CONTRACEPTION

O-YEAR FOLLOW-UP OF CHANGES IN CLINICAL CHEMISTRY 'IN SIN POREAN NORPLAN ACCEP RS: M BOLIC C NGES

Dr Kul d ip Si ngh ~BS (S'pore), ED (S'pore), MRCOG (Lond)

MA Popu ]a t lon Research with Di s t i nctlon (Exon), AId

Dr 0 A C Viegas ChB, , ~COG, (B'ham)

Mrs Diana Liew Cl in ica l Biochemist

B Sc

Mr Piara. Singh Biostat i s t i ca l Of f i ce r

B (Hon) (Can)

Professor S S Ratnam aS, , FRCS, FRCS Ed, FRCSG, F CS,

FRCOG, FRACOG (Hon), S (Hon)

Department of Obstetrics and Gynaecology Nat i ona~ tin i vers i of S i ngapore

National Univers.~,:. "~sp l ta l Lo r Kent F- -- Road

Si ngapo- : : .i

In th i s longitudlna] study involving 100 Singaporean acceptors, the e f f e~s of N0rplant ® on metabolic function was evaluated. With respect to l i v e r nction, the results at the end of two years s t i l l ind ic e the presence o f possible hepatocel lular dysfunction. Despite a 4.5~; decrease in serum b l ] i r u b i n during the second year, the mean serum bl I i rubi n st i ] ] remained sl gn i f i cant l y ra i sed (p < O. 001) Furthe ore there s a s ign i f i can t f a l l in to ta l proteins and globul in during t period under review (p < 0.001). However mean levels of a ] ] three parameters remained wlthin the normal cl i n i ca l range.

As regards l i p i d metabolism, the to ta l t r i g l yce r ides and to ta l chol esterol s t i 1 ] remai ned S ign i f i can t l y decreased at the end of ~ two years (p < 0.001). The LDL-choIesterol, which was s i g n i f i c a n t l y decreased at the end of one year, i ncreas by 2. during the second year. However, t h i s r i se i n L-cho I esterol s not s i gn i f i can t and was s t i l l below the prein~3rt lon leve]. The L-cho]esterol levels r~urned to t he i r preinsert ion ] eve] s a f te r a s ign i f i can t increase in the f i r s t year. The L-chol esterol /Total chole~ero l - HDL-cholestero] ra t i o returned to i t s pre inse~ion value at the end of two ars whi]e the LDL-cholesterol/HDL-cho]estero] r a t i o s s t i l l s ign i f i can t ]y lower than the preinsert ion ~atio (p < 0.05). As we have

Submitted for pub] i ca t ion October 21, 1988 Accepted f o r p u b l i c a t i o n November 29, 1988

FEBRUARY 19 VOL. 39 NO. 2 I

C O N T R A C E P T I O N

not incorporated a simultaneous non-steroid treated control group, the observed changes in 11 poprotein I ipids could be a t t r ibu ted to extraneous factors such as d ie t , exercise or other environmenta] changes. Nevertheless, the f indings at the end of two years, though speculative, s t i l l indicate Norplant® use to be non-contributory to cardiovascular r i sk and indeed i t may be a protect ive Factor against such r isks in th i s population. lhe use of Norplant® was not associated wit.h any s ign i f i can t e f fec t on glucose tolerance a f te r two years of use.

I ION

The No~lant ® system (subdermal implants) is one of the most modern ho nal f e r t i l i t y regulat ing methods, that is Currently undergoing c l i n i c a l t r i a l s at F e r t i l i Control C l in i c , National Universi ty Hospital, Singapore. Given the long-acting nature of Norplant® and the potent ial for i t s widespread use, i t is of pa r t i cu la r importance to assess i t s ef fects on I ip ld , and other metaboiic functions in acceptors from d i f f e ren t population subsets. Much of the information avai lable to date has been in Caucasians from developed countries in whom disease patterns and r isk factors fo r disease d i f f e r substant ia l ly from those in developing countries. Thus, i t was decided to study the effects of Norplant® on metabolic function in a representative South-East Asian population as part of the long-term survei l lance of th i s r e l a t i v e l y new contraceptive.

The f i r s t year experience in Singapore was presented in 19 (1). In th i s paper the results the end of t years are presented.

IALS S

One-hundred women we recrui ted for th i s study from the attendees of the Fert i I i t y Control CI i nic at the National Universi ty Hospltal, Singapore. These volunteers were healthy, non-smoking, non-alcohol dr i nkers and non- 1 a t ta r i ng women who had chosen No rp I an t ® fo r contracept ive purposes. Cl i nical assess nt and blood sampl ing fo r the study were done pr io r to inse ion of the implants and a f te r 6, 12 and 24 months of use. The subjects were used as t h e i r own controls. Blood samples were obtained from the r igh t antecubital vein at the same time of day (9 am to I I am)on each occasion a f te r an overnight fast .

Blood samples were taken f o r glucose (glucose oxidase meth ) (2), to ta l protein (Blur method) (3), albumin (BCG method) (4), tota I b ~ 1 i rub in ( 5), and a I ka I i ne phosphatase (6) ( i f ied Bessey-Lo Brock method) on the ASTRA-8. HT)L-chol esterol was prec ip i ta t by gnesium phosphotungstate (7) . Cholesterol (8,9) (C P-PAP method) and t r i g l yce r ides (10) (enz a t i c hydrolysis and determinants) were dete in by k i t methods from Boehringer-Manheim. LDL-cholesterol was calculated'using the method emp]oyed by Friedewald et al. (11)(LDL-cho]esterol = Tota] cholesterol - (HDL-cholesterol +

lycer l.des . . . . 1/1 ), 2 . 1 9

1 F E B R U A R Y 1 VOL. NO. 2

CONTRACEPTION

Quality control in l lp ld assays were monitored by running 2 serum control pools co r c i a l l y available (normal and high levels) in evePy assay. For cholesterol, CV between assays at both normal (about 3 o1/ I ) and high (about 8 o1/1) was less than Z~ while fo r t r l g l ycer ides , at the normal level (about 2 o i / I ) , CV was about 3~ and at the higher l i m i t (about 8 I / I ) was about 5.5%. External qua l i t y control s assessed by an internat ional qual i ty control sch e ( Internat lonal External Quality Assessment Scheme or IEQAS) conducted by O Collaboratlng Centre fo r Research and Reference Servlces In Cl in ica l Ch Is t ry , Birmingham, UK.

To assess the signif icance of differences from mean values~ the paired ' t ' tes t was used. Further comparisons were also made by observing percentage changes in mean post- insert ion level s c par to preinsert lon levels which were designated as I00%. Where d i s t r i bu t l on of the results was skewed, the Wilcoxon signed rank test of s t a t i s t i c a l signif icance us ing median d is t r i bu t ion was used.

R ULTS

(a) Liver Funct ion (Table I )

an b i l i r u b i n levels, which had increased s ign i f i can t l y by about 5 in the f i r s t year, showed a f a l l of about 4.5% in the second year of use However, the value at 0.85 /d] at the end of the second year was s t i l ] highly s ign i f i can t ] y raised when compared to the preinsert lon mean. There was a s t a t i s t i c a l l y s ign i f i can t f a l l in to ta l proteins and g]obulins over the two years. The albumin and a lkal ine phosphatase showed no s ign i f i can t changes a f te r 2 years of use. I t is important to note that no individua] values f e ] ] outside the normal range.

(b) Lip|d Hetabolism (Table I I )

Total t r ig !ycer ides and to ta l cholesterol s t i l l remained highly s i gn i f i can t l y decreased at the end of two )-ears of use. H ever there was no s ign i f i can t change between the values at the end of the f i r s t and second years. Simi lar ]y the LDL-cholesterol values was s t i ] ] s i gn i f i can t l y ]ower than the preinsert ion value. However during the second year, there was a 2.2% increase in the mean L-cho]esterol level but th i s increase was not s ign i f i can t . The L-cholesterol leve]s which showed a s ign i f i cant r ise during the f i r s t year of use returned to preinsert ion levels by the end of year 2.

The L-cholesterol/Total cholesterol - L-chol esterol r a t l o, which had shown a ~ ign i f i can t increase durlng the f i r s t year of use, returned a l st to i t s preinsert ion value at the end of 2 years, have a I so cons i der the L-cho I e stero I to

L-cholesterol ra t i o since th is is a more pe~inent ra t i o The L-cholesterol / L-cholesterol r a t i e sho a 9~ increase during

the second year of use a f te r having shown a hlghly s ign i f i can t decrease of 19. in the f i r s t year. Despite th i s , the ra t io was s t i l l s i g n i f i c a n t l y lower as c pared to the preinsert lon value.

FEBRUARY 1 VOL. NO. 2 131

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LE I . BIO I L ES IN SI P A E S: t.IV i TES

Parameters Studied

Total Protein

n

mean ± SD (g/cl l ) P value

% di f ference from pre- inser t i on mean

Albumin

n

mean ± SD (g/d l ) P value

d i f ference from pre- i n sert i on mean

Globulin

n

mean ± SO (g /d l ) P value

% di f ference from pre- i n ser t i on mean

Bi I i rub in

n

mean ± SD (mg/dl) P value

d i f ference from pre- inser t ion mean

Pre- Inser t i on Af ter

1 year of use

97 7.24 ± 0 .37

I00

96 7.08 ± 0.69

< 0.05

97 4.ZZ ± 0.22

100

96 4.Z7 ± 0.42 < 0.05

101

97 3 .03 ± 0 .35

I00

96 Z.81 ± 0.45

< 0.001

93

97 0.55 ± 0.19

100

96 0.87 ± 0.31

< 0.001

158

A1 ka I i ne Phosphatase

n mean ± SO ( i u / l )

P value % di f ference from pre- i n sert i on mean

97 61.1 • 17 .3

100

96 60.3 ± 15.7

NS

99

A f te r 2 years of use

76 6 .87 ± 0 .43

< 0 .001

98

76 4.24 ± 0.24

< 0.001

101

76 2 .63 ± 0 .35

< 0 .001

76 0 .85 ± 0 . 2 8

< 0 . 0 0 1

150

76 61.2 ± 14.7

NS

99

1 F E B R U A R Y 1989 V O L . 39 N O . 2

C O N T R A C E P T I O N

TPgLE I I . BI EI41CAL S IN S I P E S- LIPID LI

Parameters Stud led

Tr ig lycer ides

n

mean ± SO ( o l / l ) P value

% di f ference from pre - inser t i on mean

Cho I estero I

n ̧

mean ± SO ( o l l l ) P value

% di f ference from pre- inser t ion mean

HDL-Cho I ester01

n

mean ± SD ( o 1 / 1 ) P value

% di f ference from pre- inser t ion mean

LDL-Cho I ester01

n

mean ± SD ( 0111) P v a l u e

% di ffccence from pre - inser t ion mean

HDL-Cho I estero ] /Tota ] - Cholesterol -

L- Cho ] esterol Ratio

n

mean ± SD P value

% di f ference from pre- i nsert i on mean

~L-Cho I estero ] / L- Cholestero] Rat i o

n mean ± SD

P value % di f ference from pre - i nsert i on mean

Pre- lnser t ion I I year of l, sel 2 years of use

97 1 .21 ± 0 .74

100

97 5 .05 ± 1 .02

100

97 1.08 ± 0 . 3 4

100

97 3 . 3 8 ± 0 . 9 9

100

97 0 . 3 0 8 +_ 0 .164

100

97 3 . 5 6 _+ 1 . 7 3

100

96 0 . 8 9 ± 0 .45

< O. 0 0 1

74

76 0 .90 ± 0 .52

< 0 .001

76

96 4 . 6 4 ± 0.81

< 0.001

92

96 1 . 1 3 ± 0 .24

< 0 . 0 i

105

96 3 . 1 0 ± 0.72

< 0.001

92

96 0 .342 +_ 0 . 1 1 1

< 0 . 0 0 1

111

96 2.85 ± 0.90

< O. 001

81

76 4 . 7 3 + 0 .83

< 0 .001

94

76 1.08 ± 0 . 2 3

NS

98

76 3 . 2 5 ± 0 . 7 3

< 0 .05

95

76 0 .311 ± 0 .0

NS

101

76 3 .17 ± 1 .13

< 0 . 0 5

9 1

F E B R U A R Y 1 V O L . 39 NO. 2 133

CONTRACEPTION

(c) Glucose Tolerzlnce (Table I l l )

Oral glucose tolerance tests using 50 gm loads were perfo ed before and af ter 1 and Z years of Norpl an t® use . No c l i n i c a l l y s ign i f icant changes were observed. Differences in the one hour level did reach s ta t i s t i ca l significance at the end of two years when caIcul ated on a paired basis; however, the i r a b.~lute level s remained within the normal range.

BLE I I I . ORAL GLUCOSE TOLERANCE ST C NGES IN SIN PO N HORP ACCEP1FORS

i ............................ I 1 hour ............................................... Z ....... -hour-s-

............................ n --~-~97 ............................................................................ J ................................... I 131.3 ± 3Z.1 I 112.7 ± 27.6 Glucose 0 year(pre-insert ion) I 89.9 ± 9.9

n = 97 1 144.0 ± 34.0 I 110.4 ± Z8.Z G l u c o s e 1 y e a r a f t e r u s e I 9 1 . 5 ± 9 . 8 ......................... p v,.1._,e ................ i ........... ..... ,0.00 I Glucose 2 years af ter use 9.1 ± 10 .4 143.8 ± 35.5 112.1 ± 29.8

P value NS < O. 001 1 NS ....................... ........... - - , , , ~ _ , , ........... _~:-- _-... .. = = - ~ -:..~...m ...... ,, ~--- ~: ~ - .~ ~ . J

Ol SSl

Specific studies on Norplant® and i ts effect on l i p i d metabolism have y i e] ded i ncon s i s(:ent resu ] ts However, d i f fe ren t methodologies, techniques of qual l ty control, study populations and varying r isk factors might account for some of these inconsistencies. I t is for these reasons at studies are bei ng undertaken i n d i f fe ren t

pu ] at ions to c I ar i fy th is i ssue.

The highly s ign i f icant decrease in total tr iglyce?ides and to ta l cholesterol seen in the f i r s t year remained in the second year though the mean values did not decrease further. This is compatible with the results of other investigators (12-14). Similar ly the LDL-cholesterol which decreas s ign i f i cant ly during the f i r s t year of use, showed a 2.2~; increase in the second year. However despite th is , the

L-cholesterol values at the end of 2 years were st i I 1 s ign i f i can t l y lower than the preinsertion values. The HDL-cholestero] levels returned to almost preinsertion values af ter having risen s ign i f i can t ly in the f i r s t year.

I t is known t h a t a h igh H D L - c h o l e s t e r o l / T o t a l c h o l e s t e r o l - HDL- c h o l e s t e I r a t i o and a low L - c h o l e s t e r o l / . ~ O L - c h o l e s t e r o l r a t i o i s p r o t e c t i v e a g a i n s t rd iovascu~.ar compl i ions ( 1 1 ) . In t h i s s tudy we have shown t h a t t he L - cho l e s t e r o l / T o t a l ~ chol e s t e r o ] - L -cho l e s t e r o l r e t u r n e d t o a l s t i t s preinser t ion v~ lue a t the end o f 2 yea rs a f t e r having increased significantly during the first year. The LDL-cho lestero 1/~L-cho I estero I ra t io showed a highly s igni f icant decrease in the f i r s t year of Norpiant® use. During the second year, there was a increase in th is rat io . Despite th is increase, i t is reassuring to know th the ra t io was s t i l l s ign i f ica ly I er

1 FEBRUARY 1 VOL. NO. 2

CONTRACEPTION

as c pared to the preinsert ion ra t io . Thus, i t would appear that at least in Si~gapor~an women the effects of No~-plant ® on l i p i d metabolism a~ non-contributory to cardiovascular r isk . Indeed the resul ts suggest that i t may be a protect ive fa or against cardiovascular c p l icat ions. We must add that in the absence of non-steroid treated controls in the study, the observed changes could be at t r ibuted to extraneous environ ntal effects.

Most studies to da h a v e shown no s ign l f i can t changes in l l ve r function with Norp] ant® use (12,15,16). Shaaban f r Egypt show only a sinai] increase in b i ] i r ub in which reverted to normal by 6 nths. In our study have seen a highly s ign i f i can t 5 r ise in b i l i r u b i n during the f i r s t year of use (1). There was only a 4.5~ decrease in b i l i bin in the second year and as such the levels are s t i l l s i gn i f i can t l y raised as compared to preinsert lon levels. Total protein and globulins continued to f a ] l in the second year and th is f a ] ] r ined s t a t i s t i c a l ] y high]y s ign i f i can t . Serum albumin and alkal ine phosphatase showed no change a f te r 2 years of use. I t thus appears that at the end of two years of Norplant® use, there is s t i l l s degree of l i v e r dysfunction. However, despite the changes seen, the mean levels of b i l i r u b i n , to ta ] protein and globulin remained within the no a] c l i n i ca l range.

The ef fects of r-10rplant® on carbohydrate metabolism as re f l ec t by changes in the OGTT at the end of two years is minimal and no d i f fe ren t from those at the end of one year. The changes in the one hour glucose levels, although s t a t i s t i c a l l y s ign i f i can t , remain within the norma] range.

!

The f indings of th is study continues to B~phasise the compe11ing need fo r continued c] ose survei I lance of th i s rel a t i vely n cont ceptive system. We hope to make annual assessments of a l l parameters measu~"ed f the f ive,year duration of use of Norplant®.

We are gra fu] to FCmily Health International and The Population Council for i n t r ucing Norplant® to Singapore. are also in ted to t Department of Obstetrics and Gynaecology, National University of Singapore, fo r f inancia l support fo r the ent i re investigation. F ina l ly we would l i ke to thank Miss Pre for her s re tar ia l assistance in preparing th i s manuscript and the Norplan acceptors for the i r co-operat i on.

I.. Singh ~K, Viegas 0, Liew ' D, Singh P, R nam SS. The E f f e ~ s o f RPLA on Cl in ica l Ch i s t in Singaporean Acceptors a f te r 1

Year o f Use- M e t a b o l i c Changes. Con t racep t ion 38.79-89 (19 ) .

2. Cr . New Aut ted Method For asu r ing Glucose by Glucose. Oxidase. J - C l l n Patho] Z0-910-912 (1967) .

FEBRUARY 1 VOL. NO. 2 135

CON AC TION

Q

Q

O

6 ~

W

t

Q

10.

11.

12.

13.

14.

15.

16.

Weichselbaum TE: An Accurate and Rapid Determination of Proteins in Small P=mounts Plasma. J CI in Pathol 16-40-49 ( !946).

Method for the of Blood Serum and

Spencer K, Pr ice CP I nf ] uence of Reagent Qua I i ty and React i on Conditions on the Determination of Serum Albumin by the Bromocresol Green-Dye Binding Method. Ann Clin Biochem 14"105-115 ( 1977).

Jendrassik L, Grof P. Simplif ied Photometric Methods for the Determination of the Blood B i l i rub in . Biochemische Ze i tschr i f t 297-81-89 ( I 938).

Bessey OA, Lowry OH, Brock . A Method for the Rapid Dete ination of Alkaline Phosphatase with 5 Cubic Mill imetres of Serum. J Bi o] Chem i64- 321-329 (1946).

Burstein M, Scholnick HR, Morfin R. Rap id Method for the Zsolation of L ipoproteinsfr Human Serum by Precipitation with Polyanions. J Lipid Res 11 583-595 (1970).

Stahler F, Gruber W, Stinohoff K. E ine Praxi sgerechte Enz a t i c Cholesterin-Besti ng. Med Lab (Stuttg)30-29-37 (1977).

Trinder P. Determination of Glucose in Blood using Glucose Oxidase with an Alternative Oxygen Acceptor. Ann Cl in Biochem 6" 24-27 (1969)

h f ie ld . In- Bergm er, HU (ed). thods of Enz a t ic Analysis. 2nd English ed. Verlag Chemie Welnheim and Academic Press, New York and London" 4 vols; p 1831 (1974).

Fr i edewa ld WI, Hyper I i p i daema.

Levy RI, Fr rickson DS. Cl inic Chem 18:499-502 (1982).

Management of

Croxatto HB, Diaz S, Robertson DN, Pavez M. Women Treat~ with Levonorgestrel Implants I . Contraceptio~ 27-281-2 (1983).

C] inical Chemi in {Norp!ant®) or a TCU

Shaaban , Elwan SI, Abdallah SA, Darwish HA. Effect Subdermal Levonorgestre] Contraceptive Implants, Norplant®, Serum L i p iris. Contracept i on 30- 413-419 (1984).

of o n

Roberston DN. Norgestrel-releasing si last ic rods" Cl in ical Ef cts, Bioch ical Ef cts a ~ In Vivo Release Rates. In-Mishel] DR, J r , ed. Advances in Human Fert i I i and Repr u ive Endocrinology, Vol 2- Long Acting Steroid Contraception. Raven Press, New York, pp 127-147 (19~3).

Shaaban , El nt SI, EI-Sharka , Fargh AS. Effect Subdermal Levonorgestrel Contraceptive Implants, Norplant®, Liver Function. Contraception 30-407-412 (19 ).

of on

Croxatto , Diaz S, Pavez M. C] i nical Chemistry i n Women Treated with P gestogen Implants. Contraception 18-441-450 (19 ).

1 FEBRUARY 1 VOL. NO. 2