Diagnostic value of amino acid consumption test on exocrine pancreatic insufficiency

Abstract

Key Words: pancreatic insufficiency/diagnosis; aminoacid consumption test; cerulein cholelithiasis

INTRODUCTION

Amino acid consumption test (AACT) has a high sensitivity and specificity in evaluating exocrine pancreatic insufficiency[1,2], but its diagnostic value to exocrine pancreatic insufficiency in Chinese has not been well understood. In this study, the oral reagent stimulating pancreatic secretion (O-AACT) was used instead of cerulein (I-AACT) for amido acid consumption test and the dignostic efficiency of O-AACT was evaluated and compared with I-AACT on the exocrine pancreatic insufficiency in Chinese.

MATERIALS AND METHODS

General data

A total of 176 volunteers and patients, divided into three groups, were studied after giving written informed consent.

Group 1. Normal coutrols consisting of 12 males and 8 females, aged 20-51 years. None of them had evidence of digestive diseases, and were alcoholics.

Group 2. Cholelithiasis group consisting of 31 males and 45 females, aged 22-60 years, including 44 cases with gallstone, 5 with common bile duct stone and 27 with intrahepatic lithiasis. Those who had any evidence of pancreatitis, acute cholangitis, or diabetes mellitus had been excluded.

Group 3. Pancreatic disease group consisting of 49 males and 31 females, aged 23-71 years, including 28 cases of pancreatic cancer, 19 of peri-ampulla of vater tumor, 12 of chronic pancreatitis (CP) complicated with pancreatic cysts, 12 with common bile duct cyst complicated with CP, and 9 of CP. Those diagnoses were supported by the typical clinical manifestations and based on generally accepted morphologic alteration suggestive of pancreatic disease found in at least one of the following procedures: CT, surgery and/or pathology.

All patients received I-AACT and N-benzoyl-L-tyrosyl-para-aminobenzoic acid (purchased from Chongqing Medicine Industrial Laboratory) test (BT-PABA test). Among them, 12 in Group 1, 36 in Group 2 and 48 in Group 3, were also examined by O-AACT. Each test was made at an interval of two or three days. Gallbladder volume was determined by ultrasonography (US) at 0, 30, 45, 60, 90 and 120 min in 6 volunteers in Group 1 after drinking one of the three kinds of reagents stimulating pancreatic secretion at an interval of two days.

Methods

BT-PABA test This test was performed according to Imondi′s method[3]. The cut off limit for normal results was 45.0% of the orally administered dose.

I-AACT test According to Gullo′s method[2], the test was made in the morning after an overnight fast. Each of 176 subjects received a continuous intravenous infusion of cerulein (50 ng/kg·h) (Purchased from Sigma Com., USA.) for an hour dissolved in 0.9% NaCl solution. During each study blood samples for amino acid determination were taken before the infusion was started at 0 min and during the infusion at 45 min, 60 min and 30 min after the infusion was stopped. Total plasma amino acid was estimated by the sodium-β-naphthoquinone-4-sulfonate salt colorimetric method. The individual basal amino acid level was taken as 100% standard for final calculations, the relative reduction rate of the amino acid levels at each time was calculated. Decrease by over 14.0% in plasma amino acid concentration was used as an normal limit index of pancreatic function.

O-AACT Oral reagent stimulating pancreatic secretion was used instead of infusion cerulein in AACT, the other procedure was similar to I-AACT. The oral reagent stimulating pancreatic secretion consists mainly of:

A. Salad oil 1 mL/kg, 0.1 mol/L HCl 0.2 mL/kg, saccharose 0.5 g/kg added with water to 100 mL.

B. Salad oil 1.5 mL/kg, 0.1 mol/L HCl 0.2 mL/kg, saccharose 0.5 g/kg, added with water to 200 mL.

C. Salad oil 2.0 mL/kg, 0.1 mol/L, HCl 0.2 mL/kg, saccharose 0.5 g/kg added with water to 200 mL.

Gallbladder volume Gallbladder volume (Cllipsoid formula V = 0.52 × l × b × h)-was measured by US each time before and after drinking the reagent stimulating pancreatic secretion.

Statistical analyses were made using Chi-square test, and analysis of variance.

RESULTS

The change of plasma amino acid level in I-AACT

Basal amino acid (AA) levels of all subjects varied between 1.12 mmol/L and 6.8 mmol/L. During intravenous infusion of cerulein, the levels of plasma AA in all subjects of three groups gradually decreased, the 13, 45 and 5 cases had relative decline rate over 14.0% in Groups 1-3 at 45 min respectively, and 18, 68 and 11 cases at 60 min. Thereafter, plasma AA levels returned slowly toward initial AA values. The relative decline rate of plasma AA levels of 8 cases in Group 2 and 67 cases in Group 3 were still lower than 14.0% at 90 min.

The selection of oral reagent stimulating pancreatic secretion

When reagent A or B was selected, all volunteers, except two people who had slight flatulency, could drink all the reagent without discomfort. But after drinking a cup of reagent C, three volunteers had vomit and exited from the study, and gallbladder contraction occurred earlier in other three volunteers (Table 1). Reagent A was finally selected as the reagent stimulating pancreatic secretion in O-AACT because it caused steady gallbladder constriction.

Table 1 Gallbladder volume change after taking reagent (mean ± SD, mL).

Time	Reagent A n = 3	Reagent B n = 6	Reagent C n = 6
0 min	17.22 ± 3.55	16.58 ± 3.04	14.97 ± 5.23
30 min	12.85 ± 6.34	13.78 ± 2.69	8.87 ± 2.10
45 min	12.12 ± 2.25a	13.27 ± 2.96	8.70 ± 1.13
60 min	9.68 ± 3.30a	6.71 ± 2.66a	12.03 ± 3.05
90 min	6.28 ± 3.51a	4.93 ± 1.89a	13.97 ± 9.06
120 min	12.27 ± 5.42	11.40 ± 4.79	15.47 ± 5.94

^aP ＜ 0.05 in comparison with 0 min in the same group. Analys is of variance: F_A = 4.2799 (F₄₅ = 8.8133, F₆₀ = 14.4901, F₉₀ = 28.7727), F_B = 12.1610 (F₆₀ = 35.8538, F₉₀ = 63.6581)

The change of plasma amino acid level in O-AACT

During O-AACT, plasma AA levels gradually decreased, then returned slowly toward initial AA values. The change was similar to that in I-AACT (Table 2).

Table 2 The change of plasma amino acid level in O-AACT and I-AACT (mean ± SD, mmol/L).

AACT	Time (min)	Group 1 (n = 12)	Group 2 (n = 36)	Group 3 (n = 48)
I-AACT	0	4.23 ± 0.78	4.64 ± 1.49	3.91 ± 1.32
	45	3.13 ± 0.068a1	3.53 ± 1.49a6	3.65 ± 1.19
	60	2.68 ± 0.85a2	3.09 ± 1.21a7	3.49 ± 1.19
	90	3.67 ± 0.90	4.08 ± 1.47	3.66 ± 1.24
O-AACT	0	4.24 ± 1.19	4.67 ± 1.45	3.99 ± 1.26
	45	2.73 ± 0.99a3	4.05 ± 1.43	3.57 ± 1.24b1
	60	2.05 ± 0.94a4	3.01 ± 1.17a8	3.60 ± 1.20b2
	90	2.94 ± 1.12a5	3.84 ± 1.49a9	3.89 ± 1.29

^aP ＜ 0.05, in comparison with 0 min in the same group and the same method. Analysis of variance:

^a1F = 13.679,

^a2F = 21.45,

^a3F = 11.487,

^a4F = 25.095,

^a5F = 7.637,

^a6F = 9.938,

^a7F = 23.792,

^a8F = 28.622,

^a9F = 5.799. ^bP ＜ 0.05, in comparison with Group 1, analysis of variance:

^b1F = 4.746,

^b2F = 17.379.

Comparison of the diagnostic value of BT-PABA, O-AACT and I-AACT

Table 3 shows the number of cases of each group diagnosed as exocrine pancreatic insufficiency by BT-PABA, O-AACT or I-AACT. The diagnostic value of BT-PABA, O-AACT and I-AACT to exocrine pancreatic insufficiency is summarized in Table 4.

Table 3 The number of cases in each group diagnosed as exocrine pa ncreatic insufficiency.

%	Group 1	Group 2	Group 3
O-AACT	12	36	48
＞ 14.0%	11	30	13
14.0%	1	6	35a
I-AACT	20	76	80
＞ 14.0%	20	68	13
＞ 14.0%	0	8	67b
BT-PABA	20	76	80
＞ 45.0%	20	62	16
＞ 45.0%	0	14	64c

P ＜ 0.05 in comparison with groups 1 and 2. Chi-square test (χ²): a = 33.439, b = 102.208, c = 77.848.

Table 4 The diagnostic value of BT-PABA, O-AACT and I-AACT in exocrine pancreatic insufficiency.

	Sensitivity (%)	Specificity (%)	Accuracy (%)
O-AACT	72.9	85.4	79.2
I-AACT	83.8	91.7	88.1
BT-PABA	80.0	85.4	83.0

Chi-square test: P ＞ 0.05.

DISCUSSION

Exocrine pancreatic function test is generally divided into direct test (e.g., pancreozymin secretin test, cerulein test, etc.) and indirect test (pancreolauryl test, BT-PABA test, etc.). Although it is more sensitive and accurate in diagnosis of exocrine pancreatic insufficiency, the direct test is not widely used in clinical practice because it is complex, time consuming, and bringing more sufferings to patients. Indirect test is simple, but with low specificity and accuracy in diagnosis of exocrine pancreatic insufficiency. Domschke detected the change of plasma AA level before and after intravenous infusion of cholecystokinin and secretin to diagnose exocrine pancreatic insufficiency, and the accuracy being 91.0%, which was confirmed further by Gullo who used cerulein instead of cholecystikinin and secretin stimulating pancreatic secretion.

The diagnostic value of AACT in exocrine pancreatic insufficiency in Chinese has not been well understood because of the difference in race, living custom, geography, and the high fat and protein food and higher incidence of chronic pancreatitis in Europe and America, but more carbohydrate food and lower incidence of chronic pancreatitis in this country. Our study shows that the diagnositic accuracy of AACT for exocrine pancreatic insufficiency in Chinese was similar to that in European and American people. But attention should be paid to the following points: ① The relative decline rate over 14.0% of plasma AA was found only in 13 and 45 cases of Groups 1 and 2 during intravenous infusion of cerulein at 45 min, and in 18 and 68 cases at 60 min, the maximum decrease of plasma AA occurred later than that of Gullo′s report. Therefore, the results should be analyzed in combination at 45 min, 60 min and 90 min. ② Venous blood should be collected gently to avoid hemolysis. Incorrect deproteinization or not done in time will lead to bias in results.

To further reduce the test cost and illumed by the idea of Lundh test, we used oral reagent free from amino acid instead of intravenous infusion of cerulein in AACT. Because the volunteers would not like intubation in duodenum, gallbladder contraction function was observed after drinking of reagent A to view indirectly the effect of the oral reagent stimulating pancreatic secretion, based on the theory that hydrochloric acid and fat stimulate bile and pancreatic fluid secretion, and gallbladder contraction.

In this study, we found that O-AACT, based on the same design theory of I-AACT, is simple, inexpensive and easy to accept and plays a good screening role in evaluating severe exocrine pancreatic insufficiency. But whether it is applicable to diagnosis for slight and moderate exocrine pancreatic insufficiency should be further studied.