hyaluronic acid plus silver sulfadiazine in second degree burns © 2005 LIBRAPHARM LTD – Curr Med Res Opin 2005; 21(8)
Efficacy
Healing
The burn lesions of all patients in both groups were healed
at the end of the study, except for one patient in the SSD
group. Comparison between treatment groups showed a
marked difference in time to healing in favor of the fixed
combination cream. Survival analysis showed that median
time to healing was 9.5 days (95% CI 7,14) in the HA-SSD
group vs. 14 days (95% CI 13,14) in the SSD group (a
difference of 4.5 days, p-value = 0.0073) (Figure 1).
Rescue medication
The mean number of tablets and the mean days of use of
rescue medications (analgesics other than acetylsalicylic
acid) administered during the study were greater in the
SSD group than in the HA-SSD group (0.69 ± 1.08 vs.
0.57 ± 0.89; 4.0 ± 6.1 vs. 2.4 ± 3.9, respectively); however,
this difference was not statistically significant (Fisher’s
exact test: p = 0.5830 and p = 0.1764, respectively).
Pain evaluation
Pain was scored between mild to moderate in severity
at baseline in all patients. This symptom decreased
very rapidly after the first week of treatment, with no
statistically significant difference between groups (95%
confidence interval –0.43,1.48).
Itching, impairment to movement and global
comfort
No statistically significant differences between treat-
ments were observed (Fisher’s exact test: p = 0.2701
and p = 0.3551, respectively) between the start and the
end of the treatment period.
Adverse events
Only one patient in the SSD group showed adverse
events (shivering, fever, and headache) whose
relationship with the drug were judged as possible.
There were no important changes in vital signs or in
laboratory evaluations from baseline.
No statistically significant difference between treat-
ment groups was recorded in the physician’s judgment
of local and systemic tolerability (Fisher’s exact test:
p = 0.2054 and p = 0.3122, respectively).
Discussion
In this clinical trial, the comparative evaluation of a topically
applied fixed combination of silver sulfadiazine and
© 2005 LIBRAPHARM LTD – Curr Med Res Opin 2005; 21(8) Hyaluronic acid plus silver sulfadiazine in second degree burns Costagliola & Agrosì
Probability of survival
Survival time (days)
Figure 1. Time to complete healing for two patient
treatment groups with IIa- and IIb-degree burns treated
with topically applied creams. HA-SSD: fixed combination
of silver sulfadiazine plus hyaluronic acid (n = 55)
versus SSD: silver sulfadiazine (n = 54). Kaplan-Meier
cumulative survival curve
hyaluronic acid with silver sulfadiazine alone demonstrated
a statistically significant shortening of the length of time-
to-healing in IIa- and IIb-degree burns. This parameter
is a major indicator of a successful therapy of second-
degree burns, allowing the comparison of potential risks
owing to late re-epithelialization of the burn wound. In
addition to this finding, the study results showed that the
fixed combination product was a safe topical medication,
well tolerated at both the systemic and local level. No
important adverse effects, such as painful application or
allergic reactions, occurred during the study.
Results obtained in this clinical study, which confirm
those previously obtained in a recent randomized,
double-blind study in 33 patients with burns with similar
characteristics12, and in a previous trial10, favor the use
of this combination product and highlight the response-
enhancing properties of hyaluronic acid when topically
applied to these patients. The significant difference
produced by the addition of hyaluronic acid to silver
sulfadiazine is consistent with the rationale supporting
the use of hyaluronic acid to stimulate granulation tissue
formation and wound healing13.
Conclusion
In conclusion, the fixed combination of HA plus SSD
and SSD alone were well tolerated and effective
topical treatments for second-degree burns, but the
fixed combination promoted significantly faster healing
of the burn than SSD alone and thus may enrich the
assortment of topical medications available for the
treatment of burns.
1239

Acknowledgements
Declaration of interest: Study support was provided
by Fidia Farmaceutici (Abano Terme, Italy)
The authors wish to thank A. Maria Menegus (Fidia
Farmaceutici, Italy) for data management.
Study investigators and centers
Principal investigator: Prof. M. Costagliola, Department
of Plastic and Reconstructive Surgery, Centre Hospitalier
Universitaire, Toulouse Rangueil, France.
Co-investigators: Prof. Alfred Berger, Klinik f.
Plastische, Hand-und Wiederherstellungschirurgie,
Hannover, Germany; Prof. Tomaz Janezic, Department
of Plastic Surgery and Burns, Ljubljana, Slovenia; Prof.
Zdenko Stanec, Dubrava University Ospital, Zagreb,
Croatia; Prof. Daniel Wassermann, Service des Brûlès-
Hôpital Cochin–Paris, Paris, France.
References
1. Weed RO, Berens DE. Basics of burn injury: implications for
case management and life care planning. Lippincotts Case Manag
2005; Jan-Feb;10:22-9
2. Fox CL Jr. Silver sulfadiazine – a new topical therapy for
Pseudomonas in burns. Therapy of Pseudomonas infection in
burns. Arch Surg 1968;Feb;96:184-8
3. Akahane T, Tsukada S. Electron-microscopic observation on
silver deposition in burn wounds treated with silver sulphadiazine
cream. Burns Incl Therm Inj 1982;Mar;8:271-3
4. Hoffmann, S. Silver sulfadiazine: an antibacterial agent for
topical use in burns. A review of the literature. Scand J Plast
Reconstr Surg 1984;18:119-26
5. Sibbald RG, Orsted H, Schultz GS, Coutts P, Keast D.
International Wound Bed Preparation Advisory Board; Canadian
Chronic Wound Advisory Board. Preparing the wound bed 2003:
focus on infection and inflammation. Ostomy Wound Manage
2003;49:23-51
6. Carsin H, Wassermann D, Pannier M, Dumas R, Bohbot
S. A silver sulphadiazine-impregnated lipidocolloid wound
dressing to treat second-degree burns. J Wound Care 2004;13:
145-8
7. Lee AR, Moon HK. Effect of topically applied silver sulfadiazine
on fibroblast cell proliferation and biomechanical properties of
the wound. Arch Pharm Res 2003;26:855-60
8. Cho Lee AR, Leem H, Lee J, Park KC. Reversal of silver sulfa-
diazine-impaired wound healing by epidermal growth factor.
Biomaterials 2005;Aug;26:4670-6
9. Chen WYJ, Abatangelo G. Functions of hyaluronan in wound
repair. Wound Repair Reg 1999;7:79-89
10. Baux S. Etude clinique de l’activité et de la tolérance de Ialuset®
Plus dans le traitement des brûlures. Etude comparative (vs
sulfadiazine argentique), randomisée et multicentrique. Brûlures
2004;4:230-6
11. Brown JA. The role of hyaluronic acid in wound healing’s prolifer-
ative phase. J Wound Care 2004;13:48-51
12. Koller J. Topical treatment of partial thickness burns by silver
sulfadiazine plus hyaluronic acid compared to silver sulfadiazine
alone: a double-blind, clinical study. Drugs Exptl Clin Res
2004;30:183-90
13. Hadjiiski OG, Lesseva MI. Comparison of four drugs for local
treatment of burn wounds. Eur J Emerg Med 1999;6:41-7

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Paper CMRO-2970_2, Accepted for publication: 28 June 2005
Published Online: 00 July 2005
doi:10.1185/030079905X56510

1240 Hyaluronic acid plus silver sulfadiazine in second degree burns © 2005 LIBRAPHARM LTD – Curr Med Res Opin 2005; 21(8)