世界規模でのグリーンパパイヤの研究、土壌細菌叢の研究

TRIALPROTOCOL (Confidential)

Title:   

   INFLUENCE OF BIO –NORMALIZER ON TOXICITY INDUCED BY

THERAPEUTIC IRRADIATION

 

 

Phase III

Professor A G. Rumyanzev
Director of Russian Institute for
Pediatric Hematology

Professor L.G Korkina
Research Supervisor

Professor I.B.Afanas’ev
Organizer Monitor

 

Research Investigators:

Dr. T.B. Suslova, Ph.D.
Dr. Z.P. Cherernisina, Ph. Dr. I.B. Deeva, Ph.D.
Dr. I. Yu. Begma

Clinical Investigators: Dr. E.V. Samochatova, D.Sci., Ph.D.
Dr. AA. Maschan., Ph.D.

 

Table of Contents

 

1.0         Checklist for Patient Eligibility and Necessary Information

2.0         Objectives and Rationale

3.0         Background

4.0         Patient Eligibility

5.0         Treatment Plan

6.0         Patients Evaluation

 

Appendices

 

A1.0      Case Form

A2.0      Consent Form

A3.0      Decision of the Ethics Committee

 

 

1.0 Checklist for Patient Eligibility and Necessary Information

 

 

Age between 2-15 years.

Cytological  evidence  of  acute  lymphoblastic  leukemia  with  clinical neuroleukemia  or  in relapse.

Cytological evidence of acute myeloblastic leukemia.

Evidence of measurable disease.

Recovered from toxic effects after previous chemotherapy.

Cranial irradiation.

HIV infection

Bone marrow total cytosis > 50000, blast cells < 5%.

Peripheral absolute neutrophil count > 500/mm³

Platelet count > 100000 mm³

Bilirubin < 2.0 mg/dL, SGPT < 500 U/L

Normal coagulation screen

Creatinine < 2.0 mg/dL.

Normal urinanalysis.

Informed consent explained to and signed by patient/guardian.

 

 

 

 

 

 

 

 

2.0ObjectivesandRationale

 

2.1 To  determine the toxicity of  cranial irradiation following the  polychemotherapy  of pediatric  patients  with acute lymphoblastic  leukemia  with  clinical neuroleukemia or  in relapse and with acute myeloleukemia.

2.2 To determine the influence of Bio-normalizer (BN) on adverse effects of therapeutic irradiation.

2.3 To determine the effect of BN on the biochemical and cytological parameters of spinal fluid and on the free radical status of a child’s organism.

 

3.0 Background

The intensive worldwide researches of free radicals of the past 15 years have shown that oxygen free radicals play an important role in development of various pathologic processes such as inflammation, tumors, allergy, degenerative disorders, age- and environment­ induced damages. It is now well known that unusually large amounts of free radicals can be formed in pediatric patients with many blood-related diseases (leukemias, aplastic anemias, hemolytic anemias, thalassemia, etc.). Moreover, there is a large body of evidence that the action of many antitumor drugs, for example anthracycline antibiotics and bleomycin, as well as therapeutic irradiation is mediated by oxygen radicals. At the same time, an excess of these toxic species in an organism may he an origin of serious deleterious effects in normal cells and tissues. Therefore, it is believed that oxygen radicals are responsible for the adverse side effects of these drugs and of other therapeutic treatments.

For  example,  in  order  to  prevent  neurolekemia development,  therapeutic  cranial irradiation  was  applied  to  the   patients  with  different   hemoblastosis  after   previous polychemotherapy.  (Neuroleukemia  is  known  to  be  one  of  the  most  dangerous  life­ threatened features of leukemias). However, the irradiation of the child head resulted in mild or strong encephalopathy developing 4-8 weeks after irradiation. Such a postponed toxicity may be due to free radical damage of endothelium of brain vessels and developing edema or possibly myelin degradation.

Under physiological conditions, the level of organism’s oxygen radicals is controlled by powerful antioxidant systems including enzymes (SOD, catalase, glutathione peroxidase, etc.) and low-molecular antioxidants such as glutathione, ascorbic acid, and α-tocopherol. If endogenous antioxidant systems deteriorate as a result of permanent oxidative stress, the intervention of highly toxic agents, or the malfunctions of cellular organelles, a dangerous elevated level of oxygen radicals may be rapidly achieved.

We supposed that BN administration can significantly improve the free radical status of patients by suppressing damaging post-effects of irradiation and diminishing the risk of the development of encephalopathy. Indeed,  it has been  shown  that BN is a hydroxyl  radical scavenger  (Santiago,  et al., 1991)  and powerful  inhibitor  of the release of highly  reactive oxygen species  by phagocytes (Osato, et al., 1994). In addition, it has been recently found (Korkina,  et al., 1994) that BN is able to enhance  the level  of reduced  glutathione  (one  of the most important  endogenous antioxidants)  and to stimulate  the SOD  activity. On these grounds,  in  this pilot  study   we  were   testing  the  efficiency   of  BN   in  prevention   of irradiation-induced brain pathology  and  were studying the  possibility of  normalizing  free radical processes in blood and spinal fluid of patients during and after cranial irradiation.

 

4.0 Patient Eligibility

4.1  All  the  patients  from   2  to  15   years   of  age  previously   treated   with  multidrug chemotherapy, who at the risk of neuroleukemia  development and need therapeutic  cranial irradiation in dose more than 18 Gy.

4.2  Cytological evidence of diagnosis of lymphoblastic leukemia with initial neuroleukemia or in relapse; acute myelolcukemia.

4.3  Patients must have recovered from the toxic effects of all prior chemotherapy.

4.4  Patients  must  have  adequate  bone  marrow  function  (defined   as  a  total  cytosis  > 50000/ml and the blast cells content < 5%).

4.5  Patients  must  have  normal  parameters  of  peripheral  blood  (defined  as a  peripheral absolute  neutrophil  count  (ANC)  of >500/mm³, platelet count  of  > 100000/mm³, and hemoglobin content  > 10 g/dL).

4.6  Patients must have _______________________                             

 

5.0 Treatment plan

Patients  will be given  1 sachet  of  BN  3  times  a  day  during  one  month  just  after  the completion of irradiation treatment.

6.0 Patient Evaluation

7.1    A complete history and physical examination   with documentation of measurable disease is necessary for admission to the study.

7.2 other studies. Sequences.

                                                                                                                                                           

 

Physical examination

Total blood count including RBC, WBC, platelets, blast, Hb 

Biochemical blood analysis (bilirubin, urea, total protein, ALT, AST, electrolytes, iron)
Urinalysis Spinal fluid analysis (cytosis, total protein, cytopreparate) 

The determination of free radical status in blood (the intensity of leukocyte chemiluminescence, the content of glutathione, the activity of SOD, etc.)

EthicsCommission

 

Pursuant to the Declaration of Helsinki, the present study program was submitted to an independent Ethics Commission of the Russian Institute of Pediatric Hematology, Moscow, for review. The decision of this Ethics Commission must be presented in writing to the study director and organizing monitor. Any recommendations and requirements made by the Commission must be incorporated into the study schedule.

The start of the study is dependent on a positive decision by the Ethics Commission.

 

PatientInformation andConsent

 

The   clinical investigator must instruct the  patients about  the  nature, importance and scope of the trial He (she) must  obtain  and countersign the written  declaration of informed consent of each patient before his/her participation in the study. A copy of the declaration of informed consent must be handed out to every study participant.

 

 

Signatures

Professor A.G. Rumyanzev
Director of Russian Institute for Pediatric Hematology

 

Professor L.G. Korkina
Research Supervisor

 

Professor I.B. Afanas’ev
Organization Monito

 

 

CLINICAL TRIAL

INFLUENCE OF BIO-NORMALIZER ON NEUROTOXICITY INDUCED BY THERAPEUTIC IRRADIATION

 

(Pilot study)

 

FINAL REPORT

 

 

 

1. CHILDREN

 

Children suffering from acute lympholeukemia (11 children) and acute myeloleukemia (19 children). All of them have been examining and treated at the Russian Institute of Pediatric Hematology. General characteristics of the children studied are presented in Tables 1-6. Children taking part in the clinical trial were divided into 6 randomized groups:

Group A:  4 children suffering from acute lympholeukemia were given standard chemotherapy for 1 month and then were subjected to cranial irradiation in dose 12 Gy (Control 1).

Group B: 7 children were subjected to the same treatment as the children of Group A, but in addition they were given 1-3 sachets of BN a day for two weeks before irradiation.

Group C:  3 children suffering from acute myeloleukemia were given standard chemotherapy plus cranial irradiation in dose 18 Gy (Control2).

Group D: 3 children were treated as those of Group C, but in addition they were given 1-3 sachets of BN a day for 1 month after irradiation treatment.

Group E: 7 children suffered from acute myeloleukemia were under the intensive course of chemotherapy (Control 3).

Group F: 6 children were treated as those of Group E, but in addition they were given 1-3 sachets of BN a day for 1 month after the chemotherapy completing.

 

During the clinical trial traditional hematological  and biochemical analyses essential for the  evaluation  of  efficacy  of  the  chosen  treatment  and  the  main  living functions  were performed  several   times.   They   were   a full blood   count   (the   hemoglobin   content, differential cell count, hematocrit, cell size distribution), a special histological analysis of the blood and bone marrow smears, a total protein billirubin and creatinine contents and ALT and AST activities. For measurement of the organism’s oxidative stress, intensities of spontaneous and latex-activated luminol-dependent chemiluminescence in isolated leukocytes. CuZnSOD activities in erythrocytes, leukocytes, plasma and spinal fluid and MnSOD activity in leukocytes were determined. The corresponding analytical techniques are given below. There were no drop-outs in the clinical trial due to the adverse effects of drug administration.

                                                             

 

 

 

2.EXPERIMENTAL METHODS

2.1BloodSamples

 

Venous blood was drawn by disposed syringes early in the morning before breakfast   into three tubes: one of them was with 20 U/ml heparin, second was with sodium citrate and the third was without any anticoagulant. 1.5 ml blood was collected into the first tube, 0.5 ml was collected into the second tube, and 9 ml was collected into the third one. Serum was obtained from the 9 ml blood sample by centrifugation and then used for biochemical assay. The 0.5 m1 blood sample was used for differential cell count, and the 1.5 ml blood sample was used in CL, SODs, and glutathione assays.

 

 2.2PreparationofLeukocytes

 

The blood samples (1.5 ml) were anticoagulated with 0.2 m1 of heparin (20 U) in Hanks’ balanced salt solution (HBSS) and sedimented with 1.5 m1 of dextran-metrizoate mixture (50  m1  6.2%  dextrane/20  m1  38%  metrizoate)  at  25°C for  30  min.  The cell-rich supernatant was centrifuged at 150xg for 10 min. Cell pellets were washed twice in HBSS. The final suspension of 2-3×10⁶ leukocytes was prepared in the cultural medium 199. The cells were counted with a microscope, and their viability was assessed by exclusion of 0.1% trypan blue dye. Cell differential count was confirmed in the cell smears using Giemsa staining.

 

2.3ChemiluminescenceAnalysis

 

Luminol-dependent chemiluminescence was measured on a LKB luminometer mod. 1251 (Sweden).   Leukocyte suspension   (20 μl 2×10⁶ cells/ml), luminol   (50 μl final concentration), and 0.85 m1 of HBSS were mixed in the 1 m1 cuvette at 37°C.  After 5 min, the 0.1% suspension of latex (Sigma) in 0.95% NaCl solution was added, and the light emission was recorded continuously.  The intensity of spontaneous CL and the difference between the maximal values of the cellular CL response to latex and of spontaneous CL were measured.

 

2.4 Measurement of Glutathione Metabolism in Red Blood Cells

 

Erythrocytes were obtained after blood sedimentation on dextran-metrizoate gradient and washed twice in the cold phosphate buffer, pH 7.2. The content of reduced glutathione was determined by the Beutler method⁶.

 

2.5 Measurement of SODs and catalase activities

 

Superoxide   dismutase     (CuZnSOD)      activity     was determined by the adrenaline method⁷, in which the rate of superoxide production   was    measured    by lucigenin­ amplified CL⁸. Heparinized venous blood (0.2 ml) was hemolyzed with ice-cold water. Lysate was added to the equal volume of ethanol-chloroform mixture (1:1 v/v) and centrifuged at 1500 x g for 30 min. Protein content in supernatant was determined by the Lowry method⁹. To measure the total SOD activity, 50 mL top clear  supernatant was added to  carbonate  buffer  (pH  10.2, a total volume of 900 mL) containing EDTA (100 mM) and lucigenin (100 mM) in the polysterene CL  cuvette ,and the  level of  basal CL was registered continuously. Reaction was started by adding 50 mL adrenaline (100 mM) through the dispenser. CL light sum for 5 min was recorded and compared with that of a control sample (50 mL water: ethanol: chloroform solution (2:1:1 v/v/v)).  The total SOD activity was calculated from calibration curve using commercial SOD as a standard and expressed as U/mg protein. For the determination of: MnSOD activity, CuZnSOD was inhibited by the addition of NaCN (4 mM) to the supernatant. After that, the CL measurement of: MnSOD activity was performed as described above. CuZnSOD activity was estimated as a difference between the total and MnSOD activities.

Catalase activity was determined in the same lysates according to Aebi¹⁰. Enzyme activity was expressed as units per mg protein.

 

2.6 Statistical Analysis

The clinical trial was a randomized open study on 30 children with daily oral doses of Bio­ normalizer. Student’s t-test for mean values was used for evaluation of significance between different experimental groups. Paired significance test was used for connected values obtained before and after clinical trial. 5% significance level was assumed. Results were presented as mean ± SEM.

 

3. RESULTS AND DISCUSSION

The examination of children by pediatricians during and after the completion of the clinical trial has shown that there were no toxic side effects, allergic reactions and other adverse events during the BN administration. Biochemical analyses confirmed these observations. Pediatricians concluded that there was a good tolerability to the BN application. More than that, in accord with previous findings, there was significant beneficial effect of BN administration on the liver functions (Table 7). As is seen from this Table, even short-term BN administration statistically significantly improved liver enzyme activities (ALT and AST) that reflected the activation of the reparative processes in liver which certainly was damaged during chemotherapy.

3 of 4 children (75%) of Group A exhibited severe neurotoxic symptoms after irradiation exposure. One of them died of the relapse of ALL (Table 8). At the same time, only 2 of 7 children (28%) of Group B treated with BN before irradiation cycle exhibited moderate encephalopathy, all of them achieved a remission and until now they are alive (Table 8).

The more strong beneficial effect of BN application was found in a comparative study of the children from Groups C and D (Table 9). All children of Group C (100%) exhibited a moderate  encephalopathy,  and  no  one  of  Group  D  (0%)  treated  with  BN  after  the irradiation had any neurotoxic symptoms.

Comparison of children from Groups E and F shows that in the control Group two of 7 children (28%) died, being non-responsive to the applied chemotherapy. In contrast, children of Group F who were given BN after chemotherapy, exhibited no relapses (0%), and all of them are alive (Table 10).  However, BN did not affect the agranulocytosis duration. An average duration of the drug-induced a plastic syndrome was 17.6 days in the control Group and 33 days in the experimental Group E (Difference was statistically insignificant). These results could be explained in terms of cytokine production activation during BN administration. It has been previously shown in the in vitro experiments that BN can stimulate γ-interferon and α-TNF production by monocytes and macrophages. These particular cytokines are capable of suppressing the bone marrow cell proliferation. That may cause a slight prolongation of the symptoms of the drug-induced aplasia.

One of the causative reasons of the toxic effects induced by chemotherapy and therapeutic irradiation may be the overproduction of oxygen free radicals in an organism. It is now well established that these very reactive species are able to damage DNA, proteins, lipids, polysaccharides, and various biological structures including neurons and myelin. Due to that, the disturbances in the hematopoietic tissues, the immune system, and the central and peripheral nervous systems occur. There are at least four major pathways of suppressing free radical processes and reducing the state of oxidative stress:

a) Scavenging of free radicals by antioxidants and oxygen radical scavengers;

b) The stimulation of the activities of the endogenous antioxidant systems;

c) Suppressing the activities of the enzymes catalyzing the oxygen radical generation;

d) Chelating and inactivating active ions of transition metals, which catalyze the production of the most aggressive hydroxyl and hydroxyl-like radicals.

There is a body of evidence that Bio-normalizer can influence free radical processes in organism due to its significant hydroxyl radical scavenging activity [    ]. Furthermore, it was shown by us in the previous study that BN enhanced CuZnSOD activity in the macrophages [   ] and to some extent inhibited the NADPH-oxidase of neutrophils and macrophages. It could     be suggested also that BN possessed a weak chelating activity because it contains a large amount of polysaccharides. Therefore, BN can interfere with endogenous free radical processes by different metabolic pathways.

Until  now,  there  were  no  data  whether  BN   can  affect  glutathione  metabolism. Glutathione is one of the major blood antioxidant responsible for the inactivation of various oxidants formed during drug and other xenobiotics metabolism or as results of the environmental hazard. In present work it was found that irradiation and chemotherapy substantially decreased the level of reduced glutathione in the erythrocytes, while BN given before or after the irradiation treatment increased its level up to normal values (Table 11) at the same time, BN did not affect the level of glutathione when it was given after chemotherapy (Group F). These data allowed to suggest that a very strong protocol treatment of lympho- and myeloprolipherative malignancies caused a suppression of glutathione system, that can be an origin of the oxidative stress- induced deleterious effects of therapeutic irradiation and chemotherapy. BN can prevent such deleterious effects of irradiation but not chemotherapy by inducing glutathione synthesis and /or reducing oxidizing glutathione. What is a reason for the diverse effects of BN on irradiation and drug-induced glutathione depletion? That may be discovered only in course of further studies.

      It was found that BN administration before and after irradiation induced the CuZnSOD activity in children’ leukocytes and the catalase activity in erythrocytes (Tables 12). There were no significant changes in erythrocyte CuZnSOD activity (Table 13). Surprisingly, leukocyte MnSOD activity was suppressed by BN administration at its high level and was stimulated by BN at low level (Table 14). These results may be explained in terms of the capacity of BN to increase the organism potency to be adapted to oxidative stress. There is growing body of evidence that one of the genetically regulated properties of mammalian cells is the ability to be adapted to oxidative stress. Some proteins exhibit increased expressions during such an adaptation among of them are antioxidant enzymes, DNA repair enzymes, DNA damage-specific gene products, heme oxygenase, heme binding proteins, etc.^11,12. Oxidative stress can affect enzymes involved in intracellular signal transduction that in turn lead to blocking cell proliferation and activating apoptosis in bone marrow stem cells¹³. Moreover, it is now well established that strong prooxidants such as hydroxyl radicals and peroxynitrite are involved in carcinogenesis inducing protooncogene expression influencing tumor and promotion^{14, 15}. On the basis of the recognized role for the reactive oxygen species in tumor initiation and promotion, it has been hypothesized that antioxidant enzymes genes possess anti-oncogenic properties¹⁶. A decreased of the activity of antioxidant enzymes may enhance the oxidative stress, which allows the tumors to arise and develop. Therefore, it is important that BN administration allow to maintain the normal activities of main antioxidant enzymes such as CuZnSOD, MnSOD and catalase. It seems that the target for the BN stimulating activity is predominantly blood circulating monocytes and tissue macrophages.

         It should be stress that reactive oxygen species do not exhibit deleterious effects but also possess very important physiological properties. Thus, it has been shown many times that superoxide and hydrogen peroxide can defend an organism against bacterial infections, play a role as adaptongenes inducing endogenous free radical defense system, and stimulate growth, maturation, and differentiation in a variety of a mammalian cells¹⁷.While hydroxyl radicals are known as the most aggressive agents in capable initiating the irreversible damage, superoxide and hydrogen peroxide can interfere with physiological processes and regulate them. Therefore, deficiency of these physiologically important reactive oxygen products can suppress cell growth and maturation^{17, 18}. As is seen from Table 15, BN administration markedly increased spontaneous and latex-activated luminol –dependent CL in the blood leukocytes of the most of the patients. This effect may be easily interpreted taking into account that due to agranulocytosis (a granulocyte deficiency) after the intensive chemotherapy, the main source of the essential oxygen free radicals became blood monocytes. BN is known as potent activator of monocyte-macrophage system, and therefore it can stimulate the blood monocytes to produce more radicals in order to compensate the granulocyte deficiency. 

 

Table1.Generalcharacteristicsofpatients

 

 

 GroupA(control 1)

 

Number	Name	Diagnosis, group of risk	Sex	Age, years
1	Ganin	ALL, mild	M	6
2	Kopylov	ALL, moderate	M	2
3	Sitnikov	ALL, mild	M	4
4	Dovlatyan	ALL Moderate	F	2

 

Group B: BN for 2 weeks before the 12Gy irradiation

Number	Name	Diagnosis group of risk	Sex	Age
1	Kchabalaeva	ALL, mild	F	5
2	Kulchenko	ALL, moderate	M	5
3	Malofeeva	ALL, moderate	F	16
4	Bandura	ALL, mild	M	4
5	Vasinov	ALL moderate	M	4
6	Frolov	ALL, mild	M	4
7	Laptev	ALL, moderate	M	8

 GroupC(control2)

 

Number	Name	Diagnosis, group of risk	Sex	Age, years
1	Sedov	AML, M5, II	M	11
2	Knyazeva	AML,M2, I	F	15
3	Gerasimova	AML, M2, I	F	3

 

GroupD:BNfor1monthafterthe18Gyirradiation

Number	Name	Diagnosis, group of risk	Sex	Age, years
1	Bubnov	AML,M2, I	M	13
2	Mormel	AML, M5,II	F	5
3	Sushkov	AML,M4,II	M	8

 

 

 

Group E (control3)

 

Number	Name	Diagnosis, group of risk	Sex	Age
1	Smirnov	AML, M2,I	M	12
2	Mishina	AML, M5, II	F	6
3	Kotova	AML, Ml, I	F	15
4	Sidenko	AML, M5, II	M	11
5	Bazylina	AML, M4, II	F	10
6	Sautina	AML, M4, II	F	10
7	Gerasimova	AML, M2, I	F	3

 

GroupF:BNafter theintensive chemotherapy

 

Number	Name	Diagnosis, group of risk	Sex	Age, years
1	Shurupov	AML, M5a, II	M	8
2	Zmyrko	AML, relapse	F	16
3	Ivanova	AML, relapse	F	13
4	Sazonova	AML, M1, II	F	10
5	Verlina	AML, M5a, II	F	6
6	Petrov	AML, M5a, II	M	6

 

Table 7.
ALT and AST activities in the blood plasma before and after the BN administration. Group B

Number	ALT		AST
	before	after	before	after
1	261	22.5	176	23.5
2	357	41.4	41.1	46.2
3	150	53.5	64.4	24.8
4	200	42.7	93.1	31.6
5	106	76.8	58.2	31.0
6	29.8	31.8	34.8	41.1
7	200	142	99.3	62.8
M	104.4	58.7	81	37.3

GroupD

 

 

Table 8. The clinical effect of BN administration in patients of Group B
Comparing with control l

 Group A (control 1)

 

Number	Adverse effects	Encephalopathy	Remission
1	No	0	+
2	Yes	2.0	+
3	Yes	1.5	+
4	Yes	2.5	-,ex. let

Averagescore 1.5 

 

Group B

 

Number	Adverse effects	Encephalopathy	Remission
1	no	0	+
2	fever	1.0	+
3	no	1.5	+
4	fever	0	+
5	no	0	+
6	no	0	+
7	no	0	+

Averagescore 0.35 

Table9.TheclinicaleffectofBNadministrationinpatientsofGroup D

comparingwithcontrol2

 

Group C

 

Number	Adverseeffects	Encephalopathy	Remission
1	no	1.5	+
2	yes	2.0	+
3	yes	2.0	+

 

Averagescore1.9

 

Group D

 

Number	Adverseeffects	Encephalopathy	Remission
1	no	0	+
2	no	0	+
3	no	0	+

 

Averagescore 0

Table10.TheclinicaleffectofBNadministrationinpatientsofGroupF comparingwithcontrol 3

 

Group E

 

Number	Adverse effects	A plastic syndrome, days		Remission
1	no	16		+
2	no	20		+
3	no	17		+
4	no	19		+
5	no	14		+
6	no	15		-ex. let
7	no	22		-ex. let.

 

Averagevalue17.6

 

 

Group F

Number	Adverse effects	A plastic syndrome, days	Remission
1	no	15	+
2	no	35	+
3	no	71	+
4	no	16	+
5	no	29	+
6	no	32	+

Averagevalue 33.0

 

Table11.Reducedglutathionecontent(GSH)intheerythrocytesbeforeandaftertheBN administrationandbeforeandafterirradiation.

 

Group B

 

                                                                

 

Group D

 

Number	Irradiation		BN
	before	after	before	after
1	7.8	8.8	8.8	9.0
2	6.9	4.9	4.9	6.8
3	7.1	6.3	6.3	7.8
_M	7.3	6.7	6.7	7.9

Group F 

Number	Chemotherapy		BN
	before	after	before	after
1	6.9	7.0	7.0	7.9
2	7.5	7.1	7.1	7.9
3	7.0	7.5	7.5	7.3
4	7.9	6.9	6.9	6.7
5	11.8	11.9	11.9	10.4
6	6.8	7.3	7.3	11.9
M	7.95	7.95	7.95	8.7

Table 12. SODs and catalase activities before and after BN administration.

GROUP B

	Leukocyte	CuZnSOD	Leukocyte	MnSOD	Erythrocyte	catalase
Number	before	after	before	after	before	after
1	333	810	116	455	17	37
2	304	380	246	76	24	42
3	176	250	250	66	29	32
4	132	112	44	62	32	48
5	141	205	41	122	26	39
6	01	42	40	14	37	38
7	234	382	525	26	25	30
M	197.3	311.6	181.2	117.3	27.2	38

 

GROUP D

	Leukocyte	CuZnSOD	Leukocyte	MnSOD	Erythrocyte	catalase
Number	before	after	before	after	before	after
1	326	442	276	509	n.a.	n.a.
2	245	455	177	531	n.a.	n.a.
3	140	304	70	61	26	38
M	237	400.3	174.3	367

 

GROUP F

	Leukocyte	CuZnSOD	eukocyte	MnSOD	Erythrocyte	catalase
Number	before	after	before	after	before	after
1	31	304	28	26	17	35
2	59	317	68	26	24	32
3	88	414	49	242	n.a.	n.a.
4	104	152	15	66	n.a.	n.a.
5	129	52	129	37	21	46
6	80	80	80	50	14	27
M	81.8	219.8	61.5	74.5	19	35

 

 

Table 13. Luminol-depemdent CL intensities before and after BN       

administration
		GROUP B
Number		Spontaneous        before                  after		Latex-activated        before                         after
1		1222                                 154		11156                                    285
2		386                                   203		8237                                    8707
3		2279                                 220		13308                                 12737
4		792                                 2168		5472                                   13439
5		630                                   741		3167                                     6935
6		360                                 2012		3257                                    1821
7		929                                  118		5714                                      634

 

 

Group D

 

Number	Spontaneous              before             after	Latex-activated before        after
1	138                      452	4070               7928
2	296                      321	10112             1175
3	1016                    1285	14371             55869

 

 

 

 

 

 

 

GROUP F

 

Number	Spontaneous    before                           after	Latex-activated before                       after
1	72                              120	1496                        2700
2	482                             6	1841                            10
3	71                              433	1434                         4425
4	810                             9	7840                          24
5	400                            1105	3640                         7995
6	4860                          762	51300                       16269